Microglia and tumor (excluding TNF)
Aoki K, Uchihara T, Sanjo N, Nakamura A, Ikeda K, Tsuchiya K,
Wakayama Y (2003) Increased expression of neuronal apolipoprotein E
in human brain with cerebral infarction. Stroke 34:875-880
Abstract:
BACKGROUND AND PURPOSE: Cellular origin of apolipoprotein E (ApoE)
in the human brain and its roles in physiological and pathological
conditions remain to be clarified. METHODS: Immunolocalization of
ApoE was investigated in a series of autopsied human brains with or
without infarction. ApoE expression was also estimated on immunoblot
on protein extracts from autopsied brains and a cultured
neuroblastoma cell line of human origin (GOTO) subjected to an
oxidative stress induced by exposure to hydrogen peroxide (0.2
mmol/L). RESULTS: In addition to astrocytes and Microglia,
neurons and degenerated axons in and around the ischemic foci
contained ApoE-like immunoreactivity, which was more intense in
recent ischemic foci. Immunoblot demonstrated an increase in
expression of ApoE in brain extracts from ischemic lesion, and this
increase was also pronounced in the cultured neuroblastoma cell line
after the stress. CONCLUSIONS: Accumulation of ApoE in neurons in
and around ischemic foci of the human brain is related to an
increase in ApoE synthesis in neurons, as seen in cultured neuronal
cells after oxidative stress. Intrinsic regenerative activity of
neuron in reaction to external insults may be related to this
increase in ApoE of neuronal origin
Castilla EA, Prayson RA, Kanner AA, Rybicki LA, Tubbs RR,
Vogelbaum MA, Barnett GH (2003) Cyclooxygenase-2 in oligodendroglial
neoplasms. Cancer 98:1465-1472
Abstract: BACKGROUND: Although
increased expression of cyclooxygenase-2 (COX-2) has been described
in association with a variety of neoplasms, including tumors of
astrocytic derivation, limited data are available on COX-2
expression in oligodendrogliomas. METHODS: The current study
retrospectively reviewed 53 oligodendrogliomas and 7
oligodendroglioma-predominant oligoastrocytomas (mixed gliomas) for
COX-2 expression and MIB-1 proliferative index (by
immunohistochemistry) and for chromosome 1p status (by fluorescence
in situ hybridization). RESULTS: Patients included 35 males and 25
females, with a mean age of 41 years (range, 12-73 years) at the
time of surgery. Forty-four tumor
specimens were classified as World Health Organization (WHO)
Grade II neoplasms and 16 as WHO Grade III tumors. MIB-1 labeling
indices (marker of cell proliferation) ranged from 0 to 22.3 (mean
4.5). Twenty-eight tumor specimens
demonstrated allelic loss on chromosome 1p. Positive staining was
observed in 17 tumor specimens
with COX-2 antibody. COX-2-positive tumor
specimens were also evaluated with CD68
(macrophage/Microglial
cell marker) by coimmunolabeling to confirm that the observed COX-2
immunostaining was not due to immunoreactive macrophages or
Microglial
cells. COX-2 expression, lack of allelic loss at chromosome 1p, and
high proliferation indices were associated with decreased survival
(P = 0.002, P = 0.009, and P = 0.015, respectively). No correlation
with outcome was found with patient gender, age at diagnosis, or
histologic grade. CONCLUSIONS: Chromosome 1p, COX-2
immunoreactivity, and MIB-1 labeling indices correlated with outcome
and were associated with decreased survival. There was not a
one-to-one correspondence between COX-2 immunoreactivity and lack of
allelic loss at chromosome 1p. Tumors with expression of COX-2 by
immunohistochemistry may, in theory, benefit from treatment with
therapeutic agents that inhibit COX-2
Das P, Estephan R, Banerjee P (2003) Apoptosis is associated
with an inhibition of aminophospholipid translocase (APTL) in
CNS-derived HN2-5 and HOG cells and phosphatidylserine is a
recognition molecule in Microglial
uptake of the apoptotic HN2-5 cells. Life Sci.
72:2617-2627
Abstract: A balance of the activities of multiple
enzymes maintains the typical asymmetry of plasma membrane lipids in
healthy cells. Such enzyme activities are (a) the aminophopholipid
translocase (APTL) (a lipid-selective P-type ATPase that catalyzes
inward movement of aminophospholipids), (b) the scramblase (a
calcium-dependent and ATP-independent enzyme that catalyzes both
inward and outward movement of lipids), (c) the floppase (an
ATP-dependent enzyme that catalyzes only outward movement of
lipids). Activation or inhibition of any one of these enzymes would
lead to a loss in this asymmetry. Apoptosis-associated
externalization of phophatidylserine has been reported for many
different cell-types, but the exact mechanism involved in this loss
of membrane asymmetry has not been identified yet. In this report we
demonstrate concurrence of APTL inhibition, caspase-3 activation and
apoptosis in CNS-derived HN2-5 and HOG cells. Additionally, we
provide data to demonstrate that the phagocytosis of apoptotic,
CNS-derived HN2-5 cells by the Microglial
cells requires recognition through phosphatidylserine (PS). Thus the
enzyme aminopholipid translocase is inhibited during apoptosis of
CNS-derived cells and this alone could account for the loss of
plasma membrane lipid-asymmetry observed in these cells
Deininger MH, Weinschenk T, Meyermann R, Schluesener HJ
(2003) The allograft inflammatory factor-1 in Creutzfeldt-Jakob
disease brains. Neuropathol.Appl.Neurobiol. 29:389-399
Abstract:
The allograft inflammatory factor-1 (AIF-1) is a 17-kDa IFN-gamma
inducible Ca(2+)-binding EF-hand protein that is encoded within the
HLA class III genomic region and is involved in immune dysfunction
and smooth muscle cell activation. We used immunohistochemistry
double labelling experiments to analyse the spatial distribution and
cell-type-specific localization of AIF-1 in the brains of patients
who died as a result of sporadic Creutzfeldt-Jakob disease (CJD) and
neuropathologically unaltered controls. Significantly more AIF-1
immunoreactive macrophages/Microglial
cells and, interestingly, neurones were observed in CJD patients
compared to controls. Western blotting confirmed more prominent
AIF-1 immunoreactive bands of approximately 50 kDa in four CJD
patients compared to three controls. Chaotropic SDS-PAGE of the
recombinant AIF-1 resulted in almost complete reduction of the 50
kDa band and mass spectrometry revealed only AIF-1-specific tryptic
protein fragments suggesting that trimerized AIF-1 is the
predominant form in vivo. Finally, we analysed mechanisms of
neuronal AIF-1 induction. Following H2O2 challenge, a model of
general cell stress, we observed the gradual induction of AIF-1 and,
more interestingly, release to the supernatant of SKNSH neurones.
Parallel reverse transcriptase polymerase chain reaction and
sequencing was used to confirm AIF-1 mRNA expression
Fiano V, Ghimenti C, Schiffer D (2003) Expression of cyclins,
cyclin-dependent kinases and cyclin-dependent kinase inhibitors in
oligodendrogliomas in humans. Neurosci.Lett. 347:111-115
Abstract:
Cyclins are regulatory proteins of the cell cycle which bind and
activate kinases. In gliomas, contrary to many malignancies, cyclin
D1 is rarely amplified, but together with other cyclins, it
increases with anaplasia. In a series of 23 surgical biopsies of
grade II and III oligodendroglioma, cyclin D1, E, A, B1, CDK4-6,
CDK2, Cdc2 and p27/Kip.1 have been studied by immunohistochemistry
and Western blot. Cyclin D1 and A increased with anaplasia, showing
a linear correlation with MIB.1 labeling index and an inverse
correlation with p27/Kip.1 expression. Cyclin E and B1 and kinases
were almost only expressed in grade III tumors. Normal
oligodendrocytes and Microglia
cells of the cortex and white matter showed a clear positivity for
cyclin D1, but not for other cyclins or kinases
Flannery T, Gibson D, Mirakhur M, McQuaid S, Greenan C,
Trimble A, Walker B, McCormick D, Johnston PG (2003) The clinical
significance of cathepsin S expression in human astrocytomas.
Am.J.Pathol. 163:175-182
Abstract: Early local invasion by
astrocytoma cells results in tumor
recurrence even after apparent total surgical resection,
leading to the poor prognosis associated with malignant
astrocytomas. Proteolytic enzymes have been implicated in
facilitating tumor cell
invasion and the current study was designed to characterize the
expression of the cysteine proteinase cathepsin S (CatS) in
astrocytomas and examine its potential role in invasion.
Immunohistochemical analysis of biopsies demonstrated that CatS was
expressed in astrocytoma cells but absent from normal astrocytes,
oligodendrocytes, neurones and endothelial cells. Microglial
cells and macrophages were also positive. Assays of specific
activity in 59 astrocytoma biopsies confirmed CatS expression and in
addition demonstrated that the highest levels of activity were
expressed in grade IV tumors. CatS activity was also present in
astrocytoma cells in vitro and the extracellular levels of activity
were highest in cultures derived from grade IV tumors. In vitro
invasion assays were carried out using the U251MG cell line and the
invasion rate was reduced by up to 61% in the presence of the
selective CatS inhibitor 4-Morpholineurea-Leu-HomoPhe-vinylsulphone.
We conclude that CatS expression is up-regulated in astrocytoma
cells and provide evidence for a potential role for CatS in invasion
Giri RK, Selvaraj SK, Kalra VK (2003) Amyloid peptide-induced
cytokine and chemokine expression in THP-1 monocytes is blocked by
small inhibitory RNA duplexes for early growth response-1 messenger
RNA. J.Immunol. 170:5281-5294
Abstract: In Alzheimer's disease
(AD) one finds increased deposition of A beta and also an increased
presence of monocytes/macrophages in the vessel wall and activated
Microglial
cells in the brain. AD patients show increased levels of
proinflammatory cytokines by activated Microglia.
Here we used a human monocytic THP-1 cell line as a model for
Microglia
to delineate the cellular signaling mechanism involved in amyloid
peptides (A beta(1-40) and A beta(1-42))-induced expression of
inflammatory cytokines and chemokines. We observed that A beta
peptides at physiological concentrations (125 nM) increased mRNA
expression of cytokines (TNF-alpha, and IL-1 beta) and chemokines
(monocyte chemoattractant protein-1 (MCP-1), IL-8, and macrophage
inflammatory protein-1 beta (MIP-1 beta)). The cellular signaling
involved activation of c-Raf, extracellular signal-regulated
kinase-1 (ERK-1)/ERK-2, and c-Jun N-terminal kinase, but not p38
mitogen-activated protein kinase. This is further supported by the
data showing that A beta causes phosphorylation of ERK-1/ERK-2,
which, in turn, activates Elk-1. Furthermore, A beta mediated a
time-dependent increase in DNA binding activity of early growth
response-1 (Egr-1) and AP-1, but not of NF-kappa B and CREB.
Moreover, A beta-induced Egr-1 DNA binding activity was reduced >60%
in THP-1 cells transfected with small interfering RNA duplexes for
Egr-1 mRNA. We show that A beta-induced expression of TNF-alpha,
IL-1 beta, MCP-1, IL-8, and MIP-1 beta was abrogated in Egr-1 small
inhibitory RNA-transfected cells. Our results indicate that A
beta-induced expression of cytokines (TNF-alpha and IL-1 beta) and
chemokines (MCP-1, IL-8, and MIP-1 beta) in THP-1 monocytes involves
activation of ERK-1/ERK-2 and downstream activation of Egr-1. The
inhibition of Egr-1 by Egr-1 small inhibitory RNA may represent a
potential therapeutic target to ameliorate the inflammation and
progression of AD
Grant R, Kapoor V (2003) Inhibition of indoleamine
2,3-dioxygenase activity in IFN-gamma stimulated astroglioma cells
decreases intracellular NAD levels. Biochem.Pharmacol.
66:1033-1036
Abstract: Astroglia provide essential metabolic and
neurotropic support to cells within the CNS and participate in the
cellular immune response with Microglia/macrophages
following activation by the pro-inflammatory cytokine IFN-gamma.
Activation of glial cells results in local oxidative stress and
induction of a number of proteins including the enzyme indoleamine
2,3-dioxygenase (IDO). As a rate-limiting enzyme, IDO regulates
tryptophan catabolism via the kynurenine pathway producing a series
of metabolic precursors (some of which are neurotoxic) before
complete oxidation to the essential pyridine nucleotide NAD.
Inhibition of this pathway may therefore prove therapeutic in
neuroinflammatory disease by reducing production of cell toxins.
However, kynurenine metabolism may also be cytoprotective through de
novo synthesis of cellular NAD levels. We investigated the
hypothesis that IDO activity is directly involved in maintenance of
intracellular [NAD] in activated astroglial cells through control of
de novo synthesis. Exposure to IFN-gamma increased IDO activity from
7+/-1 nmol to 129+/-11 nmol kynurenine/hr/mg protein. Inhibition of
IDO activity with either 6-chloro-D-tryptophan (competitive
inhibition), or 3-ethoxy beta-carboline (non-competitive inhibition)
resulted in a dose-dependent decrease in IDO activity that
correlated directly with decreasing [NAD] (R(2)=0.92 and 0.81,
respectively). These results support the hypothesis that one
important consequence of increasing IDO activity in astroglial cells
during inflammation is to maintain NAD levels through de novo
synthesis from tryptophan. Inhibition of kynurenine pathway
metabolism under these conditions may significantly decrease cell
viability and CNS functions unless alternate precursors for NAD
synthesis are available
Kang JQ, Chong ZZ, Maiese K (2003) Critical role for Akt1 in
the modulation of apoptotic phosphatidylserine exposure and
Microglial
activation. Mol.Pharmacol. 64:557-569
Abstract: Biological
targets for neurodegenerative disease that focus on the intrinsic
maintenance of cellular integrity and the extrinsic prevention of
phagocytic cellular disposal offer the greatest promise for
therapeutic intervention. Protein kinase B (Akt1), a
serine-threonine kinase closely involved in cell growth and
survival, offers a strong potential to address both intrinsic and
extrinsic mechanisms of neuronal injury. We demonstrate that
overexpression of a constitutively active form of Akt1
(myristoylated Akt1) in differentiated SH-SY5Y neuronal cells
provides intrinsic cellular protection against apoptotic genomic DNA
destruction and membrane phosphatidylserine (PS) exposure.
Transfection of SH-SY5Y cells with a plasmid encoding a
kinase-deficient dominant-negative Akt1 eliminates cytoprotection,
suggesting that activation of Akt1 is necessary and sufficient to
prevent apoptotic destruction. Apoptotic neuronal membrane PS
exposure provides a unique pathway for Akt1 to offer extrinsic
cellular protection and block Microglial
activation, because independent cotreatment with an anti-PS receptor
neutralizing antibody could also prevent Microglial
proliferation. Akt1 maintains nuclear DNA integrity and membrane PS
exposure through the specific inhibition of caspase 3-, 8-, and
9-like activities that were linked to mitochondrial membrane
potential and cytochrome c release. Our work elucidates a novel
capacity for Akt1 to maintain cellular integrity through a series of
cysteine protease pathways and to uniquely regulate Microglial
activation through the modulation of membrane PS residue
externalization
Kang JQ, Chong ZZ, Maiese K (2003) Akt1 protects against
inflammatory Microglial
activation through maintenance of membrane asymmetry and modulation
of cysteine protease activity. J.Neurosci.Res. 74:37-51
Abstract:
In several cell systems, protein kinase B (Akt1) can promote cell
growth and development, but the "antiapoptotic" pathways
of this kinase that may offer protection against cellular
inflammatory demise have not been defined. Given that early cellular
membrane phosphatidylserine exposure is a critical component of
apoptosis, we investigated the role of Akt1 during neuronal
apoptotic injury. By employing differentiated SH-SY5Y neuronal cells
that overexpress a constitutively active form of Akt1 (myristoylated
Akt1), free radical-induced cell injury was assessed through trypan
blue dye exclusion, DNA fragmentation, membrane phosphatidylserine
exposure, protein kinase B phosphorylation, cysteine protease
activity, and mitochondrial membrane potential. Membrane
phosphatidylserine exposure was both necessary and sufficient for
Microglial
activation, insofar as cotreatment with an antiphosphatidylserine
receptor-neutralizing antibody could prevent Microglial
activity following neuronal loss of membrane asymmetry. Furthermore,
expression of myristoylated Akt1 not only prevented cell injury
through the prevention of membrane phosphatidylserine exposure and
genomic DNA fragmentation but also inhibited Microglial
activation and proliferation that required the inhibition of caspase
9-, caspase 3-, and caspase 1-like activities linked to cytochrome c
release. Interestingly, Akt1 modulation of membrane
phosphatidylserine exposure was primarily through caspase 1
activity. Removal of Akt1 activity abolished neuronal protection,
suggesting that Akt1 functions as a critical pathway for the
maintenance of cellular integrity and the prevention of phagocytic
cellular removal during neurodegenerative insults
Klegeris A, McGeer PL (2003) Toxicity of human monocytic
THP-1 cells and Microglia
toward SH-SY5Y neuroblastoma cells is reduced by inhibitors of
5-lipoxygenase and its activating protein FLAP. J.Leukoc.Biol.
73:369-378
Abstract: To explore whether the proinflammatory
products of the 5-lipoxygenase (5-LOX) pathway are involved in
Microglia-mediated
toxicity toward neuronal cells, we evaluated the effects of 5-LOX
inhibitors using an in vitro assay system where human neuronal
SH-SY5Y cells are exposed to toxic secretions from THP-1 monocytic
cells or human Microglia.
The specific 5-LOX inhibitors, REV 5901, zileuton, and
5-hydroxyeicosatetraenoic acid lactone; the nonselective LOX
inhibitors, phenidone and dapsone; the dual 5-LOX/cyclooxygenase
inhibitor, tepoxalin; and the selective inhibitor of the
5-LOX-activating protein (FLAP), MK-886, inhibited such toxicity.
The toxicity was enhanced by the 5-LOX product leukotriene (LT)D(4)
and reduced by the selective cysteinyl LT receptor (CysLT(1))
antagonist MK-571. The mRNAs for 5-LOX and FLAP were detected in
THP-1 cells and human Microglia
but not in SH-SY5Y cells. The data suggest that inhibition of
proinflammatory LT production by 5-LOX inhibition could selectively
reduce toxicity of Microglial
cells and thus be beneficial in neuroinflammatory diseases
Lambert JC, Luedecking-Zimmer E, Merrot S, Hayes A, Thaker U,
Desai P, Houzet A, Hermant X, Cottel D, Pritchard A, Iwatsubo T,
Pasquier F, Frigard B, Conneally PM, Chartier-Harlin MC, DeKosky ST,
Lendon C, Mann D, Kamboh MI, Amouyel P (2003) Association of 3'-UTR
polymorphisms of the oxidised LDL receptor 1 (OLR1) gene with
Alzheimer's disease. J.Med.Genet. 40:424-430
Abstract: Although
possession of the epsilon 4 allele of the apolipoprotein E gene
appears to be an important biological marker for Alzheimer's disease
(AD) susceptibility, strong evidence indicates that at least one
additional risk gene exists on chromosome 12. Here, we describe an
association of the 3'-UTR +1073 C/T polymorphism of the OLR1
(oxidised LDL receptor 1) on chromosome 12 with AD in French
sporadic (589 cases and 663 controls) and American familial (230
affected sibs and 143 unaffected sibs) populations. The age and sex
adjusted odds ratio between the CC+CT genotypes versus the TT
genotypes was 1.56 (p=0.001) in the French sample and 1.92 (p=0.02)
in the American sample. Furthermore, we have discovered a new T/A
polymorphism two bases upstream of the +1073 C/T polymorphism. This
+1071 T/A polymorphism was not associated with the disease, although
it may weakly modulate the impact of the +1073 C/T polymorphism.
Using 3'-UTR sequence probes, we have observed specific DNA protein
binding with nuclear proteins from lymphocyte, astrocytoma, and
neuroblastoma cell lines, but not from the Microglia
cell line. This binding was modified by both the +1071 T/A and +1073
C/T polymorphisms. In addition, a trend was observed between the
presence or absence of the +1073 C allele and the level of
astrocytic activation in the brain of AD cases. However, Abeta(40),
Abeta(42), Abeta total, and Tau loads or the level of Microglial
cell activation were not modulated by the 3'-UTR OLR1 polymorphisms.
Finally, we assessed the impact of these polymorphisms on the level
of OLR1 expression in lymphocytes from AD cases compared with
controls. The OLR1 expression was significantly lower in AD cases
bearing the CC and CT genotypes compared with controls with the same
genotypes. In conclusion, our data suggest that genetic variation in
the OLR1 gene may modify the risk of AD
Lange-Dohna C, Zeitschel U, Gaunitz F, Perez-Polo JR, Bigl V,
Rossner S (2003) Cloning and expression of the rat BACE1 promoter.
J.Neurosci.Res. 73:73-80
Abstract: The pathogenic processing of
the amyloid precursor protein (APP) into beta-amyloid peptides,
which give rise to beta-amyloid plaques in the brains of Alzheimer's
disease patients, requires the enzymatic activity of the beta-site
APP-cleaving enzyme 1 (BACE1). We report the cloning and sequence of
a 1.5-kb DNA fragment upstream of the coding sequence of the rat
BACE1 gene and the construction of a BACE1 promoter/luciferase
reporter construct. The basal activity of this promoter construct
was highest in neuronal cell lines such as BE(2)-C and PC12 and in
the pancreatic cell line AR42J, somewhat lower in rat primary
neurons, and astrocytic and Microglial
cultures, very low in hepatocytes, and almost absent in fibroblasts
and in the monocyte-macrophage cell line RAW264.7. The first 600 bp
of this promoter are highly conserved among rat, mouse, and human,
suggesting that this region contains regulatory elements that
modulate BACE1 transcription. Indeed, this fragment contains several
putative transcription factor binding sites such as MZF1, Sp1, four
GATA-1 sites, and one YY1 site. Directed mutagenesis of GATA-1
elements led to altered luciferase expression, indicating that these
sites are involved in the regulation of BACE1 transcription.
Additionally, the analysis of promoter activities of deletion
mutants suggests the presence of activators of BACE1 transcription
between bases -514 to -753 and of suppressor elements between bases
-754 and -1541. The BACE1 promoter sequence data and the constructs
described here will be useful to identify factors that influence the
expression of BACE1 in experimental paradigms in vitro
Luder CG, Lang C, Giraldo-Velasquez M, Algner M, Gerdes J,
Gross U (2003) Toxoplasma gondii inhibits MHC class II expression in
neural antigen-presenting cells by down-regulating the class II
transactivator CIITA. J.Neuroimmunol. 134:12-24
Abstract: Major
histocompatibility complex (MHC) class II expression by Microglia
and astrocytes is critical for CD4+-mediated immune responses within
the central nervous system. Here, we demonstrate that the obligate
intracellular parasite, Toxoplasma gondii, down-regulates
activation-induced MHC class II expression in human-derived
glioblastoma cells as well as in primary astrocytes and Microglia
from cortices of rat fetuses. Down-regulation of MHC class II
proteins was predominantly observed in parasite-positive, but not
parasite-negative, host cells of T. gondii-infected cell cultures.
MHC class II transcript levels induced by IFN-gamma alone or in
combination with TNF-alpha were also clearly diminished after
parasitic infection. Furthermore, T. gondii dose-dependently
down-regulated the transcript levels of the class II transactivator
CIITA. These results suggest that T. gondii partially evade
CD4+-mediated intracerebral immune responses, a mechanism which may
contribute to long-term persistence of the parasite within the CNS
Munch G, Gasic-Milenkovic J, Dukic-Stefanovic S, Kuhla B,
Heinrich K, Riederer P, Huttunen HJ, Founds H, Sajithlal G (2003)
Microglial
activation induces cell death, inhibits neurite outgrowth and causes
neurite retraction of differentiated neuroblastoma cells. Exp.Brain
Res. 150:1-8
Abstract: Activation of glial cells has been
proposed to contribute to neuronal dysfunction and neuronal cell
death in Alzheimer's disease. In this study, we attempt to determine
some of the effects of secreted factors from activated murine N-11
Microglia
on viability and morphology of neurons using the differentiated
neuroblastoma cell line Neuro2a. Microglia
were activated either by lipopolysaccharide (LPS), bacterial cell
wall proteoglycans, or advanced glycation endproducts (AGEs),
protein-bound sugar oxidation products. At high LPS or AGE
concentrations, conditioned medium from Microglia
caused neuronal cell death in a dose-dependent manner. At sublethal
LPS or AGE concentrations, conditioned media inhibited retinoic
acid-induced neurite outgrowth and stimulated retraction of already
extended neurites. Among the many possible secreted factors, the
contribution of NO or NO metabolites in the cytotoxicity of
conditioned medium was investigated. Cell death and changes in
neurite morphology were partly reduced when NO production was
inhibited by nitric oxide synthase inhibitors. The results suggest
that even in the absence of significant cell death, inflammatory
processes, which are partly transmitted via NO metabolites, may
affect intrinsic functions of neurons such as neurite extension that
are essential components of neuronal morphology and thus may
contribute to degenerative changes in Alzheimer's disease
Phuong LK, Allen C, Peng KW, Giannini C, Greiner S, TenEyck
CJ, Mishra PK, Macura SI, Russell SJ, Galanis EC (2003) Use of a
vaccine strain of measles virus genetically engineered to produce
carcinoembryonic antigen as a novel therapeutic agent against
glioblastoma multiforme. Cancer Res. 63:2462-2469
Abstract:
Despite the most aggressive medical and surgical treatments,
glioblastoma multiforme remains incurable with a median survival of
<1 year. We investigated the antitumor
potential of a novel viral agent, an attenuated strain of
measles virus (MV), derived from the Edmonston vaccine lineage,
genetically engineered to produce carcinoembryonic antigen (CEA).
CEA production as the virus replicates can serve as a marker of
viral gene expression. Infection of a variety of glioblastoma cell
lines including U87, U118, and U251 at MOIs 0.1, 1, and 10 resulted
in significant cytopathic effect consisting of excessive syncycial
formation and massive cell death at 72-96 h from infection. terminal
deoxynucleotidyltransferase-mediated nick end labeling assays
demonstrated the mechanism of cell death to be predominantly
apoptotic. The efficacy of this approach in vivo was examined in
BALB/c nude mice by using both s.c. and intracranial orthotopic U87
tumor models. In the s.c.
U87 model, mice with established xenografts were treated with a
total dose of 8 x 10(7) plaque forming units of MV-CEA, administered
i.v. Mice treated with UV light inactivated MV, and untreated mice
with established U87 tumors were used as controls. There was
statistically significant regression of s.c. tumors (P < 0.001)
and prolongation of survival (P = 0.007) in MV-CEA treated animals
compared with the two control groups. In the intracranial orthotopic
U87 model, there was significant regression of intracranial U87
tumors treated with intratumoral administration of MV-CEA at a total
dose of 1.8 x 10(6) plaque forming units as assessed by magnetic
resonance image (P = 0.002), and statistically significant
prolongation of survival as compared with mice that received
UV-inactivated virus and untreated mice (P = 0.02). Histological
examination of brains of MV-CEA-treated animals revealed complete
regression of the tumor with
the presence of a residual glial scar and reactive changes, mainly
presence of hemosiderin-laden macrophages. In addition, CEA levels
in the peripheral blood in both the s.c. and orthotopic models
increased before tumor regression,
indicating viral gene expression, and returned to normal when the
tumors regressed. Ifnar(ko) CD46 Ge transgenic mice, susceptible to
MV infection, were used to assess central nervous system toxicity of
MV-CEA. Intracranial administration of MV-CEA into the caudate
nucleus of Ifnar(ko) CD46 Ge did not result in clinical
neurotoxicity. Pathologic examination demonstrated limited
Microglial
infiltration surrounding the injection site. In summary, MV-CEA has
potent antitumor activity
against gliomas in vitro, as well as in both s.c. and orthotopic U87
animal models. Monitoring CEA levels in the serum can serve as a
low-risk method of detecting viral gene expression during treatment,
and could allow dose optimization and individualization of treatment
Platten M, Kretz A, Naumann U, Aulwurm S, Egashira K,
Isenmann S, Weller M (2003) Monocyte chemoattractant protein-1
increases Microglial
infiltration and aggressiveness of gliomas. Ann.Neurol.
54:388-392
Abstract: Macrophages are thought to represent a first
line of defense in anti-tumor
immunity. Despite infiltration by Microglial
cells, however, malignant gliomas are still highly aggressive
tumors. We here identify monocyte chemoattractant protein-1 (MCP-1)
as a critical chemoattractant for glioma-infiltrating Microglial
cells. MCP-1-transfected rat CNS-1 gliomas were massively
infiltrated by Microglial
cells. Whereas MCP-1 did not promote the growth of CNS-1 cells in
vitro, intracerebral CNS-1-transfected tumors grew more aggressively
than control-transfected tumors. This provides the first functional
evidence that MCP-1 recruits Microglial
cells to gliomas and promotes their growth in vivo. Microglial
cells may support rather than suppress glioma growth
Sasaki A, Horikoshi Y, Yokoo H, Nakazato Y, Yamaguchi H
(2003) Antiserum against human glucose transporter 5 is highly
specific for Microglia
among cells of the mononuclear phagocyte system. Neurosci.Lett.
338:17-20
Abstract: Human monocytes and a variety of tissue
macrophages, including Microglia,
were studied immunohistochemically to determine the expression of a
novel Microglial
marker, human glucose transporter 5 (hGLUT5), in these cells. The
hGLUT5 was not expressed in most peripheral macrophages in the
normal state, but weakly expressed in some foamy macrophages in
atherosclerotic lesions. There was no hGLUT5 reactivity in blood
monocytes. In the lesions of brain infarcts, foamy macrophages
(predominantly monocyte-derived cells) in the ischemic core were
mostly negative for hGLUT5, while activated and phagocytic Microglia
in the transitional zone were consistently positive. The present
study indicated that unlike other Microglial
markers, hGLUT5 is rarely present in peripheral macrophages, and
that hGLUT5 immunohistochemistry is useful in distinguishing
Microglia-derived
macrophages from monocyte-derived macrophages in acute necrotic
lesions
Shimizu T, Sato K, Suzuki T, Tachibana K, Takeda K (2003)
Induction of plasminogen activator inhibitor-2 is associated with
suppression of invasive activity in TPA-mediated differentiation of
human prostate cancer cells. Biochem.Biophys.Res.Commun.
309:267-271
Abstract: We previously reported that
12-O-tetra-decanoylphorbol-13-acetate (TPA) induces Microglia-like
differentiation and decreases malignancy in human prostate cancer
TSU-Pr1 cells. To investigate the mechanism underlying
differentiation and decrease of malignancy in TSU-Pr1 cells treated
with TPA, we attempted to identify genes expressed differentially
during the differentiation using differential display. We
successfully detected plasminogen activator inhibitor type-2 (PAI-2)
as one gene up-regulated by TPA treatment. The change in expression
of PAI-2 by TPA was blocked by treatment with protein kinase C or
mitogen-activated protein kinase inhibitors. We also found that
secretion of PAI-2 protein was increased by TPA treatment. Moreover,
we demonstrated that suppression of invasive activity of TSU-Pr1
cells by TPA treatment was blocked by co-treatment with anti-PAI-2
antibody. These results suggest that induction of PAI-2 is
associated with suppression of invasive activity in TSU-Pr1 cells
treated with TPA
Ando H, Saio M, Ohe N, Tamakawa N, Yu H, Nakayama T,
Yoshimura S, Kaku Y, Iwama T, Shinoda J, Sakai N, Takami T (2002)
B7.1 immunogene therapy effectively activates CD(4+)
tumor-infiltrating lymphocytes in the central nervous system in
comparison with B7.2 gene therapy. Int.J.Oncol. 20:807-812
Abstract:
The B7 gene utilizing immunogene therapy is one of the most common
methods against tumor growth.
However, there is no known study that investigated the difference
between B7.1 and B7.2 with regard to B7 gene therapy in the central
nervous system (CNS). Therefore, to clarify the difference, we
established B7.1 or B7.2 gene transduced tumor
cells originating from the murine T cell lymphoma cell line
EL4 (EL4-B7.1 or EL4-B7.2). First, we observed the survival time
after intracranial inoculation of parent (IC-wt) or genetically
modified tumor cells. All
mice in control groups (IC-wt or IC-mock) were dead within 16 days.
While there was significant survival elongation in the B7.2 modified
group (IC-B7.2, p=0.0002), all mice in this group were dead of tumor
growth within 22 days. On the other hand, 60% of mice
inoculated with EL4-B7.1 (IC-B7.1) survived more than 120 days
(p<0.0001). Second, to shed light on the anti-tumor
immune response in situ, we tried to analyze CD(4+)
tumor-infiltrating T lymphocytes (CD(4+) TIL). To purify and analyze
CD(4+) TIL, we had to deplete F4/80(+) Microglia
because of the CD4 expression. In terms of activation marker
expression in CD(4+) TIL, a small population was activated (CD25,
9.8%; CD69, 15.8%) in the control group (IC-wt). In contrast, the
activation marker positive CD4+ TIL percentage both in IC-B7.1
(CD25, 25.1%; CD69, 40.1%) and IC-B7.2 (CD25, 16.2%; CD69, 28.3%)
appeared to reflect the survival curve in both groups. These
findings strongly suggest that, in the CNS, B7.1 gene therapy could
effectively introduce CD(4+) TIL activation compared with B7.2 gene
therapy. This is the first study clearly describing the difference
between B7.1 gene therapy and B7.2 gene therapy in the CNS in terms
of the activation status of CD(4+) TIL in situ
Badie B, Bartley B, Schartner J (2002) Differential
expression of MHC class II and B7 costimulatory molecules by
Microglia
in rodent gliomas. J.Neuroimmunol. 133:39-45
Abstract: To assess
the immune function of Microglia
and macrophages in brain tumors, the expression of MHC class II and
B7 costimulatory molecules in three rodent glioma models was
examined. Microglia
and macrophages, which accounted for 5-12% of total cells, expressed
B7.1 and MHC class II molecules in the C6 and 9L tumors, but not RG2
gliomas. Interestingly, the expression of B7.1 and MHC class II
molecules by Microglia
and macrophage was associated with an increase in the number of
tumor-infiltrating lymphocytes in C6 and 9L tumors. B7.2 expression,
which was present at low levels on Microglia
and macrophages in normal brain, did not significantly change in
tumors. Interestingly, the expression of all three surface antigens
increased after Microglia
were isolated from intracranial C6 tumors and cultured for a short
period of time. We conclude that Microglia
immune activity may be suppressed in gliomas and directly correlates
to the immunogenecity of experimental brain tumors
Bate C, Rutherford S, Gravenor M, Reid S, Williams A (2002)
Cyclo-oxygenase inhibitors protect against prion-induced
neurotoxicity in vitro. Neuroreport 13:1933-1938
Abstract: The
mechanisms of neuronal loss during the course of the prion diseases
are not fully understood. In this study, neurones treated with
certain non-steroidal anti-inflammatory drugs (NSAIDs) were
protected against the otherwise toxic effects of a peptide derived
from the prion protein, or extracts containing infectious prions
(PrP ). These NSAIDs inhibit the cyclo-oxygenase (cox) enzymes that
metabolise arachidonic acid to prostaglandins (PG). Conversely,
drugs that inhibited the metabolism of arachidonic acid to
leucotrienes enhanced neurotoxicity. Studies with selective
inhibitors highlighted the importance of the cox-1 isoform in
prion-induced neurotoxicity. The cox-1 inhibitors also inhibited
neuronal PGE production and protected both neuroblastoma cells and
primary cortical neurones against prions. They also reduced
Microglia-mediated
killing of prion-treated neurones
Fukumitsu H, Takase-Yoden S, Furukawa S, Nemoto K, Ikeda T,
Watanabe R (2002) Implantation of BDNF-producing packaging cells
into brain. Cell Transplant. 11:459-464
Abstract: In order to
invent a screening system to check in vivo gene function and the
efficiency of gene transfer mediated by a retroviral vector system,
we established a novel packaging cell, PacC6/A8, that is
transplantable to rat brains. The packaging cell is based on the
gene of the neuropatogenic retrovirus, A8-V. For expression in the
brain, a vector that expresses brain-derived neurotrophic factor
(BDNF) tagged by c-Myc-His6 (LxA/bdmh) was constructed. After
transfection of LxA/bdmh to PacC6/A8, a cloned cell line,
PacC6/A8/bmh, was established. PacC6/A8/bmh cells stably produced
pseudotyped retroviruses carrying LxA/bdmh. For a control, a
retroviral vector that bears the gene that codes enhanced green
fluorescent protein (EGFP) tagged by C-Mic-His6 was also created and
used for the establishment of PacC6/A8/gfmh cells that produce
pseudotyped retroviruses carrying LxA/gfmh. PacC6/A8/bmh and
PacC6/A8/gfmh cells were injected to the brain of newborn rats. A
tumor was formed in all the
rats injected that did not exhibit any symptoms until 3-4 weeks
after the injection. A histological study of the injected rats
revealed that the transferred BDNF gene was expressed in the brain
of rats injected with PacC6/A8/bmh cells, but not in rats with
PacC6/A8/gfmh cells. Interestingly, many activated Microglia
had migrated into the tumor
induced by PacC6/A8/bmh cells, and expressed a high amount of
BDNF
Johnston JB, Silva C, Power C (2002) Envelope gene-mediated
neurovirulence in feline immunodeficiency virus infection: induction
of matrix metalloproteinases and neuronal injury. J.Virol.
76:2622-2633
Abstract: The release of neurotoxins by activated
brain macrophages or Microglia
is one mechanism proposed to contribute to the development of
neurological disease following infection by lentiviruses, including
feline immunodeficiency virus (FIV). Since molecular diversity in
the lentiviral envelope gene influences the expression of host
molecules implicated in neuronal injury, the role of the envelope
sequence in FIV neuropathogenesis was investigated by using the
neurovirulent FIV strain V1CSF, the nonneurovirulent strain
Petaluma, and a chimera (FIVCh) containing the V1CSF envelope gene
in a Petaluma background. All three viruses replicated in primary
feline macrophages with equal efficiency, but conditioned medium
from V
Klegeris A, McGeer PL (2002) Cyclooxygenase and
5-lipoxygenase inhibitors protect against mononuclear phagocyte
neurotoxicity. Neurobiol.Aging 23:787-794
Abstract:
Neuroinflammation and oxidative stress are believed to be
contributing factors to neurodegeneration in normal aging, as well
as in age-related neurological disorders. Reactive Microglia
are found in increased numbers in aging brain and are prominently
associated with lesions in such age-related degenerative conditions
as Alzheimer's disease (AD), Parkinson's disease (PD) and
amyotrophic lateral sclerosis (ALS). In vitro, stimulated Microglia
or Microglial-like
cells secrete neurotoxic materials and are generators of free
radicals through their respiratory burst system. Agents that
suppress Microglial
activation are therefore candidates for neuroprotection. We have
developed quantitative in vitro assays for measuring neurotoxicity
of Microglia
or other mononuclear phagocytes. Neuronal like SH-SY5Y cells are
cultured in supernatants from activated cells of the human monocytic
THP-1 line and their survival is followed. Respiratory burst is
directly measured on the activated cells. We tested inhibitors of
the cyclooxygenase (COX) or the 5-lipoxygenase (5-LOX) pathways as
possible neuroprotective agents. The COX pathway generates
inflammatory prostaglandins, while the 5-LOX pathway generates
inflammatory leukotrienes. We found that inhibitors of both these
pathways suppressed neurotoxicity in a dose-dependent fashion. They
included the COX-1 inhibitor indomethacin; the COX-2 inhibitor
NS-398; the mixed COX-1/COX-2 inhibitor ibuprofen; the nitric oxide
(NO) derivatives of indomethacin, ibuprofen and flurbiprofen; the
5-LOX inhibitor REV 5901; and the 5-LOX activating protein (FLAP)
inhibitor MK-886. The FLAP inhibitor also reduced respiratory burst
activity in a more potent manner than indomethacin. Combinations of
COX and 5-LOX inhibitors were more effective than single inhibitors.
The data suggest that both COX inhibitors and 5-LOX inhibitors may
be neuroprotective in vivo by suppressing toxic actions of
Microglia/macrophages,
and that combinations of the two might have greater therapeutic
potential than single inhibitors of either class
Linden DR, El Fakahany EE (2002) Microglial
derived nitric oxide decreases serotonin content in rat basophilic
leukemia (RBL-2H3) cells. Eur.J.Pharmacol. 436:53-56
Abstract:
Nitric oxide (NO) and serotonin (5-hydroxytryptamine; 5-HT) are
important neuromodulators that are involved in a myriad of
biochemical reactions. In this work, we describe a novel model
co-culture system to study the interactions between NO and 5-HT. NO
derived from cytokine stimulated Bv2 Microglial
cells depleted 5-HT from RBL-2H3 cells. Reduction of 5-HT content by
NO derived from the NO donor S-nitroso-N-acetylpenicillamine (SNAP)
was concentration-dependent, independent of intracellular Ca(2+) and
inhibited by reduced glutathione (GSH). Collectively, these data
indicate that this cell co-culture system is a viable model to study
the mechanisms of interaction between nitrergic and serotonergic
pathways
Liu H, Ng CE, Tang BL (2002) Nogo-A expression in mouse
central nervous system neurons. Neurosci.Lett. 328:257-260
Abstract:
The longest central nervous system (CNS) specific isoform of the
major myelin-associated inhibitor of neurite growth, Nogo-A, has
previously been shown to be expressed largely in oligodendrocytes.
Using an antibody raised against a recombinant fusion protein
comprising amino acids 223-399 of Nogo-A, we show in this report
that Nogo-A is also expressed in the cell body of a distinct set of
CNS neurons. The antibody detects a single protein band of 220 kDa
in rat brain lysate and neuroblastoma cell lysates.
Immunofluorescent analyses reveal that Nogo-A is found largely in
the endoplasmic reticulum of neuroblastoma cell lines SH-SY5Y and
NIE-115. In the mouse CNS, Nogo-A can be found in specific subsets
of neuronal cell bodies in addition to oligodendrocytes, but not
glial fibrillary associated protein positive astrocytes or MAC-1
positive Microglia.
Our results provide a conclusive demonstration of the expression of
Nogo-A in CNS neurons, which suggests that Nogo-A may have distinct
endogenous roles in neurons other than its known ability to inhibit
neurite growth
Madigan MC, Penfold PL, King NJ, Billson FA, Conway RM (2002)
Immunoglobulin superfamily expression in primary retinoblastoma and
retinoblastoma cell lines. Oncol.Res. 13:103-111
Abstract:
Retinoblastoma (Rb) is the most common intraocular tumor
of childhood. In this study we examined primary Rb specimens
and Rb cell lines for the expression of immunoglobulin superfamily
(IgSF) antigens: MHC class I and II (MHC-I and MHC-II), neural cell
adhesion molecule (NCAM), intercellular adhesion molecule-1
(ICAM-1), and Thy-1, which play an important role in immune system
and tumor cell
interactions. MHC-I and-II, ICAM-1 (CD54), NCAM (CD56), and Thy-1
(CDw90) immunoreactivity was studied in eight primary Rb biopsy
specimens using immunohistochemistry, three using immunoelectron
microscopy, and six Rb cell lines using flow cytometry (FCM).
Parenchymal and vascular-associated cells, phenotypically similar to
retinal Microglia,
strongly expressed MHC-II immunoreactivity and were distributed
throughout primary Rb specimens. However, MHC-II expression on Rb
cell lines was similar to nonspecific control levels. tumor
cells in primary Rb specimens displayed high NCAM, moderate
Thy-1, and low MHC-I and ICAM-1 immunolabeling. tumor
vasculature expressed low to moderate MHC-I and ICAM-1
immunoreactivity and moderate Thy-1 immunoreactivity. NCAM was not
detected on the vasculature of primary Rb specimens. Rb cell lines
displayed variable expression of Thy-1, ICAM-1, and MHC-I. NCAM was
highly expressed on five of six Rb cell lines. The high levels of
constitutive NCAM immunoreactivity on Rb tumor
cells confirm the neuroectodermal origins of this tumor.
Additionally, the variable expression of Thy-1 may suggest separate
neural lineages or differences in the maturational status ofsome Rb
tumors. The presence of a population of infiltrating MHC-II-positive
cells in primary Rb tumors has implications for immunomodulation of
Rb growth
Mentlein R, Held-Feindt J (2002) Pleiotrophin, an angiogenic
and mitogenic growth factor, is expressed in human gliomas.
J.Neurochem. 83:747-753
Abstract: Pleiotrophin (PTN) is a
mitogenic/angiogenic, 15.3 kDa heparin-binding peptide that is found
in embryonic or early postnatal, but rarely in adult, tissues. Since
developmentally regulated factors often re-appear in malignant
cells, we examined PTN expression in human glioma cell lines, cell
cultures derived from solid gliomas and glioma sections. PTN mRNA or
protein was detected by reverse transcriptase-polymerase chain
reaction, immunohistochemistry, western blot or enzyme-linked
immunoassay in all WHO III and IV grade gliomas and cells analyzed
in vitro or in situ. One WHO II grade glioma investigated was PTN
negative. In vitro, PTN was synthesized in perinuclear regions of
glioma cells, secreted into the cultivation medium, but its
production varied considerably between glioma cells cultivated from
different solid gliomas or glioma cell lines. In situ, PTN
expression was restricted to distinct parts/cells of the tumour. PTN
did not influence the proliferation of glioma cells themselves, but
stimulated [3H]thymidine incorporation into DNA of Microglial
cells. Furthermore, in Boyden chamber assays, PTN showed a strong
chemotactic effect on murine BV-2 Microglial
cells. PTN is supposed to be a paracrine growth/angiogenic factor
that is produced by gliomas and contributes to their malignancy by
targeting endothelial and Microglial
cells
Perini G, Della-Bianca V, Politi V, Della VG, Dal Pra I,
Rossi F, Armato U (2002) Role of p75 neurotrophin receptor in the
neurotoxicity by beta-amyloid peptides and synergistic effect of
inflammatory cytokines. J.Exp.Med. 195:907-918
Abstract: The
neurodegenerative changes in Alzheimer's disease (AD) are elicited
by the accumulation of beta-amyloid peptides (Abeta), which damage
neurons either directly by interacting with components of the cell
surface to trigger cell death signaling or indirectly by activating
astrocytes and Microglia
to produce inflammatory mediators. It has been recently proposed
that the p75 neurotrophin receptor (p75(NTR)) is responsible for
neuronal damage by interacting with Abeta. By using neuroblastoma
cell clones lacking the expression of all neurotrophin receptors or
engineered to express full-length or various truncated forms of
p75(NTR), we could show that p75(NTR) is involved in the direct
signaling of cell death by Abeta via the function of its death
domain. This signaling leads to the activation of caspases-8 and -3,
the production of reactive oxygen intermediates and the induction of
an oxidative stress. We also found that the direct and indirect
(inflammatory) mechanisms of neuronal damage by Abeta could act
synergistically. In fact, TNF-alpha and IL-1beta, cytokines produced
by Abeta-activated Microglia,
could potentiate the neurotoxic action of Abeta mediated by p75(NTR)
signaling. Together, our results indicate that neurons expressing
p75(NTR), mostly if expressing also proinflammatory cytokine
receptors, might be preferential targets of the cytotoxic action of
Abeta in AD
Repovic P, Benveniste EN (2002) Prostaglandin E2 is a novel
inducer of oncostatin-M expression in macrophages and Microglia.
J.Neurosci. 22:5334-5343
Abstract: Oncostatin-M (OSM), a
pluripotent cytokine of the interleukin-6 (IL-6) family, is produced
in a number of inflammatory conditions. Known sources of OSM include
monocytes-macrophages and T-cells. Here we present Microglia,
the resident macrophages of the brain, as a source of OSM in the
CNS. In this context, we describe a novel inducer of OSM,
prostaglandin E(2) (PGE(2)). PGE(2) induces OSM expression in
Microglia,
monocytes, and macrophages of human and murine origin. PGE(2)
induction of OSM is mimicked by cholera toxin, an activator of
stimulatory G (G(s))-proteins; by forskolin, an activator of
adenylate cyclase; and by the cAMP analog, dibutyryl-cAMP. PGE(2)
induction of OSM gene expression is inhibited by the adenylate
cyclase inhibitor 2',5'-dideoxyadenosine, by the protein kinase A
(PKA) inhibitor H-89, and by a dominant-negative PKA construct.
These data indicate that PGE(2) signals via G(s)-protein-coupled
receptor(s), adenylate cyclase, and PKA to induce OSM expression.
Accordingly, other activators of cAMP signaling such as
norepinephrine and PGE(1) induce OSM. The ability of PGE(2) to
induce OSM expression was tested under more physiological
conditions, using cocultures of astrocytes and monocytes. Treatment
of the cocultures with IL-1beta or tumor
necrosis factor-alpha (TNF-alpha) results in production of
PGE(2) and OSM. PGE(2) produced in the cocultures is responsible for
OSM induction, because pretreatment with indomethacin, an inhibitor
of prostaglandin synthesis, as well as depletion of PGE(2), abrogate
OSM expression induced by IL-1beta or TNF-alpha. These data suggest
that in the CNS, OSM may be produced through collaboration of
astrocytes and macrophages-Microglia
Seo JH, Rah JC, Choi SH, Shin JK, Min K, Kim HS, Park CH, Kim
S, Kim EM, Lee SH, Lee S, Suh SW, Suh YH (2002) Alpha-synuclein
regulates neuronal survival via Bcl-2 family expression and PI3/Akt
kinase pathway. FASEB J. 16:1826-1828
Abstract: Alpha-synuclein
(alpha-SN) is a ubiquitous protein that is especially abundant in
the brain and has been postulated to play a central role in the
pathogenesis of Parkinson's disease, Alzheimer's disease, and other
neurodegenerative disorders. However, little is known about the
neuronal functions of alpha-SN and the molecular and cellular
mechanisms underlying neuronal loss. Here, we show that alpha-SN
plays dual roles of neuroprotection and neurotoxicity depending on
its concentration or level of expression. At nanomolar
concentrations, a-SN protected neurons against serum deprivation,
oxidative stress, and excitotoxicity through the PI3/Akt signaling
pathway, and its protective effect was increased by Bcl-2
overexpression. Conversely, at both low micromolar and overexpressed
levels in the cell, alpha-SN resulted in cytotoxicity. This might be
related to decreased Bcl-xL expression and increased bax expression,
which is subsequently followed by cytochrome c release and caspase
activation and also by Microglia-mediated
inflammatory responses via the NFkappaB and mitogen-activated
protein kinase pathways
Serhan CN, Hong S, Gronert K, Colgan SP, Devchand PR, Mirick
G, Moussignac RL (2002) Resolvins: a family of bioactive products of
omega-3 fatty acid transformation circuits initiated by aspirin
treatment that counter proinflammation signals. J.Exp.Med.
196:1025-1037
Abstract: Aspirin (ASA) is unique among current
therapies because it acetylates cyclooxygenase (COX)-2 enabling the
biosynthesis of R-containing precursors of endogenous
antiinflammatory mediators. Here, we report that lipidomic analysis
of exudates obtained in the resolution phase from mice treated with
ASA and docosahexaenoic acid (DHA) (C22:6) produce a novel family of
bioactive 17R-hydroxy-containing di- and tri-hydroxy-docosanoids
termed resolvins. Murine brain treated with aspirin produced
endogenous 17R-hydroxydocosahexaenoic acid as did human Microglial
cells. Human COX-2 converted DHA to 13-hydroxy-DHA that switched
with ASA to 17R-HDHA that also proved a major route in hypoxic
endothelial cells. Human neutrophils transformed COX-2-ASA-derived
17R-hydroxy-DHA into two sets of novel di- and trihydroxy products;
one initiated via oxygenation at carbon 7 and the other at carbon 4.
These compounds inhibited (IC(50) approximately 50 pM) Microglial
cell cytokine expression and in vivo dermal inflammation and
peritonitis at ng doses, reducing 40-80% leukocytic exudates. These
results indicate that exudates, vascular, leukocytes and neural
cells treated with aspirin convert DHA to novel 17R-hydroxy series
of docosanoids that are potent regulators. These biosynthetic
pathways utilize omega-3 DHA and EPA during multicellular events in
resolution to produce a family of protective compounds, i.e.,
resolvins, that enhance proresolution status
Tan J, Town T, Crawford F, Mori T, DelleDonne A, Crescentini
R, Obregon D, Flavell RA, Mullan MJ (2002) Role of CD40 ligand in
amyloidosis in transgenic Alzheimer's mice. Nat.Neurosci.
5:1288-1293
Abstract: We have shown that interaction of CD40 with
CD40L enables Microglial
activation in response to amyloid-beta peptide (Abeta), which is
associated with Alzheimer's disease (AD)-like neuronal tau
hyperphosphorylation in vivo. Here we report that transgenic mice
overproducing Abeta, but deficient in CD40L, showed decreased
astrocytosis and microgliosis associated with diminished Abeta
levels and beta-amyloid plaque load. Furthermore, in the PSAPP
transgenic mouse model of AD, a depleting antibody against CD40L
caused marked attenuation of Abeta/beta-amyloid pathology, which was
associated with decreased amyloidogenic processing of amyloid
precursor protein (APP) and increased circulating levels of Abeta.
Conversely, in neuroblastoma cells overexpressing wild-type human
APP, the CD40-CD40L interaction resulted in amyloidogenic APP
processing. These findings suggest several possible mechanisms
underlying mitigation of AD pathology in response to CD40L
depletion, and validate the CD40-CD40L interaction as a target for
therapeutic intervention in AD
Tanimukai S, Hasegawa H, Nakai M, Yagi K, Hirai M, Saito N,
Taniguchi T, Terashima A, Yasuda M, Kawamata T, Tanaka C (2002)
Nanomolar amyloid beta protein activates a specific PKC isoform
mediating phosphorylation of MARCKS in Neuro2A cells. Neuroreport
13:549-553
Abstract: Myristoylated alanine-rich C kinase
substrate (MARCKS), a protein associated with cell growth,
neurosecretion and macrophage activation, is activated by protein
kinase C (PKC) phosphorylation. We reported that amyloid beta
protein (Abeta) activated MARCKS through a tyrosine kinase and
PKC-delta in rat cultured Microglia.
Here we report that Abeta signaling pathway through a specific PKC
isoform is involved in the phosphorylation of MARCKS in Neuro2A
cells. Selective PKC inhibitors but not tyrosine kinase inhibitors
significantly inhibited the phosphorylation of MARCKS induced by
Abeta. Abeta selectively activated PKC-alpha among the four PKC
isoforms localized in Neuro2A cells. PKC-alpha activated by Abeta
directly phosphorylated a recombinant MARCKS in vitro, Translocation
of PKC-alpha from the cytoplasm to the membrane and accumulation of
phospho-MARCKS in the cytoplasm were induced by Abeta. These results
suggest involvement of a phosphoinositide signaling system through
PKC-alpha in the phosphorylation of MARCKS in neurons, an event
which may be associated with mechanisms underlying neurotrophic and
neurotoxic effects of Abeta
Traister RS, Lynch WP (2002) Reexamination of amphotropic
murine leukemia virus neurovirulence: neural stem cell-mediated
Microglial
infection fails to induce acute neurodegeneration. Virology
293:262-272
Abstract: The 4070A amphotropic murine leukemia virus
(A-MuLV) has been variably reported to harbor neurovirulence
determinants within its env gene. In this report we reexamined this
issue by applying two approaches previously demonstrated to amplify
murine leukemia virus neurovirulence. The first approach involved
introducing the 4070A env gene into the background of Friend virus
clone FB29 to enhance peripheral virus replication kinetics and
central nervous system entry. The resulting chimeric virus, FrAmE,
exhibited widespread vascular infection throughout the central
nervous system (CNS); however, parenchymal infection was quite
limited. Neither clinical neurological signs nor spongiform
neurological changes accompanied FrAmE CNS infection. To overcome
this CNS entry limitation, 4070A and FrAmE were delivered directly
into the CNS via transplantation of infected C17.2 neural stem cells
(NSCs). Significantly, NSC dissemination of either 4070A or FrAmE
resulted in widespread, high-level amphotropic virus expression
within the CNS parenchyma, including the infection of Microglia,
the critical target required for inducing neurodegeneration. Despite
the extensive CNS infection, no associated clinical neurological
signs or acute neuropathological changes were observed.
Interestingly, we observed the frequent appearance of circulating
polytropic (MCF) virus in the serum of amphotropic virus-infected
animals. However, neither peripheral inoculation of an
amphotropic/MCF virus mixture nor transplantation of NSCs expressing
both amphotropic and MCF viruses induced acute clinical neurological
signs or spongiform neuropathology. Thus, the results generated in
this study suggest that the 4070A env gene is not inherently
neurovirulent. However, the frequent appearance of endogenous MCF
viruses suggests the possibility that the interactions of
amphotropic viruses with endogenous retroviral elements could
contribute to the development of retrovirus-induced
neurodegenerative disease
Warntges S, Friedrich B, Henke G, Duranton C, Lang PA,
Waldegger S, Meyermann R, Kuhl D, Speckmann EJ, Obermuller N,
Witzgall R, Mack AF, Wagner HJ, Wagner A, Broer S, Lang F (2002)
Cerebral localization and regulation of the cell volume-sensitive
serum- and glucocorticoid-dependent kinase SGK1. Pflugers Arch.
443:617-624
Abstract: The serum- and glucocorticoid-dependent
kinase SGK1 is regulated by alterations of cell volume, whereby cell
shrinkage increases and cell swelling decreases the transcription,
expression and activity of SGK1. The kinase is expressed in all
human tissues studied including the brain. The present study was
performed to localize the sites of SGK1 transcription in the brain,
to elucidate the influence of the hydration status on SGK1
transcription and to explore the functional significance of altered
SGK1 expression. Northern blot analysis of human brain showed SGK1
to be expressed in all cerebral structures examined: amygdala,
caudate nucleus, corpus callosum, hippocampus, substantia nigra,
subthalamic nucleus and thalamus. In situ hybridization and
immunohistochemistry in the rat revealed increased expression of
SGK1 in neurons of the hippocampal area CA3 after dehydration,
compared with similar slices from brains of euvolaemic rats.
Additionally, several oligodendrocytes, a few Microglial
cells, but no astrocytes, were positive for SGK1. The abundance of
SGK1 mRNA in the temporal lobe, including hippocampus, was increased
by dehydration and SGK1 transcription in neuroblastoma cells was
stimulated by an increase of extracellular osmolarity. Co-expression
studies in Xenopus laevis oocytes revealed that SGK1 markedly
increased the activity of the neuronal K+ channel Kv1.3. As
activation of K+ channels modifies excitation of neuronal cells,
SGK1 may participate in the regulation of neuronal excitability
Badie B, Schartner J, Prabakaran S, Paul J, Vorpahl J (2001)
Expression of Fas ligand by Microglia:
possible role in glioma immune evasion. J.Neuroimmunol.
120:19-24
Abstract: The immune-privileged status of the central
nervous system is thought to limit the application of immunotherapy
for treatment of malignant brain tumors. Because the Fas pathway has
been proposed to play a role in immune evasion, we examined the
effect of tumor environment
on the expression of Fas ligand (FasL) in a mouse glioma model.
Immunoblotting revealed the expression of membrane-bound FasL to
nearly double when murine G26 gliomas were propagated intracranially
(IC) as compared to subcutaneously (SC). Further analysis by flow
cytometry revealed Microglia,
which were absent in the SC tumors, to account for half of the FasL
expression in the IC tumors. Interestingly, when FasL activity was
inhibited in IC tumors, the proportion of tumor-infiltrating
leukocytes increased three-fold, reaching the same frequency as the
SC tumors. These observations suggest that Microglia
are a major source of FasL expression in brain tumors and possibly
contribute to the local immunosuppressive milieu of malignant
gliomas
Bate C, Reid S, Williams A (2001) Killing of prion-damaged
neurones by Microglia.
Neuroreport 12:2589-2594
Abstract: The loss of neurones that
occurs in the transmissible spongiform encephalopathies, or prion
diseases, can be reproduced in vitro by incubating neuronal cultures
with either peptides derived from the prion protein or with
partially purified prion preparations. In the present studies, the
extent of neuronal loss on exposure to these prions or prion
peptides was increased by the addition of Microglia,
a process that was dependent upon the number of Microglia
added, the concentration of prions/peptides present and the degree
of fibrillarity of the prion peptides. Microglia
also killed scrapie-infected neuroblastoma cells expressing
infectious PrP(SC). Microglia
secreted low amounts of interleukin (IL)-6 when incubated with
peptides alone but up to 10 times as much IL-6 when incubated with
peptide-treated neurones, suggesting that Microglia
recognise peptide-induced changes in neurones
Bu J, Akhtar N, Nishiyama A (2001) Transient expression of
the NG2 proteoglycan by a subpopulation of activated macrophages in
an excitotoxic hippocampal lesion. Glia 34:296-310
Abstract:
Cells that express the NG2 proteoglycan (NG2+ cells) constitute a
large glial population in the normal mature rodent brain. They can
differentiate into oligodendrocytes but are distinct from mature
oligodendrocytes, astrocytes, Microglia,
and neurons. Changes in NG2+ cells were examined in kainic
acid-induced excitotoxic lesions of the hippocampus, and the
relationship between NG2+ cells and reactive astrocytes and
Microglia
was investigated between 1 and 90 days after lesioning. Two types of
reactive NG2+ cells with altered morphology and increased NG2
immunoreactivity were observed in the lesion. Early changes,
consisting of an increase in NG2 immunoreactivity and the number of
processes, were apparent 24 h after lesioning and persisted through
3 months. These cells were distinct from reactive astrocytes or
activated Microglia/macrophages.
A second type of reactive NG2+ cells appeared 2 weeks after
injection, following an influx of macrophages. They had large, round
cell bodies with short processes and expressed the
Microglia/macrophage
antigens OX42 and ED1. Single cells coexpressing NG2 and
macrophage/Microglial
antigens could be isolated from the lesion. The number of NG2+/OX42+
cells gradually declined and disappeared by 3 months after
injection. They did not express glial fibrillary acidic protein or
the alpha receptor for platelet-derived growth factor, indicating
that they are distinct from astrocytes or oligodendrocyte progenitor
cells. Cells that coexpressed NG2 and OX42 were never observed in
hippocampal slice cultures treated with kainic acid, suggesting that
NG2+/OX42+ cells are not derived from endogenous resident brain
cells. These findings demonstrate that NG2 expression is transiently
upregulated on activated macrophages/Microglia
that appear during the chronic stage in an excitotoxic lesion in the
adult CNS
Eales-Reynolds LJ, Laver H, Mojtahedi H (2001) Evidence for
the expression of the EGF receptor on human monocytic cells.
Cytokine 16:169-172
Abstract: Several malignancies over-express
the epidermal growth factor receptor, ligation of which results in
cellular differentiation and multiplication. Mononuclear phagocytes
secrete this cytokine and its receptor has been detected on
Microglial
cells. This communication describes the expression (and its
regulation) of epidermal growth factor receptor (EGFR) on U937
cells. We have shown that a few are EGFR-positive, with expression
being up regulated by interleukin 6 (IL-6). Also, when cultured in
the presence of serum with the monoclonal anti-EGFR, ICR62, U937s
showed a reduced growth rate. By contrast, ICR9 caused a significant
increase in cellular proliferation. Both antibodies induced cycle
arrest in late G(1)/S phase. When the cells were cultured in the
absence of serum, low antibody concentration (10 microg/ml) showed
an early inhibitory effect on cell proliferation. By contrast, at
high antibody concentrations (50 micro/ml), ICR62 significantly
increased the proliferation of U937 cells. We suggest that these
results provide indirect evidence for an autocrine action of EGF on
U937 cells
Elmlinger MW, Deininger MH, Schuett BS, Meyermann R, Duffner
F, Grote EH, Ranke MB (2001) In vivo expression of insulin-like
growth factor-binding protein-2 in human gliomas increases with the
tumor grade. Endocrinology
142:1652-1658
Abstract: Human central nervous system tumors and
glioma cell lines highly express the insulin-like growth
factor-binding protein (IGFBP)-2. As IGFBP-2 can affect tumor
growth, we studied the relationship between IGFBP-2
expression and the malignancy of brain tumors in vivo. To do so, we
investigated by immunohistochemistry the accumulation of IGFBP-1,
-2, and -3 in 50 human gliomas classified by the WHO Malignancy
Scale. Double labeling using anti-CD68 (monocytes/macrophages),
antiglial fibrillary acidic protein, and anti-CD3 (T cells)
antibodies was performed to further characterize the IGFBP-1, -2,
and -3(+) cells. The expression of IGFBP messenger RNAs (mRNAs) was
tested by RT-PCR in tumor samples
from nine gliomas of different grades and in eight cell lines
representing the cellular composition of human glioma. As controls,
the accumulation of IGFBP-2 was investigated in normal brain and in
the rat C6 glioblastoma model. IGFBP-1 and -3 accumulated in
endothelial and macrophage/Microglial
cells. IGFBP-2(+) macrophage/Microglial
and glioma cells clustered in the immediate vicinity of focal
necrosis of the human gliomas as well as of the rat C6 glioblastoma.
The labeling score of IGFBP-1 accumulation in endothelial cells
correlated negatively (P: = 0.0229), and that of IGFBP-2
accumulation in glioma cells correlated positively (P: < 0.0006)
with the tumor grade of the
gliomas. In addition, RT-PCR analysis confirmed mRNA expression of
IGFBP-1, -2, and -3 by the gliomas and glial cells. Small amounts of
IGFBP-1 and -3 mRNA, but high amounts of IGFBP-2 mRNA, were
detectable in macrophage-like and glioma cell lines. The results
suggest cell type-specific accumulation of IGFBP-1, -2, and -3 in
human glial tumors of the brain. The increase in IGFBP-2 expression
with this malignancy suggests a role of IGFBP-2 in the biology of
human gliomas
Fischer FR, Luo Y, Luo M, Santambrogio L, Dorf ME (2001)
RANTES-induced chemokine cascade in dendritic cells. J.Immunol.
167:1637-1643
Abstract: Dendritic cells (DC) are the most potent
APCs and the principal activators of naive T cells. We now report
that chemokines can serve as activating agents for immature DC.
Murine bone marrow-derived DC respond to the CC chemokine RANTES
(10-100 ng/ml) by production of proinflammatory mediators. RANTES
induces rapid expression of transcripts for the CXC chemokines KC
and macrophage inflammatory protein (MIP)-2, the CC chemokines
MIP-1beta and MIP-1alpha, and the cytokines TNF-alpha and IL-6.
Synthesis of KC, IL-6, and TNF-alpha proteins were also
demonstrated. After 4 h, autoinduction of RANTES transcripts was
observed. These responses are chemokine specific. Although DC
demonstrated weak responses to eotaxin, DC failed to respond to
other chemokines including KC, MIP-2, stromal-derived factor-1alpha,
MIP-1beta, MIP-1alpha, monocyte chemoattractant protein-1, T cell
activation gene 3, or thymus-derived chemotactic agent 4. In
addition, RANTES treatment up-regulated expression of an orphan
chemokine receptor termed Eo1. Chemokine induction was also observed
after treatment of splenic DC and neonatal Microglia
with RANTES, but not after treatment of thymocytes or splenocytes
depleted of adherent cells. TNF-alpha-treated DC lose responsiveness
to RANTES. DC from mice deficient for CCR1, CCR3, and CCR5 respond
to RANTES, indicating that none of these receptors are exclusively
used to initiate the chemokine cascade. RANTES-mediated chemokine
amplification in DC may prolong inflammatory responses and shape the
microenvironment, potentially enhancing acquired and innate immune
responses
Fleige G, Nolte C, Synowitz M, Seeberger F, Kettenmann H,
Zimmer C (2001) Magnetic labeling of activated Microglia
in experimental gliomas. Neoplasia. 3:489-499
Abstract:
Microglia,
as intrinsic immunoeffector cells of the central nervous system
(CNS), play a very sensitive, crucial role in the response to almost
any brain pathology where they are activated to a phagocytic state.
Based on the characteristic features of activated Microglia,
we investigated whether these cells can be visualized with magnetic
resonance imaging (MRI) using ultrasmall superparamagnetic iron
oxides (USPIOs). The hypothesis of this study was that MR Microglia
visualization could not only reveal the extent of the tumor, but
also allow for assessing the status of immunologic defense. Using
USPIOs in cell culture experiments and in a rat glioma model, we
showed that Microglia
can be labeled magnetically. Labeled Microglia
are detected by confocal microscopy within and around tumors in a
typical border-like pattern. Quantitative in vitro studies revealed
that Microglia
internalize amounts of USPIOs that are significantly higher than
those incorporated by tumor cells
and astrocytes. Labeled Microglia
can be detected and quantified with MRI in cell phantoms, and the
extent of the tumor can be
seen in glioma-bearing rats in vivo. We conclude that magnetic
labeling of Microglia
provides a potential tool for MRI of gliomas, which reflects tumor
morphology precisely. Furthermore, the results suggest that
MRI may yield functional data on the immunologic reaction of the CNS
Hentze H, Schwoebel F, Lund S, Keel M, Ertel W, Wendel A,
Jaattela M, Leist M, Kehl M (2001) In vivo and in vitro evidence for
extracellular caspase activity released from apoptotic cells.
Biochem.Biophys.Res.Commun. 283:1111-1117
Abstract: While
caspases play an established role as intracellular executors of
apoptosis, little is known about extracellular activities of this
ubiquitously expressed family of proteases. We demonstrate here that
recombinant caspase-3 retained enzymatic activity in various
extracellular fluids. Experiments with cell lines, primary cells,
and mice with fulminant CD95-triggered hepatitis showed that
significant amounts of DEVD-aminofluoromethylcoumarine-cleaving
activity, indicative of active effector caspases, were released into
the medium/plasma during apoptosis. Furthermore, caspase activities
were detected in liquor samples from human head trauma patients.
These findings warrant closer investigation of DEVDase activity as a
diagnostic marker, and of potential extracellular substrates for
caspases
Hodges JL, Ireland DD, Reiss CS (2001) The role of
interleukin-18 in vesicular stomatitis virus infection of the CNS.
Viral Immunol. 14:181-191
Abstract: Intranasal application of
vesicular stomatitis virus (VSV) results in the initial infection of
the olfactory receptor neurons and a rapid progression of the virus
through the mouse central nervous system (CNS). Interleukin-18
(IL-18) is an 18.3-kd cytokine that induces interferon gamma
(IFN-gamma) production in mice. IL-18 is synthesized as an inactive
precursor that is cleaved and activated by
caspase-1/interleukin-1beta converting enzyme (ICE). IL-18 shares
several biological properties with IL-12, including the ability to
induce IFN-gamma production in T lymphocytes and natural killer (NK)
cells. In the CNS, Microglia
and astrocytes produce IL-18 and IL-12. We have previously shown
that IL-12 promotes recovery from VSV encephalitis. This led us to
examine the potential role of IL-18 in the pathogenesis of VSV
encephalitis. We show that both IL-18 and caspase-1 mRNA are
consistently present in the CNS of mice. The addition of exogenous
IL-18 to cell cultures does not affect the production of VSV, and
addition of exogenous IL-18 at the time of infection does not alter
the morbidity or mortality of BALB/c mice. In vitro studies with
neutralizing monoclonal antibody to IL-18 had no effect. From these
results we conclude that in this system and under the experimental
conditions used, unlike IL-12 and IFN-gamma, IL-18 does not play a
significant role in the host response to VSV infection
Hoozemans JJ, Rozemuller AJ, Janssen I, De Groot CJ, Veerhuis
R, Eikelenboom P (2001) Cyclooxygenase expression in Microglia
and neurons in Alzheimer's disease and control brain. Acta
Neuropathol.(Berl) 101:2-8
Abstract: Epidemiological studies
suggest that non-steroidal anti-inflammatory drugs (NSAIDs) lower
the risk of developing Alzheimer's disease (AD). Most NSAIDs act
upon local inflammatory events by inhibiting the expression or
activation of cylooxygenase (COX). In the present study the
expression of COX-1 and COX-2 in AD and non-demented control
temporal and frontal cortex was investigated using
immunohistochemistry. COX-1 expression was detected in Microglial
cells, while COX-2 expression was found in neuronal cells. In AD
brains, COX-1-positive Microglial
cells were primarily associated with amyloid beta plaques, while the
number of COX-2-positive neurons was increased compared to that in
control brains. No COX expression was detected in astrocytes. In
vitro, primary human Microglial
and astrocyte cultures, and human neuroblastoma cells (SK-N-SH) were
found to secrete prostaglandin E2 (PGE2), especially when
stimulated. PGE2 synthesis by astrocytes and SK-N-SH cells was
stimulated by interleukin-1beta. Microglial
cell PGE2 synthesis was stimulated by lipopolysaccharide only.
Although astrocytes are used in studies in vitro to investigate the
role of COX in AD, there are no indications that these cells express
COX-1 or COX-2 in vivo. The different distribution patterns of COX-1
and COX-2 in AD could implicate that these enzymes are involved in
different cellular processes in the pathogenesis of AD
Ito A, Saito S, Masuko T, Oh-Eda M, Matsuura T, Satoh M,
Nejad FM, Enomoto T, Orikasa S, Hakomori SI (2001) Monoclonal
antibody (5F3) defining renal cell carcinoma-associated antigen
disialosyl globopentaosylceramide (V3NeuAcIV6NeuAcGb5), and
distribution pattern of the antigen in tumor
and normal tissues. Glycoconj.J. 18:475-485
Abstract:
Renal cell carcinoma (RCC) has been characterized by high expression
of three types of disialogangliosides: two based on lacto-series
type 1 structure (disialosyl Lc(4), GalNAc disialosyl Lc(4)), the
other based on globo-series structure (disialosyl
globopentaosylceramide; disialosyl Gb5). The present study
established a mAb, 5F3, directed to disialosyl Gb5. 5F3 was
established after immunization with RCC cell line ACHN. The major
disialoganglioside antigen isolated from ACHN cells, showing
specific reactivity with 5F3, was characterized unequivocally as
disialosyl Gb5 (V(3)NeuAcIV(6)NeuAcGb5) by identification of the
core structure as globopentaosylceramide (Gb5) after enzymatic and
acid hydrolysis, and by 2-dimensional (1)H-NMR spectroscopy. 5F3
does not react with monosialosyl Gb5 (V(3)NeuAcGb5), Gb5, or any
lacto-series structures. 5F3 strongly stained 19 of 41 cases of
primary RCC tissue. It reacted with proximal tubules (but not distal
tubules) of kidney, Microglial
cells of cerebrum and cerebellum, goblet cells of stomach and
intestine, smooth muscle of various organs. It did not react with
parenchymatous cells of various organs, except for kidney epithelia
and prostate stroma. Immunostaining of RCC tissue by mAb 5F3, in
combination with staining by other antibodies directed to
globo-series and lacto-series structures, has prognostic
significance in defining metastatic potential of RCC
Keppler OT, Horstkorte R, Pawlita M, Schmidt C, Reutter W
(2001) Biochemical engineering of the N-acyl side chain of sialic
acid: biological implications. Glycobiology 11:11R-18R
Abstract:
N-Acetylneuraminic acid is the most prominent sialic acid in
eukaryotes. The structural diversity of sialic acid is exploited by
viruses, bacteria, and toxins and by the sialoglycoproteins and
sialoglycolipids involved in cell-cell recognition in their highly
specific recognition and binding to cellular receptors. The
physiological precursor of all sialic acids is N-acetyl
D-mannosamine (ManNAc). By recent findings it could be shown that
synthetic N-acyl-modified D-mannosamines can be taken up by cells
and efficiently metabolized to the respective N-acyl-modified
neuraminic acids in vitro and in vivo. Successfully employed
D-mannosamines with modified N-acyl side chains include N-propanoyl-
(ManNProp), N-butanoyl- (ManNBut)-, N-pentanoyl- (ManNPent),
N-hexanoyl- (ManNHex), N-crotonoyl- (ManNCrot), N-levulinoyl-
(ManNLev), N-glycolyl- (ManNGc), and N-azidoacetyl D-mannosamine
(ManNAc-azido). All of these compounds are metabolized by the
promiscuous sialic acid biosynthetic pathway and are incorporated
into cell surface sialoglycoconjugates replacing in a cell
type-specific manner 10-85% of normal sialic acids. Application of
these compounds to different biological systems has revealed
important and unexpected functions of the N-acyl side chain of
sialic acids, including its crucial role for the interaction of
different viruses with their sialylated host cell receptors. Also,
treatment with ManNProp, which contains only one additional
methylene group compared to the physiological precursor ManNAc,
induced proliferation of astrocytes, Microglia,
and peripheral T-lymphocytes. Unique, chemically reactive ketone and
azido groups can be introduced biosynthetically into cell surface
sialoglycans using N-acyl-modified sialic acid precursors, a process
offering a variety of applications including the generation of
artificial cellular receptors for viral gene delivery. This group of
novel sialic acid precursors enabled studies on sialic acid
modifications on the surface of living cells and has improved our
understanding of carbohydrate receptors in their native environment.
The biochemical engineering of the side chain of sialic acid offers
new tools to study its biological relevance and to exploit it as a
tag for therapeutic and diagnostic applications
Le Y, Yazawa H, Gong W, Yu Z, Ferrans VJ, Murphy PM, Wang JM
(2001) The neurotoxic prion peptide fragment PrP(106-126) is a
chemotactic agonist for the G protein-coupled receptor formyl
peptide receptor-like 1. J.Immunol. 166:1448-1451
Abstract: Prion
diseases are transmissible and fatal neurodegenerative disorders
which involve infiltration and activation of mononuclear phagocytes
at the brain lesions. A 20-aa acid fragment of the human cellular
prion protein, PrP(106-126), was reported to mimic the biological
activity of the pathologic isoform of prion and activates
mononuclear phagocytes. The cell surface receptor(s) mediating the
activity of PrP(106-126) is unknown. In this study, we show that
PrP(106-126) is chemotactic for human monocytes through the use of a
G protein-coupled receptor formyl peptide receptor-like 1 (FPRL1),
which has been reported to interact with a diverse array of
exogenous or endogenous ligands. Upon stimulation by PrP(106-126),
FPRL1 underwent a rapid internalization and, furthermore,
PrP(106-126) enhanced monocyte production of proinflammatory
cytokines, which was inhibited by pertussis toxin. Thus, FPRL1 may
act as a "pattern recognition" receptor that interacts
with multiple pathologic agents and may be involved in the
proinflammatory process of prion diseases
Platten M, Wick W, Weller M (2001) Malignant glioma biology:
role for TGF-beta in growth, motility, angiogenesis, and immune
escape. Microsc.Res.Tech. 52:401-410
Abstract: Characteristics of
human malignant glioma are excessive proliferation, infiltrative
growth, angiogenesis and suppression of anti-tumor
immune surveillance. Transforming growth factor-beta
(TGF-beta), a versatile cytokine, is intimately involved in the
regulation of these processes. Here, we discuss the interactions of
TGF-beta with growth factors, such as basic fibroblast growth factor
(bFGF), epidermal growth factor (EGF) and platelet derived growth
factor (PDGF), metalloproteinases (MMP-2, MMP-9) and their
inhibitor, plasmin activator inhibitor-1 (PAI-1), and immune cells,
like natural killer cells, T-cells and Microglia.
The differential effects of TGF-beta in glioma biology are outlined
with emphasis on the induction of a survival advantage for glioma
cells by enforced cell growth, migration, invasion, angiogenesis and
immune paralysis. By virtue of its growth regulatory and
immunomodulatory properties, TGF-beta promises to become a novel
target for the experimental therapy of human malignant glioma
Portis JL (2001) Genetic determinants of neurovirulence of murine oncornaviruses. Adv.Virus Res. 56:3-38
Proescholdt MA, Merrill MJ, Ikejiri B, Walbridge S, Akbasak
A, Jacobson S, Oldfield EH (2001) Site-specific immune response to
implanted gliomas. J.Neurosurg. 95:1012-1019
Abstract: OBJECT:
Immunotherapy for glioblastoma has been uniformly ineffective. The
immunological environment of the brain, with its low expression of
major histocompatibility complex (MHC) molecules and limited access
for inflammatory cells and humoral immune effectors due to the
blood-brain barrier (BBB), may contribute to the failure of
immunotherapy. The authors hypothesize that brain tumors are
protected from immune surveillance by an intact BBB at early stages
of development. To investigate the immunological characteristics of
early tumor growth, the
authors compared the host response to a glioma implanted into the
brain and into subcutaneous tissue. METHODS: Samples of tumors
growing in the brain or subcutaneously in rats were obtained for 7
consecutive days and were examined immunohistochemically for MHC
Class I & II molecules, and for CD4 and CD8 lymphocyte markers.
Additionally, B7-1 costimulatory molecule expression and
lymphocyte-specific apoptosis were examined. CONCLUSIONS: On Days 3
and 4 after implantation, brain tumors displayed significantly lower
MHC Class II expression and lymphocytic infiltration (p < 0.05).
After Day 5, however, no differences were detected. The MHC Class II
expressing cells within the brain tumors appeared to be infiltrating
Microglia.
Minimal B7-1 expression combined with lymphocyte-specific apoptosis
were detected in both brain and subcutaneous tumors. Low MHC Class
II expression and low lymphocytic infiltration at early time points
indicate the importance of the immunologically privileged status of
the brain during early tumor
growth. These characteristics disappeared at later time
points, possibly because the increasing perturbation of the BBB
alters the specific immunological environment of the brain. The lack
of B7-1 expression combined with lymphocyte apoptosis indicates
clonal anergy of glioma-infiltrating lymphocytes regardless of
implantation site
Roth W, Isenmann S, Nakamura M, Platten M, Wick W, Kleihues
P, Bahr M, Ohgaki H, Ashkenazi A, Weller M (2001) Soluble decoy
receptor 3 is expressed by malignant gliomas and suppresses CD95
ligand-induced apoptosis and chemotaxis. Cancer Res.
61:2759-2765
Abstract: Decoy receptor 3 (DcR3) is a newly
identified soluble protein that binds to CD95 ligand (CD95L) and
inhibits its proapoptotic activity. Here we report that DcR3 is
expressed by the majority of long-term and ex vivo malignant glioma
cell lines as well as in human glioblastoma in vivo. Expression of
DcR3 correlates with the grade of malignancy: 15 of 18 (83%)
glioblastomas (WHO grade IV) but none of 11 diffuse astrocytomas
(WHO grade II) exhibited DcR3 immunoreactivity. We also demonstrate
that human malignant glioma cells engineered to release high amounts
of DcR3 into the cell culture supernatant are protected from
CD95L-induced apoptotic cell death. In contrast, DcR3 does not
confer protection from the death ligand Apo2 ligand (TRAIL).
Importantly, ectopic expression of DcR3 resulted in substantial
differences in immune cell infiltration in the 9L rat gliosarcoma
model. Thus, the infiltration of CD4+ and CD8+ T cells as well as
Microglia/macrophages
into glioma was substantially decreased in DcR3-producing tumors
compared with control tumors. Chemotaxis assays revealed that DcR3
counteracts the chemotactic activity of CD95L against Microglial
cells in vitro. These findings suggest that DcR3 may be involved in
the progression and immune evasion of malignant gliomas
Sugibayashi R, Shimizu T, Suzuki T, Yamamoto N, Hamada H,
Takeda K (2001) Upregulation of p21(WAF1/CIP1) leads to morphologic
changes and esterase activity in TPA-mediated differentiation of
human prostate cancer cell line TSU-Pr1. Oncogene
20:1220-1228
Abstract: We reported previously that human prostate
cancer cell line TSU-Pr1 can differentiate into Microglia-like
cells by 12-O-tetra-decanoylphorbol-13-acetate (TPA) treatment. In
this study, we identified a signal transduction pathway involved in
TPA-induced TSU-Pr1 cell differentiation and investigated the
mechanism of growth arrest that accompanies this differentiation.
TPA-induced differentiation and growth arrest of TSU-Pr1 cells were
inhibited by treatment with Protein kinase C (PKC) inhibitor
GF109203X and mitogen-activated protein (MAP) kinase inhibitor
PD98059. Treatment of TSU-Pr1 cells with TPA for 15 min or longer
resulted in translocation of PKCalpha, PKCgamma, and PKCepsilon from
cytosolic to membrane fraction. Our results suggest that TPA-induced
TSU-Pr1 cell differentiation is associated with activation of MAP
kinase and PKCalpha, PKCgamma, and PKCepsilon. The mechanism of
growth arrest in TSU-Pr1 cells that underwent TPA-induced
differentiation were examined for factors in the signaling pathway
downstream of MAP kinase that control the cell cycle. Upregulation
of p21(WAF1/CIP1) cyclin-dependent kinase inhibitor protein was
observed in a manner dependent on PKC or MAP kinase. Moreover,
adenovirus-mediated overexpression of recombinant p21(WAF1/CIP1) in
TSU-Pr1 cells result in growth arrest, morphological change to
Microglia-like
cells, and increased alpha-naphthyl acetate esterase activity, all
of which are associated with cellular differentiation. Thus, our
results indicate that p21(WAF1/CIP1) mediates TPA-induced growth
arrest and differentiation of TSU-Pr1 cells
Synowitz M, Ahmann P, Matyash M, Kuhn SA, Hofmann B, Zimmer
C, Kirchhoff F, Kiwit JC, Kettenmann H (2001) GABA(A)-receptor
expression in glioma cells is triggered by contact with neuronal
cells. Eur.J.Neurosci. 14:1294-1302
Abstract: The expression of
functional GABA(A)-receptors in glioma cells correlates with low
malignancy of tumours and cell lines from glioma lack these
receptors. Here we show that contact with neurons induces the
expression of functional GABA(A)-receptors. C6 and F98 glioma cell
lines were labelled by recombinant expression of enhanced green
fluorescent protein injected into rat brain and studied in acute
slices after two to three weeks of tumour growth. The cells
responded to GABA or the specific agonist, muscimol with a current
typical for GABA(A)-receptors, as studied with the patch-clamp
technique. To get insight into the mechanism of GABA(A) receptor
induction, the C6 or F98 cells were co-cultured with neurons,
astrocytes, oligodendrocytes and Microglia.
Glioma cells expressed functional GABA(A) receptors within 24 h only
in cultures where physical contact to neurons occurred. Activation
of GABA(A)-receptors in the co-cultures attenuated glioma cell
metabolism while blockade of the receptors increased metabolism. We
conclude that with this form of interaction, neurons can influence
tumour behaviour in the brain
Carpentier AF, Xie J, Mokhtari K, Delattre JY (2000)
Successful treatment of intracranial gliomas in rat by
oligodeoxynucleotides containing CpG motifs. Clin.Cancer Res.
6:2469-2473
Abstract: Phosphorothioate oligodeoxynucleotides with
CpG motifs (CpG-ODNs) activate various immune cell subsets and
induce production of numerous cytokines. To evaluate whether
CpG-ODNs can induce rejection of established tumors, Lewis rats were
inoculated intracerebrally with syngeneic CNS-1 glioma cells and
subsequently injected with CpG-ODNs into the tumor
bed. Although all of the control rats (n = 14) died within 23
days, 88% of the animals (n = 8) treated with a single CpG-ODN
injection 5 days after tumor
inoculation showed long-term survival (>90 days; P <
0.002). CpG-ODNs increased tumoral infiltration with
macrophage/Microglial
cells, CD8, and natural killer lymphocytes. CpG-ODN-cured animals
were further protected against a second tumor
challenge. CpG-ODNs had no effect on a s.c. CNS1 tumor
in nude mice, which suggested that CpG-ODN is not directly
cytotoxic and that immunostimulation is required for the antitumoral
effect. These findings suggest that intratumoral injections of
CpG-ODNs represent a new immunotherapeutic approach in human
gliomas, which overcome the need for the selection and purification
of a tumoral antigen
Cohen J, Sugita Y, Chader GJ, Schwartz JP (2000) Recombinant
forms of the neurotrophic factor pigment epithelium-derived factor
activate cellular metabolism and inhibit proliferation of the RAW
macrophage cell line. Neuroimmunomodulation. 7:51-58
Abstract:
Recombinant forms of the neurotrophic factor pigment
epithelium-derived factor (PEDF) activate metabolism of RAW
macrophage cells while simultaneously inhibiting their
proliferation. The recombinant forms (rPEDF) acted with EC(50)s of
0.1-1 nM while full-length native bovine PEDF was inactive. Urea,
which is the buffer used to extract recombinant PEDF, stimulated RAW
cell proliferation, the first report of an effect of urea on
non-kidney cells. PEDF acted within 12 h and its effects persisted
up to 72 h with continuous exposure. Although rPEDF had no direct
action on glioma cell lines, it increased the amount of a soluble
factor released by RAW cells which was capable of blocking glioma
cell division. Thus PEDF may function as a neuroimmune modulator,
affecting both neural and immune system cells
Dallasta LM, Wang G, Bodnar RJ, Draviam R, Wiley CA, Achim
CL, Hamilton RL (2000) Differential expression of intercellular
adhesion molecule-1 and vascular cell adhesion molecule-1 in chronic
murine retroviral encephalitis. Neuropathol.Appl.Neurobiol.
26:332-341
Abstract: The cell adhesion molecules, intercellular
adhesion molecule (ICAM)-1 and vascular cell adhesion molecule
(VCAM)-1, are important mediators of immune interactions within the
central nervous system (CNS). A wide variety of pro-inflammatory
insults to the brain, including viral infection, result in
upregulation of these molecules on brain endothelial cells,
astrocytes, and Microglia.
This study investigated the expression of ICAM-1 and VCAM-1 in
chronic encephalitis induced by infection with a temperature
sensitive (ts-1) strain of Moloney murine leukaemia virus (MoMuLV),
an ecotropic murine retrovirus. During the late stages of disease,
viral antigen was present in both endothelial cells and Microglia,
but not astrocytes, in regions of spongiform change and gliosis. In
these areas, ICAM-1 staining was detected on activated Microglia,
but not on endothelial cells or astrocytes. In contrast, no cells
showed increased VCAM-1 expression in the CNS. These findings
demonstrate that there is cell-specific, differential expression of
these adhesion molecules in ts-1 retroviral encephalitis. The lack
of endothelial cell expression correlates with the characteristic
lack of lymphocytic infiltrate in this chronic retroviral
encephalitis and suggests that increased Microglial
ICAM-1 expression may play a role in the pathogenesis of MoMuLV
(ts-1)-mediated neurodegeneration
Deininger MH, Seid K, Engel S, Meyermann R, Schluesener HJ
(2000) Allograft inflammatory factor-1 defines a distinct subset of
infiltrating macrophages/Microglial
cells in rat and human gliomas. Acta Neuropathol.(Berl)
100:673-680
Abstract: Allograft inflammatory factor-1 (AIF-1) is
a Ca2+-binding peptide that constitutes a potential modulator of
macrophage activation and function during the immune response of the
brain. Peptides termed Microglia
response factor-1 or ionized calcium-binding adaptor molecule- have
been reported to be identical with AIF-1. We have investigated the
expression of AIF-1 in the rat C6 glioblastoma and 9L gliosarcoma
tumor models and
additionally assessed AIF- expression in a diverse range of human
astrocytomas by immunohistochemistry. AIF-1 was expressed by
activated Microglial
cells and a subset of infiltrating macrophages in areas of
infiltrative tumor growth
and in compact tumor areas
in both rat and human gliomas. Double-labeling experiments in rats
and humans characterized the nature and the functional status of
AIF-1+ cells. AIF-1 expression was detected in cells expressing
major histocompatibility complex class II molecules and in a subset
of activated macrophages/Microglial
cells. All MRP-8+ cells coexpressed AIF-1. In humans, there was a
strong correlation of AIF-1-expressing activated
macrophages/Microglial
cells with tumor malignancy
(P < 0.0001). These results suggest that AIF-1 defines a distinct
subset of tumor-associated activated macrophages/ Microglial
cells
Deininger MH, Meyermann R, Trautmann K, Duffner F, Grote EH,
Wickboldt J, Schluesener HJ (2000) Heme oxygenase (HO)-1 expressing
macrophages/Microglial
cells accumulate during oligodendroglioma progression. Brain Res.
882:1-8
Abstract: Heme oxygenase (HO-1, HSP32) catalyzes the
oxidation of heme to biliverdin and carbon monoxide, a putative
neurotransmitter. In the brain, HO-1 expression has been associated
with neuroprotection during oxidative stress and hypoxia. However,
consecutive downstream mediation is involved in neoangiogenesis and
consequent neoplastic outgrowth. We have analyzed HO-1 expression in
69 oligodendroglioma tissue samples, in rat intracranially
transplanted C6 gliomas, and neuropathologically unaltered control
brains by immunohistochemistry. Double labeling experiments
confirmed the nature of HO-1 expressing cells. Reverse
transcription-polymerase chain reaction was used to demonstrate HO-1
gene expression. HO-1 immunoreactivity was predominantly observed in
macrophages/Microglial
cells. The number of HO-1 expressing macrophages/Microglial
cells was significantly lower in primary oligodendrogliomas than in
their matched relapses (P<0.0001) and lower in primary anaplastic
oligodendrogliomas than in their relapses (P=0.0006). Prominent
accumulation of HO-1 expressing macrophages/Microglial
cells was observed in perinecrotic areas of both experimental rat
and human glioblastoma relapses. HO-1 expressing neurons,
macrophages/Microglial
cells and astrocytes were scattered in areas of infiltrative tumor
growth. Surprisingly, HO-1 mRNA was detected in only one
glioblastoma multiforme relapse. We conclude from these data that
HO-1 expressing macrophages/Microglial
cells accumulate during oligodendroglioma progression in areas of
focal necrosis. However, overall biological function of this
phenomenon remains to be determined
Kingham PJ, Pocock JM (2000) Microglial
apoptosis induced by chromogranin A is mediated by mitochondrial
depolarisation and the permeability transition but not by cytochrome
c release. J.Neurochem. 74:1452-1462
Abstract: Chromogranin A is
up-regulated in the senile plaques of Alzheimer's brain and is a
novel activator of Microglia,
transforming them to a neurotoxic phenotype. Treatment of primary
cultures of rat brain Microglia
or the murine N9 Microglial
cell line with chromogranin A resulted in nitric oxide production,
which triggered Microglial
apoptosis. Exposure of Microglia
to chromogranin A resulted in a fall in mitochondrial membrane
potential. Mitochondrial depolarisation and apoptosis were reduced
significantly by cyclosporin A, but not by the calcineurin inhibitor
FK506. Cytochrome c did not translocate from the mitochondria to the
cytosol, but its expression became significantly enhanced within the
mitochondria. Inhibition of caspase 1 attenuated chromogranin
A-induced Microglial
apoptosis, but did not prevent mitochondrial depolarisation,
indicating that apoptosis occurred downstream of mitochondrial
depolarisation. Conversely, staurosporine-induced Microglial
apoptosis led to mitochondrial cytochrome c release, but not caspase
1 activation. Our findings provide insight into the pathways
controlling activation-triggered Microglial
apoptosis and may point to routes for the modulation of Microglial
evoked neurotoxicity
Prat E, Baron P, Meda L, Scarpini E, Galimberti D, Ardolino
G, Catania A, Scarlato G (2000) The human astrocytoma cell line
U373MG produces monocyte chemotactic protein (MCP)-1 upon
stimulation with beta-amyloid protein. Neurosci.Lett.
283:177-180
Abstract: Astrocytes associated with beta-amyloid
(Abeta) accumulate in senile plaques of Alzheimer's disease (AD). To
investigate the biological effects of Abeta/astrocyte interaction,
we examined chemokine production by the human astrocytoma cell line
U373MG stimulated with Abeta peptides. Northern blot analysis and
specific immunoassays demonstrate that Abeta [1-42] and Abeta
[25-35] induce mRNA expression and release of monocyte chemotactic
protein (MCP)-1 but not of gamma-interferon inducible protein
(IP)-10 by U373MG cells. The observation that Abeta induces
astrocyte production of the potent Microglia
chemoattractant MCP-1 contributes to understanding mechanism of
damage exerted by Abeta in AD senile plaques
Sandmair AM, Turunen M, Tyynela K, Loimas S, Vainio P,
Vanninen R, Vapalahti M, Bjerkvig R, Janne J, Yla-Herttuala S (2000)
Herpes simplex virus thymidine kinase gene therapy in experimental
rat BT4C glioma model: effect of the percentage of thymidine
kinase-positive glioma cells on treatment effect, survival time, and
tissue reactions. Cancer Gene Ther. 7:413-421
Abstract: Herpes
simplex virus thymidine kinase (HSV-tk) gene transfer and
ganciclovir (GCV) administration have been suggested for the
treatment of malignant gliomas. To understand tissue responses and
possible ways to improve the treatment effect, we studied tumor
growth, tissue reactions, and survival time after HSV-tk/GCV
treatment in a syngeneic BT4C rat glioma model by mixing various
ratios of stably transfected HSV-tk-expressing BT4C-tk glioma cells
with wild-type BT4C glioma cells (percentage of BT4C-tk cells: 0%,
1%, 10%, 30%, 50%, and 100%), followed by injection into BDIX rat
brains (n = 79). With the exception of some animals with end-stage
tumors, very little astroglia or Microglia
reactivity was detected in the wild-type tumors as analyzed by
immunocytochemistry using glial fibrillary acid protein (GFAP)-,
vimentin-, human histocompatibility leukocyte antigen-DR-, OX-42-,
and CD68-specific monoclonal antibodies. After 14 days of GCV
treatment, tumors induced with > or = 10% BT4C-tk cells showed a
significant reduction in tumor
size (P < .05) and prolonged survival time (P < .01).
Astrogliosis, as indicated by a strong GFAP and vimentin
immunoreactivity, was seen in the tumor
scar area. GFAP and vimentin reactivity was already present
after the GCV treatment in tumors induced with 1% BT4C-tk cells.
Much less human histocompatibility leukocyte antigen-DR-positive
Microglia
was seen in the treated animals, indicating low Microglia
reactivity and immunoactivation against the tumor. However,
GCV-treated tumors were positive for apoptosis, indicating that
apoptosis is an important mechanism for cell death in the BT4C-tk
glioma model. Our results suggest that > or = 10% transfection
efficiency is required for a successful reduction in BT4C glioma
tumor size with HSV-tk/GCV
treatment in vivo. Tissue reactions after 14 days of GCV treatment
are characterized by astrogliosis and apoptosis, whereas Microglia
response and immunoactivation of the brain cells appear to play a
minor role. Stimulation of the Microglia
response by gene transfer or other means might improve the efficacy
of the HSV-tk/GCV treatment in vivo
Satoh J, Kuroda Y (2000) Amyloid precursor protein
beta-secretase (BACE) mRNA expression in human neural cell lines
following induction of neuronal differentiation and exposure to
cytokines and growth factors. Neuropathology. 20:289-296
Abstract:
Recently, a novel amyloid precursor protein beta-secretase
(designated BACE) was identified. Because activated Microglia
and astrocytes play a role in amyloidogenesis in Alzheimer's
disease, the constitutive and glial cytokine/growth factor-regulated
expression of BACE was studied in human neural cell lines. By
reverse transcription-polymerase chain reaction (RT-PCR) analysis,
BACE mRNA expression was identified in various human neural and
non-neural cell lines. By northern blot analysis, the expression of
BACE mRNA composed of five distinct transcripts (>8.0, 7.0, 6.0,
4.4 and 2.6 kb) was elevated markedly in NTera2 teratocarcinoma
cells following retinoic acid-induced neuronal differentiation. But
the levels of three major BACE mRNA species (7.0, 6.0 and 4.4 kb)
were not significantly altered in NTera2-derived neurons, SK-N-SH
neuroblastoma or U-373MG astrocytoma following exposure to tumor
necrosis factor-alpha, interleukin (IL)-1beta, IL-6,
interferon-gamma, transforming growth factor-beta1, epidermal growth
factor, basic fibroblast growth factor, brain-derived neurotrophic
factor, dibutyryl cyclic adenosine monophosphate or phorbol
12-myristate 13-acetate. These results indicate that BACE mRNA is
expressed constitutively in human neural cells and its expression is
upregulated during neuronal differentiation, but it is unlikely to
be regulated by activated glia-derived cytokines and growth factors
Satoh J, Kuroda Y (2000) Beta-catenin expression in human
neural cell lines following exposure to cytokines and growth
factors. Neuropathology. 20:113-123
Abstract: Beta-catenin acts
as a key mediator of the Wnt/Wingless signaling pathway involved in
cell proliferation, differentiation and survival. Recent studies
have shown that an unstable interaction between beta-catenin and the
mutant presenilin-1 induces neuronal apoptosis, and that
beta-catenin levels are decreased in the brains of patients with
Alzheimer's disease (AD). Since activated Microglia
and astrocytes play a role in the process of neuronal degeneration
in AD, the cytokine/growth factor-regulated expression of
beta-catenin in human neural cell lines, including NTera2
teratocarcinoma-derived differentiated neurons (NTera2-N), IMR-32
neuroblastoma, SKN-SH neuroblastoma and U-373MG astrocytoma, was
studied quantitatively following exposure to epidermal growth factor
(EGF), basic fibroblast growth factor (bFGF), brain-derived
neurotrophic factor (BDNF), tumor
necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta,
IL-6, interferon (IFN)-gamma, transforming growth factor
(TGF)-beta1, dibutyryl cyclic adenosine 3',5'-cyclic monophosphate
(cAMP) (dbcAMP) or phorbol 12-myristate 13-acetate (PMA).
Beta-catenin mRNA expressed constitutively in all of these cell
lines was unaffected by treatment with any factors examined. In
contrast, beta-catenin protein levels were reduced markedly in
NTera2-N cells by exposure to dbcAMP, EGF or bFGF, and in U-373MG
cells by treatment with dbcAMP or PMA, but were unaffected in any
cell lines by BDNF, TNF-alpha, IL-1beta, IL-6, IFN-gamma or
TGF-beta1. These results indicate that beta-catenin is expressed
constitutively in human neural cells and downregulated at a protein
level by a set of growth factors in a cell type-specific manner
Shieh JT, Martin J, Baltuch G, Malim MH, Gonzalez-Scarano F
(2000) Determinants of syncytium formation in Microglia
by human immunodeficiency virus type 1: role of the V1/V2 domains.
J.Virol. 74:693-701
Abstract: Microglia
are the main reservoir for human immunodeficiency virus type 1
(HIV-1) in the central nervous system (CNS), and multinucleated
giant cells, the result of fusion of HIV-1-infected Microglia
and brain macrophages, are the neuropathologic hallmark of HIV
dementia. One potential explanation for the formation of syncytia is
viral adaptation for these CD4(+) CNS cells. HIV-1(BORI-15), a virus
adapted to growth in Microglia
by sequential passage in vitro, mediates high levels of fusion and
replicates more efficiently in Microglia
and monocyte-derived-macrophages than its unpassaged parent (J. M.
Strizki, A. V. Albright, H. Sheng, M. O'Connor, L. Perrin, and F.
Gonzalez-Scarano, J. Virol. 70:7654-7662, 1996). Since the
interaction between the viral envelope glycoprotein and CD4 and the
chemokine receptor mediates fusion and plays a key role in tropism,
we have analyzed the HIV-1(BORI-15) env as a fusogen and in
recombinant and pseudotyped viruses. Its syncytium-forming phenotype
is not the result of a switch in coreceptor use but rather of the
HIV-1(BORI-15) envelope-mediated fusion of CD4(+)CCR5(+) cells with
greater efficiency than that of its parental strain, either by
itself or in the context of a recombinant virus. Genetic analysis
indicated that the syncytium-forming phenotype was due to four
discrete amino acid differences in V1/V2, with a single-amino-acid
change between the parent and the adapted virus (E153G) responsible
for the majority of the effect. Additionally, HIV-1(BORI-15)
env-pseudotyped viruses were less sensitive to decreases in the
levels of CD4 on transfected 293T cells, leading to the hypothesis
that the differences in V1/V2 alter the interaction between this
envelope and CD4 or CCR5, or both. In sum, the characterization of
the envelope of HIV-1(BORI-15), a highly fusogenic glycoprotein with
genetic determinants in V1/V2, may lead to a better understanding of
the relationship between HIV replication and syncytium formation in
the CNS and of the importance of this region of gp120 in the
interaction with CD4 and CCR5
Taniguchi Y, Ono K, Yoshida S, Tanaka R (2000)
Antigen-presenting capability of glial cells under glioma-harboring
conditions and the effect of glioma-derived factors on antigen
presentation. J.Neuroimmunol. 111:177-185
Abstract: The
antigen-presenting capability of syngeneic rat glial cells was
investigated under glioma-harboring conditions. Microglia
induced a significant proliferation of glioma-primed splenocytes,
but astrocytes did not. Furthermore, astrocytes suppressed the
accessory cell function of Microglia.
The presence of both indomethacin and anti-interleukin (IL)-10
neutralizing antibody during priming of Microglia
enhanced splenocyte proliferation. The glioma culture supernatants
down-regulated the interferon-gamma-induced expression of major
histocompatibility complex class II molecules on Microglia.
The down-regulation was blocked by indomethacin and anti-IL-10
antibody. The results suggest that Microglia
but not astrocytes may function as antigen-presenting cells in
glioma, and that glioma may suppress the antigen-presenting
abilities of Microglia
Thomas A, Gasque P, Vaudry D, Gonzalez B, Fontaine M (2000)
Expression of a complete and functional complement system by human
neuronal cells in vitro. Int.Immunol. 12:1015-1023
Abstract: We
demonstrate in vitro expression of complement components, i.e. C3,
factor H (FH), factor B (FB), C4, C1-inhibitor (C1-inh), C1q, C5,
C6, C7 and C9, by four human neuroblastoma cell lines IMR32, SKNSH,
SH-SY5Y and KELLY. Activating proteins C4, C9 and C1q, and
regulatory proteins FH and C1-inh were produced constitutively by
the four cell lines. C3, C6 and FB were mainly produced by SKNSH and
SH-SY5Y. Western blot experiments showed that secreted proteins were
structurally similar to their serum counterparts. An additional
polypeptide of 43 kDa with FH immunoreactivity was detected, which
could correspond to the N-terminal truncated form found in plasma.
Regulation of complement expression by inflammatory cytokines,
lipopolysaccharide and dexamethasone was tested in vitro. These
factors had no significant effects on activating synthesis of
components C3, FB and C4, but expression of regulating components
C1-inh and FH was strongly increased particularly by IFN-gamma and
tumor necrosis
factor-alpha. The rate of synthesis of complement components was
dependent on the differentiation of neuroblastoma cells. This effect
of differentiation was also observed on normal rat neurons. Rat
cerebellar granule cells constitutively expressed mRNA for C4 and
C1q, but expression of C3 mRNA was induced by differentiation. This
study shows that neurons could be another local source of complement
in the brain, besides astrocytes and Microglia.
Human neuroblastoma cell lines can constitute an interesting model
to analyze complement biosynthesis by human neurons. Local
complement expression by neurons in vivo may be implicated in some
physio-pathological processes
Zhou X, Espey MG, Chen JX, Hofseth LJ, Miranda KM, Hussain
SP, Wink DA, Harris CC (2000) Inhibitory effects of nitric oxide and
nitrosative stress on dopamine-beta-hydroxylase. J.Biol.Chem.
275:21241-21246
Abstract: Dopamine-beta-hydroxylase (DbetaH) is a
copper-containing enzyme that uses molecular oxygen and ascorbate to
catalyze the addition of a hydroxyl group on the beta-carbon of
dopamine to form norepinephrine. While norepinephrine causes
vasoconstriction following reflex sympathetic stimulation, nitric
oxide (NO) formation results in vasodilatation via a guanylyl
cyclase-dependent mechanism. In this report, we investigated the
relationship between NO and DbetaH enzymatic activity. In the
initial in vitro experiments, the activity of purified DbetaH was
inhibited by the NO donor, diethylamine/NO (DEA/NO), with an IC(50)
of 1 mm. The inclusion of either azide or GSH partially restored
DbetaH activity, suggesting the involvement of the reactive nitrogen
oxide species, N(2)O(3). Treatment of human neuroblastoma cells
(SK-N-MC) with diethylamine/NO decreased cellular DbetaH activity
without affecting their growth rate and was augmented by the
depletion of intracellular GSH. Co-culture of the SK-N-MC cells with
interferon-gamma and lipopolysaccharide-activated macrophages, which
release NO, also reduced the DbetaH activity in the neuroblastoma
cells. Our results are consistent with the hypothesis that
nitrosative stress, mediated by N(2)O(3), can result in the
inhibition of norepinephrine biosynthesis and may contribute to the
regulation of neurotransmission and vasodilatation
Arbour N, Ekande S, Cote G, Lachance C, Chagnon F, Tardieu M,
Cashman NR, Talbot PJ (1999) Persistent infection of human
oligodendrocytic and neuroglial cell lines by human coronavirus
229E. J.Virol. 73:3326-3337
Abstract: Human coronaviruses (HuCV)
cause common colds. Previous reports suggest that these infectious
agents may be neurotropic in humans, as they are for some mammals.
With the long-term aim of providing experimental evidence for the
neurotropism of HuCV and the establishment of persistent infections
in the nervous system, we have evaluated the susceptibility of
various human neural cell lines to acute and persistent infection by
HuCV-229E. Viral antigen, infectious virus progeny and viral RNA
were monitored during both acute and persistent infections. The
astrocytoma cell lines U-87 MG, U-373 MG, and GL-15, as well as
neuroblastoma SK-N-SH, neuroglioma H4, and oligodendrocytic MO3.13
cell lines, were all susceptible to an acute infection by HuCV-229E.
The CHME-5 immortalized fetal Microglial
cell line was not susceptible to infection by this virus. The MO3.13
and H4 cell lines also sustained a persistent viral infection, as
monitored by detection of viral antigen and infectious virus
progeny. Sequencing of the S1 gene from viral RNA after
approximately 130 days of infection showed two point mutations,
suggesting amino acid changes during persistent infection of MO3.13
cells but none for H4 cells. Thus, persistent in vitro infection did
not generate important changes in the S1 portion of the viral spike
protein, which was shown for murine coronaviruses to bear
hypervariable domains and to interact with cellular receptor. These
results are consistent with the potential persistence of HuCV-229E
in cells of the human nervous system, such as oligodendrocytes and
possibly neurons, and the virus's apparent genomic stability
Badie B, Schartner J, Klaver J, Vorpahl J (1999) In vitro
modulation of Microglia
motility by glioma cells is mediated by hepatocyte growth
factor/scatter factor. Neurosurgery 44:1077-1082
Abstract:
OBJECTIVE: Considered as immune effector cells of the central
nervous system, Microglia
represent a major component of the inflammatory cells found in
malignant gliomas. Although their role in brain tumor
biology is unclear, accumulation of Microglia
in malignant brain tumors may be mediated through active secretion
of cytokines by glioma cells. Because hepatocyte growth
factor/scatter factor (HGF/SF) has been shown to modulate glioma
motility through an autocrine mechanism, and because Microglia
have been reported to express the HGF/SF receptor Met, we
hypothesized that Microglia
recruitment by gliomas may also occur through the secretion of
HGF/SF. METHODS: The effect of glioma cells in augmenting BV-2
murine Microglia
motility was studied by using an in vitro Boyden chamber migration
assay. To determine the chemokines involved in Microglia
migration, neutralizing monoclonal antibodies against monocyte
chemotactic protein-1 and HGF/SF were tested. Immunoblotting was
used to check for the expression of HGF/SF by glioma cells, and the
expression of Met by BV-2 cells was examined by flow cytometry.
RESULTS: BV-2 migration was noted within 7 hours of incubation with
both human (U251 MG and U373 MG) and murine (GL261) glioma cell
lines. This migration corresponded to HGF/SF secretion by glioma
cells and was completely inhibited by neutralizing monoclonal
antibody against HGF/SF, but not monocyte chemotactic protein-1.
Exposure of BV-2 cells to recombinant HGF/SF, but not monocyte
chemotactic protein-1, resulted in their migration and
down-regulation of Met in a dose-dependent fashion. CONCLUSION:
HGF/SF, which plays a role in glioma motility and mitogenesis, may
also act as a chemokine for Microglia
and may be responsible for the Microglia
infiltration in malignant gliomas. This active recruitment of
Microglia
may play an important role in glioma biology
Chattopadhyay N, Ye CP, Yamaguchi T, Kerner R, Vassilev PM,
Brown EM (1999) Extracellular calcium-sensing receptor induces
cellular proliferation and activation of a nonselective cation
channel in U373 human astrocytoma cells. Brain Res.
851:116-124
Abstract: A receptor for extracellular calcium ions
(Ca2+o), cloned from parathyroid gland, serves a critical function
in Ca2+o homeostasis by regulating PTH release via "sensing"
of its physiological agonist, Ca2+o. Its cloning from rat striatum
revealed that the extracellular calcium-sensing receptor (CaR) could
be involved in sensing ambient Ca2+o within the brain, where Ca2+
plays key roles in virtually all aspects of central nervous system
(CNS) function. The CaR is expressed in neurons, oligodendrocytes,
Microglia
and the human astrocytoma cell line, U87 where its functions include
control of cellular proliferation and modulation of ion channels,
such as outward K+ channels and nonselective cation channels (NCC).
In this report, we have shown that the CaR is expressed in U373
cells as assessed by RT-PCR using CaR-specific primers followed by
sequencing of the amplified products, by Northern blot analysis
using a CaR-specific probe as well as by Western analysis utilizing
a specific polyclonal anti-CaR antiserum. Furthermore, agents known
to activate the cloned CaR induce increases in cellular
proliferation and the open probability of an NCC. Thus our study
strongly suggests that elevated levels of Ca2+o, acting via the CaR,
activate an NCC that could contribute to the associated CaR-induced
stimulation of proliferation
Czub M, Czub S, Gosztonyi G, Koutsilieri E, Sopper S, Muller
JG, Gerlach M, Riederer P, ter M, V (1999) Effects of Selegiline in
a retroviral rat model for neurodegenerative disease. J.Neurovirol.
5:458-464
Abstract: Upon inoculation into neonatal rats, murine
leukemia virus (MuLV) NT40 causes a non-inflammatory degeneration of
the central nervous system. While Microglia
cells appear to be the major target cells within the brain
parenchyma for neurovirulent MuLV, degenerating neurons do not
express retroviral gene products. In order to protect rats from
neuronal damage we treated retrovirally infected rats once with
monoamine oxidase (MAO) B inhibitor Selegiline which--under
different conditions--exerts neuroprotective effects. Unexpectedly,
when administered at 17 days post-infection (d.p.i.) a single
intraperitoneal dose of Selegilin (1 mg/kg bodyweight) significantly
shortened the incubation period for neurological disease. In
contrast, Selegiline given in a lower dosage (0.05 mg/kg bodyweight)
and/or at a different time point (13 d.p.i.) at the low (0.05 mg/kg
bodyweight) and the high dose (1.0 mg/kg bodyweight) had no effect
on the outcome of neurological disease. Animals treated with
Selegiline (1.0 mg/kg bodyweight at 17 d.p.i.) contained higher
amounts of viral loads in the CNS, higher numbers of brain cells
expressing major histocompatibility complex class II molecules, and
exhibited inhibition of MAO-B in comparison to untreated yet
infected (control) animals. Supposedly, Selegiline activated the
major target cell population of the CNS for MuLV-NT40, Microglia,
with the consequence of enhanced susceptibility for retroviral
infection and triggered endogenous mechanism(s) involved in the
pathogenesis of retroviral neurodegeneration
Davoust N, Jones J, Stahel PF, Ames RS, Barnum SR (1999)
Receptor for the C3a anaphylatoxin is expressed by neurons and glial
cells. Glia 26:201-211
Abstract: Little is known about the
expression of the receptor for complement anaphylatoxin C3a (C3aR)
in the central nervous system (CNS). In this study, we provide the
first evidence that neurons are the predominant cell type expressing
C3aR in the normal CNS. By using in situ hybridization (ISH) and
immunohistochemistry, we found that C3aR is constitutively expressed
at high levels in cortical and hippocampal neurons as well as in
Purkinje cells. Moreover, we showed that primary culture of human
astrocytes and Microglia
express the C3aR mRNA as assessed by RT-PCR. In situ hybridization
performed on rat primary astrocytes confirmed the RT-PCR result
demonstrating C3aR expression by astrocytes. In experimental
allergic encephalitis (EAE), C3aR expression was elevated on
Microglia,
infiltrating monocyte-macrophage cells and a few astrocytes, whereas
neuronal expression remained unchanged during the course of the
disease. These data demonstrate that the C3aR is expressed primarily
by neurons in the normal CNS and that its neuronal expression is not
dramatically upregulated under inflammation. This is in contrast to
the increased neuronal expression of the C5aR in several
inflammatory CNS conditions. The high constitutive expression of the
C3aR by neurons suggests this receptor may play an important role in
normal physiological conditions in the CNS
Deininger MH, Schluesener HJ (1999) Cyclooxygenases-1 and -2
are differentially localized to Microglia
and endothelium in rat EAE and glioma. J.Neuroimmunol.
95:202-208
Abstract: Cyclooxygenases (COX) mediate a wide variety
of derangements observed during diseases of the brain. Their
overexpression is involved in the mediation of inflammation,
immunomodulation, blood flow, apoptosis and fever. Here, we have
analyzed the localization of COX-1 and COX-2 in rat experimental
autoimmune encephalomyelitis (EAE), C6 glioblastoma and 9L
gliosarcoma by immunohistochemistry. In healthy brain, COX-1 was
expressed in single macrophages/Microglial
cells. Neurons and few endothelial cells expressed COX-2. In EAE, we
observed an increase in COX-1+ macrophages/Microglial
cells and COX-2+ endothelial cells that was closely linked to
disease progression. Both COX-1+ macrophages/Microglial
cells and COX-2+ endothelial cells were abundant in areas of
cellular infiltration. In C6 and 9L tumors, high numbers of COX-1+
macrophages/Microglial
cells and COX-2+ endothelial cells were found both in the tumor
parenchyma and in areas of infiltrative tumor
growth. Double labeling experiments confirmed expression of
COX-2 in vWF+ (endothelial) cells and COX-1 in ED1+ (macophages),
OX6+ (MHC class II) and in W3/13+ (lymphoblasts) cells. These data
provide further evidence that expression of COX-1 in
macrophages/Microglial
cells and COX-2 in endothelial cells might represent important
regulatory mechanisms in inflammatory processes associated with
autoimmunity and neoplasia of the rat brain
Dewey RA, Morrissey G, Cowsill CM, Stone D, Bolognani F, Dodd
NJ, Southgate TD, Klatzmann D, Lassmann H, Castro MG, Lowenstein PR
(1999) Chronic brain inflammation and persistent herpes simplex
virus 1 thymidine kinase expression in survivors of syngeneic glioma
treated by adenovirus-mediated gene therapy: implications for
clinical trials. Nat.Med. 5:1256-1263
Abstract: The long-term
consequences of adenovirus-mediated conditional cytotoxic gene
therapy for gliomas remain uncharacterized. We report here detection
of active brain inflammation 3 months after successful inhibition of
syngeneic glioma growth. The inflammatory infiltrate consisted of
activated macrophages/Microglia
and astrocytes, and T lymphocytes positive for leucosyalin, CD3 and
CD8, and included secondary demyelination. We detected strong
widespread herpes simplex virus 1 thymidine kinase immunoreactivity
and vector genomes throughout large areas of the brain. Thus,
patient evaluation and the design of clinical trials in ongoing and
future gene therapy for brain glioblastoma must address not only
tumor-killing efficiency, but also long-term active brain
inflammation, loss of myelin fibers and persistent transgene
expression
Engel S, Isenmann S, Stander M, Rieger J, Bahr M, Weller M
(1999) Inhibition of experimental rat glioma growth by decorin gene
transfer is associated with decreased Microglial
infiltration. J.Neuroimmunol. 99:13-18
Abstract: Decorin gene
therapy for experimental malignant glioma is thought to involve
antagonism of immunosuppression induced by glioma-derived
transforming growth factor-beta (TGF-beta). TGF-beta is chemotactic
for cells of the monocyte macrophage lineage but inhibits their
functional activity in many in vitro paradigms. Here, we examined
changes in the patterns of Microglial
infiltration of rat C6 gliomas expressing a decorin transgene. We
find that the number of OX42/ED-1-positive Microglial
cells is reduced rather than enhanced in the presence of decorin.
Decorin-expressing gliomas contain lower numbers of MHC class II
antigen-expressing Microglial
cells whereas the relative frequency of MHC I immunoreactivity among
Microglial
cells is increased. Interestingly, the reduction of TGF-beta levels
in the tumors by decorin is associated with the de novo expression
of inducible nitric oxide synthase (iNOS) in a minority of
Microglial
cells. These data suggest that Microglial
cells do not participate in the regression of decorin-expressing rat
C6 gliomas
Flugel A, Labeur MS, Grasbon-Frodl EM, Kreutzberg GW, Graeber
MB (1999) Microglia
only weakly present glioma antigen to cytotoxic T cells.
Int.J.Dev.Neurosci. 17:547-556
Abstract: Microglia
and brain macrophages represent a substantial fraction of the cells
present in astrocytic gliomas. Yet, the functional role of Microglia
in these tumors has remained enigmatic. We have compared rat
Microglial
cells and thymocytes with regard to their ability to present
purified CNS proteins, MBP and S100beta, as well as C6 glioma cells
to specific T lymphocytes. In addition, a new cytotoxicity assay
based on fluorescence activated cell sorting of tumor
cells carrying the green fluorescent protein was established.
This assay was used to determine the influence of Microglial
population density and activational state on C6 glioma cell survival
in vitro. Microglia
were consistently found to present MBP and S100beta less efficiently
than thymocytes and appeared to be unable to present C6 glioma cells
to cytotoxic T lymphocytes. In addition, high concentrations of
Microglial
cells attenuated the cytotoxic effects of these T cells on C6 glioma
cells whereas thymocytes significantly supported their specific
killing. It is suggested that defense functions of Microglial
cells against C6 glioma are severely compromised and that the
observed deficiency in antigen presentation may play an important
role for astrocytoma growth in vivo
Krohn K (1999) TGF-beta1-dependent differential expression of
a rat homolog for latent TGF-beta binding protein in astrocytes and
C6 glioma cells. Glia 25:332-342
Abstract: Transforming growth
factor-beta1 (TGF-beta1) is widely recognized for its multiple roles
in development, cellular maintenance, and protection against injury.
In the brain, TGF-beta1 upregulation in Microglia/macrophages
is a predominant response to lesion and during pathology. However,
the precise functions of TGF-beta1 in this context are still
enigmatic. The present study investigates changes in astroglial gene
expression as a major target of TGF-beta1 signaling in the brain.
Differential display reverse transcription-polymerase chain reaction
(DDRT-PCR) was used to identify several gene fragments
differentially regulated by TGF-beta1 in rat astrocytes and C6
glioma cells. Among the cDNAs regulated by TGF-beta1 in C6 cells two
cDNAs showed homology to alpha-tropomyosin and glycerol-3-phosphate
dehydrogenase, respectively. Cloning of a full length cDNA
corresponding to a differentially regulated gene fragment revealed
close homology to latent TGF-beta binding protein (LTBP)-2. Data
using antisense LTBP-2 oligonucleotides to decrease LTBP-2
expression suggest that LTBP-2 functions to activate TGF-beta.
Therefore, it is likely that upregulation of the rat LTBP-2 homolog
mRNA in C6 cells and cortical astrocytes by TGF-1 might lead to
self-activation and exaggeration of TGF-beta signaling. These data
will extend our current understanding of TGF-beta1 functioning on
lesion-related features of glial cells
Mizuno T, Yoshihara Y, Kagamiyama H, Ohsawa K, Imai Y,
Kohsaka S, Mori K (1999) Neuronal adhesion molecule telencephalin
induces rapid cell spreading of Microglia.
Brain Res. 849:58-66
Abstract: Telencephalin (TLCN) is a neuronal
surface glycoprotein whose expression is restricted to the
telencephalon, the most rostral segment of the brain. TLCN binds to
lymphocyte function-associated antigen-1 (LFA-1) integrin. In the
central nervous system, LFA-1 is selectively and constitutively
expressed by Microglia,
suggesting that TLCN/LFA-1 binding may mediate cell-cell
interactions between telencephalic neurons and Microglia.
In the present study, we investigated the effects of recombinant
TLCN protein on the morphology of Microglia.
TLCN induced an intensive spreading of lamellipodia, causing a rapid
change in Microglial
morphology. In contrast, TLCN induced no significant change in
morphology of neuroblastoma and fibroblasts. Furthermore, the
TLCN-induced spreading of Microglia
was accompanied by a clustering of LFA-1 on cell surface membrane.
These results provide evidence that TLCN binding to the surface of
Microglia
transduces signals into Microglia
that mediate or accelerate cell spreading and LFA-1 redistribution,
implying that neuronal TLCN may control the state and/or function of
Microglia
in both physiological and pathological conditions
Morelli L, Prat MI, Castano EM (1999) Differential
accumulation of soluble amyloid beta peptides 1-40 and 1-42 in human
monocytic and neuroblastoma cell lines. Implications for cerebral
amyloidogenesis. Cell Tissue Res. 298:225-232
Abstract:
Alzheimer's disease (AD) is characterized by the massive deposition
in the brain of the 40-42-residue amyloid beta protein (A(beta)).
While A(beta)1-40 predominates in the vascular system, A(beta)1-42
is the major component of the senile plaques in the neuropil. The
concentration of both A(beta) species required to form amyloid
fibrils in vitro is micromolar, yet soluble A(betas) found in normal
and AD brains are in the low nanomolar range. It has been recently
proposed that the levels of A(beta) sufficient to trigger
amyloidogenesis may be reached intracellularly. To study the
internalization and intracellular accumulation of the major isoforms
of A(beta), we used THP-1 and IMR-32 neuroblastoma cells as models
of human monocytic and/or macrophagic and neuronal lineages,
respectively. We tested whether these cells were able to internalize
and accumulate 125I-A(beta)1-40 and 125I-A(beta)1-42 differentially
when offered at nanomolar concentrations and free of large
aggregates, conditions that mimic a prefibrillar stage of A(beta) in
AD brain. Our results showed that THP-1 monocytic cells internalized
at least 10 times more 125I-A(betas) than IMR-32 neuroblastoma
cells, either isolated or in a coculture system. Moreover,
125I-A(beta)1-42 presented a higher adsorption, internalization, and
accumulation of undigested peptide inside cells, as opposed to
125I-A(beta)1-40. These results support that A(beta)1-42, the major
pathogenic form in AD, may reach supersaturation and generate
competent nuclei for amyloid fibril formation intracellularly. In
light of the recently reported strong neurotoxicity of soluble,
nonfibrillar A(beta)1-42, we propose that intracellular
amyloidogenesis in Microglia
is a protective mechanism that may delay neurodegeneration at early
stages of the disease
Poulsen DJ, Favara C, Snyder EY, Portis J, Chesebro B (1999)
Increased neurovirulence of polytropic mouse retroviruses delivered
by inoculation of brain with infected neural stem cells. Virology
263:23-29
Abstract: Following intraperitoneal (IP) inoculation of
neonatal mice, the polytropic recombinant murine leukemia virus
(MuLV), Fr98, induces a severe brain disease characterized by
ataxia, seizures and death. In contrast, no apparent clinical
neurological disease is seen after IP infection with Fr54, a
polytropic MuLV differing from Fr98 in its envelope gene sequences.
In the brain both Fr98 and Fr54 infect primarily capillary
endothelial cells and Microglia.
However, the level of Microglial
infection by Fr98 is twofold higher than by Fr54, which might
account for the difference in neurovirulence. In the present study,
in order to test directly whether an increase in the number of
Microglia
infected by Fr54 would be sufficient to induce clinical disease, we
attempted to increase the level of Fr54 in the brain by changing the
route of infection. After intraventricular inoculation with
Fr54-infected neural stem cells (clone C17.2), a well-established
vehicle for delivery of viruses and genes to the brain, mice became
ataxic and died 4 weeks postinfection. In these mice induction of
brain disease was correlated with a higher level of viral antigen in
the cerebrum and an increase in the number of infected Microglial
cells in all brain regions examined compared with mice inoculated
IP. In contrast, mice inoculated with neural stem cells infected
with an ecotropic nonneurovirulent murine leukemia virus, FB29,
developed no clinical disease in spite of evidence for widespread
infection of Microglia
in brain. Since the main differences between Fr54 and FB29 are in
the SU (gp70) region of the envelope gene, this region is most
likely to account for the differences in induction of CNS disease
seen in the current experiments
Sandmair AM, Loimas S, Poptani H, Vainio P, Vanninen R,
Turunen M, Tyynela K, Vapalahti M, Yla-Herttuala S (1999) Low
efficacy of gene therapy for rat BT4C malignant glioma using
intra-tumoural transduction with thymidine kinase retrovirus
packaging cell injections and ganciclovir treatment. Acta
Neurochir.(Wien.) 141:867-872
Abstract: BACKGROUND: The purpose
of this study was to test the use of Herpes Simplex virus thymidine
kinase (HSVtk) retrovirus packaging cell injections in the treatment
of malignant brain tumours. METHODS: Therapeutic effect and tissue
responses were examined in vivo in a syngeneic BT4C rat glioma model
after HSVtk-producing PA317 packaging cell injections and
intraperitoneal ganciclovir (GCV) medication. MRI was used to
visualise the tumours before and after the treatment.
Immunohistochemical stainings were performed to study astroglia and
Microglia
responses and apoptosis-mediated cell death. RESULTS: The results
suggest that only a limited treatment effect can be achieved with
HSVtk packaging cell injections with no prolonged survival rates.
Histological examination showed a strong astroglia response but only
a modest Microglia
response after the treatment. HSVtk and GCV-induced cell death was
at least partially mediated by apoptosis. It is concluded that HSVtk
packaging cell injections and GCV treatment do not lead to
eradication of malignant cells in a syngeneic BT4C rat glioma model.
The lack of efficacy is most likely due to low gene transfer
efficiency and limited life span of the injected packaging cell
inside the tumours. CONCLUSIONS: Improvements in gene transfer
efficiency, and stimulation of immunoresponse against tumour cells
might lead to a more effective therapeutic response in vivo
Satoh J, Kurohara K, Yukitake M, Kuroda Y (1999) The 14-3-3
protein detectable in the cerebrospinal fluid of patients with
prion-unrelated neurological diseases is expressed constitutively in
neurons and glial cells in culture. Eur.Neurol. 41:216-225
Abstract:
The 14-3-3 protein belongs to a family of 30-kD proteins originally
identified by two-dimensional analysis of brain protein extracts.
Recently, the detection of the 14-3-3 protein in the cerebrospinal
fluid (CSF) is utilized as a highly reliable test for the premortem
diagnosis of prion diseases such as Creutzfeldt-Jakob disease. For
the initial step, to clarify the biological implication of the CSF
14-3-3 protein in these diseases, its expression was investigated in
neural tissues and cultures and CSF samples from patients with a
variety of neurological diseases by Western blot analysis and
immunocytochemistry. The constitutive expression of the 14-3-3
protein was identified in all neural and nonneural tissues examined.
It was expressed in all neurons, astrocytes, oligodendrocytes, and
Microglia
in culture with its location in both cytoplasmic and nuclear
regions. The 14-3-3 protein was detected in the CSF of 8 out of 71
patients, including 1 Gerstmann-Straussler-Scheinker disease patient
and 7 patients with prion-unrelated neurological diseases, such as
meningoencephalitis of viral, bacterial, or tuberculous origin,
multiple sclerosis, and mitochondrial myopathy, encephalopathy,
lactic acidosis, and strokelike episodes. These results suggest that
the 14-3-3 protein expressed constitutively at substantial levels in
both neurons and glial cells might be released into the CSF as a
disease-nonspecific consequence of the extensive brain damage and
indicate that the analysis of the 14-3-3 protein in the CSF is not
useful as a screening test for prion diseases
van Den Pol AN, Mocarski E, Saederup N, Vieira J, Meier TJ
(1999) Cytomegalovirus cell tropism, replication, and gene transfer
in brain. J.Neurosci. 19:10948-10965
Abstract: Cytomegalovirus
(CMV) infects a majority of adult humans. During early development
and in the immunocompromised adult, CMV causes neurological
deficits. We used recombinant murine cytomegalovirus (mCMV)
expressing either green fluorescent protein (GFP) or
beta-galactosidase under control of human elongation factor 1
promoter or CMV immediate early-1 promoter as reporter genes for
infected brain cells. In vivo and in vitro studies revealed that
neurons and glial cells supported strong reporter gene expression
after CMV exposure. Brain cultures selectively enriched in either
glia or neurons supported viral replication, leading to process
degeneration and cell death within 2 d of viral exposure. In
addition, endothelial cells, tanycytes, radial glia, ependymal
cells, Microglia,
and cells from the meninges and choroid were infected. Although mCMV
showed no absolute brain cell preference, relative cell preferences
were detected. Radial glia cells play an important role in guiding
migrating neurons; these were viral targets in the developing brain,
suggesting that cortical problems including microgyria that are a
consequence of CMV may be caused by compromised radial glia.
Although CMV is a species-specific virus, recombinant mCMV entered
and expressed reporter genes in both rat and human brain cells,
suggesting that mCMV might serve as a vector for gene transfer into
brain cells of non-murine species. GFP expression was sufficiently
strong that long axons, dendrites, and their associated spines were
readily detected in both living and fixed tissue, indicating that
mCMV reporter gene constructs may be useful for labeling neurons and
their pathways
Cunningham TJ, Hodge L, Speicher D, Reim D, Tyler-Polsz C,
Levitt P, Eagleson K, Kennedy S, Wang Y (1998) Identification of a
survival-promoting peptide in medium conditioned by oxidatively
stressed cell lines of nervous system origin. J.Neurosci.
18:7047-7060
Abstract: A survival-promoting peptide has been
purified from medium conditioned by Y79 human retinoblastoma cells
and a mouse hippocampal cell line (HN 33.1) exposed to H2O2. A 30
residue synthetic peptide was made on the basis of N-terminal
sequences obtained during purification, and it was found to exhibit
gel mobility and staining properties similar to the purified
molecules. The peptide maintains cells and their processes in vitro
for the HN 33.1 cell line treated with H2O2, and in vivo for
cortical neurons after lesions of the cerebral cortex. It has weak
homology with a fragment of a putative bacterial antigen and, like
that molecule, binds IgG. The peptide also contains a motif
reminiscent of a critical sequence in the catalytic region of
calcineurin-type phosphatases; surprisingly, like several members of
this family, the peptide catalyzes the hydrolysis of
para-nitrophenylphosphate in the presence of Mn2+. Application of
the peptide to one side of bilateral cerebral cortex lesions
centered on area 2 in rats results in an increase in IgG
immunoreactivity in the vicinity of the lesions 7 d after surgery.
Microglia
immunopositive for IgG and ED-1 are, however, dramatically reduced
around the lesions in the treated hemisphere. Furthermore, pyramidal
neurons that would normally shrink, die, or disintegrate were
maintained, as determined by MAP2 immunocytochemistry and Nissl
staining. These survival effects were often found in both
hemispheres. The results suggest that this peptide operates by
diffusion to regulate the immune response and thereby rescue neurons
that would usually degenerate after cortical lesions. The
phosphatase activity of this molecule also suggests the potential
for direct neuron survival-promoting effects
Fiebich BL, Hull M, Lieb K, Schumann G, Berger M, Bauer J
(1998) Potential link between interleukin-6 and arachidonic acid
metabolism in Alzheimer's disease. J.Neural Transm.Suppl
54:268-278
Abstract: Prostaglandins (PGs) and cytokines, such as
interleukin-1 (IL-1) and interleukin-6 (IL-6), have been implicated
in the etiopathology of various inflammatory and degenerative
disorders, including Alzheimer's disease (AD). Previously, we
detected the presence of IL-6 in cortices of AD patients. On the
other hand, non-steroidal antiinflammatory drugs (NSAIDs), potent
inhibitors of prostaglandin synthesis, have been shown to be
beneficial in the treatment of AD. Until now, it remained unclear
whether and how these two observations were functionally connected.
Here, we show that PGs are able to induce IL-6 synthesis in a human
astrocytoma cell line. PGE1 and PGE2, but not PGD2 and PGF2 alpha,
led to a rapid and transient induction of astrocytic IL-6 mRNA,
followed by IL-6 protein synthesis. Furthermore, PGE2 potentiated
IL-1 beta-induced IL-6 mRNA synthesis. These results suggest a
possible link between the release of PGs from activated Microglia
and the astrocytic synthesis of IL-6, which itself may affect
neuronal cells, as hypothesized for Alzheimer's disease. Finally we
demonstrate that Microglia
are a strong source of PGE2 synthesis indicating that these cells
may act as the origin of the pathogenic cascade
Gasque P, Singhrao SK, Neal JW, Wang P, Sayah S, Fontaine M,
Morgan BP (1998) The receptor for complement anaphylatoxin C3a is
expressed by myeloid cells and nonmyeloid cells in inflamed human
central nervous system: analysis in multiple sclerosis and bacterial
meningitis. J.Immunol. 160:3543-3554
Abstract: The complement
anaphylatoxins C5a and C3a are released at the inflammatory site,
where they contribute to the recruitment and activation of
leukocytes and the activation of resident cells. The distribution of
the receptor for C5a (C5aR) has been well studied; however, the
receptor for C3a (C3aR) has only recently been cloned, and its
distribution is uncharacterized. Using a specific affinity-purified
anti-C3aR peptide Ab and oligonucleotides for reverse
transcriptase-PCR analysis, C3aR expression was characterized in
vitro on myeloid and nonmyeloid cells and in vivo in the brain. C3aR
was expressed by adult astrocytes, astrocyte cell lines, monocyte
lines THP1 and U937, neutrophils, and monocytes, but not by K562 or
Ramos. C3aR staining was confirmed by flow cytometry, confocal
imaging, and electron microscopy analysis. A 65-kDa protein was
immunoprecipitated by the anti-C3aR from astrocyte and monocyte cell
lysates. Our results at the protein level were confirmed at the mRNA
level. Using reverse transcriptase-PCR, Southern blot, and
sequencing we found that C3aR mRNA was expressed by fetal
astrocytes, astrocyte cell lines, and THP1, but not by K562 or
Ramos. The astrocyte C3aR cDNA was identical with the reported C3aR
cDNA. C3aR expression was not detected in normal brain sections.
However, a strong C3aR staining was evident in areas of inflammation
in multiple sclerosis and bacterial meningitis. In meningitis, C3aR
was abundantly expressed by reactive astrocytes, Microglia,
and infiltrating cells (macrophages and neutrophils). In multiple
sclerosis, infiltrating lymphocytes did not express C3aR, but a
strong staining was detected on smooth muscle cells (pericytes)
surrounding blood vessels
Itayasu T, Shimizu T, Iizumi T, Oshio S, Umeda T, Takeda K
(1998) Phorbol ester induces differentiation of a human prostatic
cancer cell line TSU-Pr1 into cells with characteristics of
Microglia.
Anticancer Res. 18:113-117
Abstract: The effects of various
reagents on the induction of differentiation of the human prostatic
cancer cell line, TSU-Pr1, were examined. Among these agents, the
phorbol ester, TPA, almost completely suppressed cell proliferation
at the concentration of 10(-8) M, and induced remarkable morphologic
changes yielding cells with the Microglial
feature of an ameboid and/or ramified shape. More than 90% of the
cells underwent the induction of morphologic changes by day 7 after
treatment with 10(-8) M TPA. The expression of reliable Microglial
markers, alpha-naphthyl acetate esterase activity and CD11b, was
observed in the differentiated cells. The data presented here
suggest that TPA induces differentiation of a human prostate cancer
cell line into cells with the characteristics of Microglia
Makman MH, Dobrenis K, Surratt CK (1998) Properties of mu 3
opiate alkaloid receptors in macrophages, astrocytes, and HL-60
human promyelocytic leukemia cells. Adv.Exp.Med.Biol.
437:137-148
Abstract: An opiate alkaloid-selective receptor,
designated mu(3), mediates inhibition by morphine of activation of
human peripheral blood monocytes and granulocytes. The mu(3)
receptor is present on several macrophage cell types including
Microglia,
on cultured astrocytes, and in brain and retina. Murine macrophage
cell lines and human HL-60 leukemia cells contain high
concentrations of these receptors. Binding of 3H-morphine to the
receptor is displaced by morphine, etorphine, naloxone,
diprenorphine and morphine 6-glucuronide, but not by morphine
3-glucuronide, fentanyl, benzomorphans, enkephalins, dynorphin,
beta-endorphin, endomorphin-1, other opioid peptides or nociceptin
(orphanin FQ). The mu(3) receptor appears to be much more sensitive
to inactivation by reduced glutathione than are classical mu, delta
and kappa receptors. Evidence is also presented for G
protein-coupling of these receptors. These and other data raise the
possibility that the mu(3) receptor is a member of a chemokine or of
another related receptor family, rather than the opioid receptor
family. The affinity for morphine of mu(3) receptors of
granulocytic-differentiated HL-60 cells is markedly enhanced in the
presence of levorphanol and certain benzomorphans. In contrast,
receptors of monocytes, macrophage cell lines, Microglia,
macrophage-differentiated HL-60 cells and astrocytes are not
affected by levorphanol or benzomorphans. It is concluded that mu(3)
receptors of granulocytic and promyelocytic cells differ from those
of macrophage and astrocyte cell types, possibly due to differences
in receptor subtype or to the presence of an additional component in
the granulocytic and promyelocytic cells
Shieh JT, Albright AV, Sharron M, Gartner S, Strizki J, Doms
RW, Gonzalez-Scarano F (1998) Chemokine receptor utilization by
human immunodeficiency virus type 1 isolates that replicate in
Microglia.
J.Virol. 72:4243-4249
Abstract: The role of human
immunodeficiency virus (HIV) strain variability remains a key
unanswered question in HIV dementia, a condition affecting around
20% of infected individuals. Several groups have shown that viruses
within the central nervous system (CNS) of infected patients
constitute an independently evolving subset of HIV strains. A
potential explanation for the replication and sequestration of
viruses within the CNS is the preferential use of certain chemokine
receptors present in Microglia.
To determine the role of specific chemokine coreceptors in infection
of adult Microglial
cells, we obtained a small panel of HIV type 1 brain isolates, as
well as other HIV strains that replicate well in cultured Microglial
cells. These viruses and molecular clones of their envelopes were
used in infections, in cell-to-cell fusion assays, and in the
construction of pseudotypes. The results demonstrate the predominant
use of CCR5, at least among the major coreceptors, with minor use of
CCR3 and CXCR4 by some of the isolates or their envelope clones
Stoica G, Barker R, Wu G, Lynn WS, Wong PK (1998)
Quantitative variation of free amino acids in the central nervous
system of MoMuLV-ts1-infected mice. In Vivo 12:395-401
Abstract:
A temperature-sensitive mutant of Moloney murine leukemia virus
(MoMuLV-ts1) induces polioencephalomyelopathy and hind limb
paralysis in highly susceptible FVB/N strains of neonatal mice. This
disease is characterized by progressive motor neurons loss and
severe gliosis within specific target areas of the central nervous
system (CNS). The mechanism(s) of this neurodegeneration is unknown.
In the neonatal infection of the CNS, the MoMuLV-ts1 virus was
reported to replicate within the endothelial, ependymal, astrocytes
and Microglial
cells. Since no virus or viral products were recognized in the
degenerating neurons, it is postulated that an indirect mechanism(s)
caused the loss of neurons in the neonatally infected mice. This
study was undertaken to investigate the possible pathogenic role of
excitatory amino acids (EAAs) such as glutamate and other
nonneurotransmitters amino acids (NAAs) in this animal model. The
free amino acids concentration was analysed by a fluorometric HPLC
method. The temporal measurements of the free amino acids
concentration, glutamate, glutamine and arginine from the brain stem
and spinal cord of MoMuLV-ts1-infected mice was significantly
decreased when compared with the control non-infected mice. The
concentration of EAAs during the course of this infection indicated
a sharp decline in glutamate and its precursor, glutamine with early
infection (10 days post infection-dpi). This deficiency persisted
(20 and 30 dpi) in the spinal cord, where the neuronal loss was most
severe, but not in the brain stem. A similar pattern occurs with the
amino acid arginine. These observations suggest that an
astrocyte-induced metabolic disturbance of glutamate and arginine in
the CNS of developing mice, could be, in part responsible for the
loss of motor neurons observed in this model
Xia MQ, Berezovska O, Kim TW, Xia WM, Liao A, Tanzi RE,
Selkoe D, Hyman BT (1998) Lack of specific association of presenilin
1 (PS-1) protein with plaques and tangles in Alzheimer's disease.
J.Neurol.Sci. 158:15-23
Abstract: Missense mutations in the
presenilin-1 (PS-1) gene are causally related to the majority of
familial early-onset Alzheimer's disease (FAD). PS-1
immunohistochemical expression in normal human brain and in brains
with Alzheimer's disease (AD) has so far been controversial. Here,
we report a study of PS-1 expression in brains, cell lines and
peripheral blood mononuclear cells using a panel of well
characterized PS-1-specific antibodies. These antibodies were
characterized by immunofluorescent staining of PS-1 transfectants
followed by flow cytometric analysis. In human brain, widespread
neuronal staining was observed. PS-1 immunoreactivity was primarily
confined to neuronal cell bodies and proximal dendrites. Weaker
staining of Microglia
was also detected, in accord with the finding of PS-1
immunoreactivity in monocytes. PS-1 expression is not particularly
associated with neurons either containing or spared from
neurofibrillary tangles, nor with senile plaques. The level of PS-1
expression does not differ between normal and AD brains.
Immunoprecipitation from AD, FAD and control brains revealed only a
32 kDa N-terminal fragment and an 18-20 kDa C-terminal fragment.
Little or no full length PS-1 was detected. The enriched presence of
PS-1 in neurons implies an important role in neuronal function,
however, the lack of apparent association of its expression with AD
pathology signifies the need for a better understanding of its
pathophysiological role
Yang F, Sun X, Beech W, Teter B, Wu S, Sigel J, Vinters HV,
Frautschy SA, Cole GM (1998) Antibody to caspase-cleaved actin
detects apoptosis in differentiated neuroblastoma and
plaque-associated neurons and Microglia
in Alzheimer's disease. Am.J.Pathol. 152:379-389
Abstract: During
apoptosis, activation of a family of cysteine proteases related to
interleukin-1beta-converting enzyme (ICE)-related proteases or
"caspases" results in endoproteolytic cleavage of multiple
substrates at specific aspartate residues. We have sought to develop
new antibody probes for the neoepitopes in protein fragments
produced by ICE-related proteolytic cleavage as specific markers of
events tightly linked to apoptotic mechanisms. Here, we demonstrate
that an antibody probe specific for the C terminus of a 32-kd actin
fragment produced by ICE-like activity specifically labels apoptotic
but not necrotic, differentiated human neuroblastoma cells in
culture. Unlike probes for nonspecific DNA strand breaks confined to
the nucleus or cell body, this method allows the detection of
cytoskeletal fragments in cell processes as well as the perikaryon
long before DNA fragmentation and cell death and therefore serves as
a novel marker of apoptosis-related events in distal parts of cells
such as axons and dendrites. To illustrate this new tool, we show
that the antibody detects the processes and cell bodies of
degenerating neurons and plaque-associated Microglia
in Alzheimer's disease. In situ detection of caspase-cleaved actin
provides a new means to evaluate the role of caspase activation in
pathological and physiological processes
Yeung MC, Geertsma F, Liu J, Lau AS (1998) Inhibition of
HIV-1 gp120-induced apoptosis in neuroblastoma SK-N-SH cells by an
antisense oligodeoxynucleotide against p53. AIDS
12:349-354
Abstract: OBJECTIVES: This study examines the
cytotoxicity potential and the mechanism of toxicity of the HIV-1
gp120 on human neuroblastoma cells. DESIGN: Previous data from our
group have suggested that the HIV-1 envelope protein gp120 promotes
the secretion of tumor necrosis
factor-alpha and other factors by astrocytes and Microglial
cells present in primary human brain cell cultures, thereby
contributing to the injury of neurons in these cultures. This study
investigates the cytotoxicity potential and the mechanism of
toxicity of gp120 on human neuroblastoma cells. METHODS: SK-N-SH
cells were treated with HIV-1 gp120, and was followed by in situ DNA
fragmentation staining and small molecular weight DNA extraction
studies to ascertain the induction of apoptosis by gp120 in these
cells. To evaluate a potential role of the growth suppressor gene
p53, gp120-treated SK-N-SH cells were subjected to reverse
transcription polymerase chain reaction (RT-PCR) and Western blot
analyses for the induction of p53. An antisense oligodeoxynucleotide
against p53 was used to investigate the role of p53 in the
gp120-induced apoptosis in these cells. RESULTS: Data from T7 DNA
polymerase staining and small molecular weight DNA extraction
studies demonstrated that gp120-induced DNA breakage in SK-N-SH
cells with fragmentation patterns characteristic of apoptosis.
RT-PCR and Western blot analyses revealed that the gp120-mediated
induction of apoptosis was dependent on a gp120-induced and
gp120-sustained upregulation of p53. The induction of p53 by gp120
was specific, since an antibody against gp120 prevented both the
induction of p53 and subsequent apoptosis in SK-N-SH cells. The
critical role of p53 was further illustrated by the effectiveness of
a p53 antisense oligodeoxynucleotide to inhibit the gp120-induced
apoptosis. As a control, the apoptosis-inducing potential of gp120
on SK-N-SH cells was not seen in the HIV-1 Gag proteins even when
used at up to 5 nM. CONCLUSIONS: These results established that
HIV-1 gp120 is potentially cytotoxic to human neuronal cells through
the induction of p53, which may eventually lead to induction of
apoptosis
Bosman GJ, Renkawek K, Van Workum FP, Bartholomeus IG, De
Grip WJ (1997) Involvement of neuronal anion exchange proteins in
cell death in Alzheimer's disease. Gerontology 43:67-78
Abstract:
Anion exchange (AE) proteins are present in human neurons in the
brain. Immunohistochemical data indicate that their apparent
expression level increases with age, and especially with
degeneration in Alzheimer's disease-affected brain areas. The
increase in immunoreactivity is probably caused by changes in AE
structure that lead to an increased accessibility of hitherto hidden
epitopes. These epitopes correspond to regions in the membrane
domain that are involved in generation of senescent cell-specific
antigen from AE1 in aging erythrocytes. Elucidation of the molecular
nature of these changes and the underlying mechanisms, will lead to
insight in the processes that govern aging- and
degeneration-associated perturbation of membrane integrity.
AE-mediated chloride/bicarbonate exchange is a major component in
the regulation of intracellular pH. The functional consequences of
changes in AE structure may range from acidosis, disturbance of
cytoskeleton integrity, and untimely or impaired recognition of
cells by components of the immune system, such as Microglia.
A molecular and physiological description of these changes will
establish AE proteins as valuable tools in elucidating the processes
of normal aging, and the disturbances in aging-related diseases such
as Alzheimer's disease
Du YS, Zhu H, Fu J, Yan SF, Roher A, Tourtellotte WW,
Rajavashisth T, Chen X, Godman GC, Stern D, Schmidt AM (1997)
Amyloid-beta peptide-receptor for advanced glycation endproduct
interaction elicits neuronal expression of macrophage-colony
stimulating factor: a proinflammatory pathway in Alzheimer disease.
Proc.Natl.Acad.Sci.U.S.A 94:5296-5301
Abstract: In Alzheimer
disease (AD), neurons are thought to be subjected to the deleterious
cytotoxic effects of activated Microglia.
We demonstrate that binding of amyloid-beta peptide (Abeta) to
neuronal Receptor for Advanced Glycation Endproduct (RAGE), a cell
surface receptor for Abeta, induces macrophage-colony stimulating
factor (M-CSF) by an oxidant sensitive, nuclear factor
kappaB-dependent pathway. AD brain shows increased neuronal
expression of M-CSF in proximity to Abeta deposits, and in
cerebrospinal fluid from AD patients there was approximately 5-fold
increased M-CSF antigen (P < 0.01), compared with age-matched
controls. M-CSF released by Abeta-stimulated neurons interacts with
its cognate receptor, c-fms, on Microglia,
thereby triggering chemotaxis, cell proliferation, increased
expression of the macrophage scavenger receptor and apolipoprotein
E, and enhanced survival of Microglia
exposed to Abeta, consistent with pathologic findings in AD. These
data delineate an inflammatory pathway triggered by engagement of
Abeta on neuronal RAGE. We suggest that M-CSF, thus generated,
contributes to the pathogenesis of AD, and that M-CSF in
cerebrospinal fluid might provide a means for monitoring neuronal
perturbation at an early stage in AD
Huettner C, Czub S, Kerkau S, Roggendorf W, Tonn JC (1997)
Interleukin 10 is expressed in human gliomas in vivo and increases
glioma cell proliferation and motility in vitro. Anticancer Res.
17:3217-3224
Abstract: Interleukin 10 (IL-10) is a cytokine with
a broad spectrum of immunosuppressive activity, but itoffs role in
the oncogenesis of solid tumors is still unclear. In previous
experiments we have shown that IL-10 specific mRNA is produced
within glial tumors in vivo. The aim of the present study was to
investigate the expression of the IL-10 protein in vivo and to
identify the cells producing IL-10 within the tumor
tissue. Expression levels significantly increased with
malignancy of the gliomas. 87.5% of grade III and IV, but only 4% of
grade II tumors expressed high levels of mRNA. Elevation of IL-10
serum levels was found in 11% of low grade and in 63.6% of high
grade glioma patients. In situ hybridization analysis with combined
immunohistochemistry revealed that: a) IL-10 is not produced by
infiltrating B- or T- lymphocytes, b) both Microglia
and astroglia contributed to IL-10 expression in malignant gliomas
in vivo. These data suggested the functional role of IL-10 in glioma
progression. Therefore, the effects of IL-10 on proliferation and
migration of glioma cells were determined in vitro. Two human glioma
cell lines were grown as monolayer as well as spheroids in the
presence of different concentrations of IL-10. IL-10 increased cell
proliferation significantly in both culture systems with a dose
optimum of 25 ng/ml. Glioma cell motility was enhanced with 25 ng/ml
as the optimal dose. Adding the IL-10 specific antibody reversed
both effects. We conclude from our data that IL-10 is involved in
the progression of glial tumors, especially in the enhancement of
tumor cell proliferation
and migration which promotes infiltration of the surrounding tissue
Li Y, Kustova Y, Sei Y, Basile AS (1997) Regional changes in
constitutive, but not inducible NOS expression in the brains of mice
infected with the LP-BM5 leukemia virus. Brain Res.
752:107-116
Abstract: Potential neurotoxins such as nitric oxide
have been implicated in the pathogenesis of acquired
immunodeficiency syndrome (AIDS) dementia complex. The LP-BM5 murine
leukemia-infected mice, which develop immunological and cognitive
deficits reminiscent of human HIV-1 infection, were employed to
investigate the changes in brain constitutive nitric oxide synthase
(cNOS) and inducible nitric oxide synthase (iNOS) expression.
Cerebellar and striatal cNOS enzymatic activity increased
approximately 70% as early as 2 weeks after infection, declining to
control levels by 12-16 weeks. In contrast, cNOS protein expression
in the striatum and cerebellum was decreased 30% at 4 weeks,
declining to 50% of control levels by 16 weeks post-infection.
Staining intensity for cNOS, but not neuron number was reduced in
the cerebral cortex, striatum, ventromedial hypothalamic nucleus and
amygdala. Although iNOS protein expression was elevated in splenic
monocytes, neither iNOS activity, mRNA nor protein was detected in
the brains of mice 12 weeks after infection. These results indicate
that neurons decrease cNOS protein expression to compensate for
chronic cNOS activation, probably resulting from glutamatergic
stimulation. The cNOS activation is contemporaneous with Microglial
activation in LP-BM5-infected mice, and precedes the development of
cognitive deficits. Moreover, the lack of iNOS induction in either
infected macrophages or glial elements suggests that iNOS is not
necessary for the development of these cognitive deficits
Malhotra SK, Luong LT, Bhatnagar R, Shnitka TK (1997)
Up-regulation of reactive astrogliosis in the rat glioma 9L cell
line by combined mechanical and chemical injuries. Cytobios
89:115-134
Abstract: The 9L rat glioma cells grown in culture,
when subjected to a mechanical injury (scratch wound) and/or a
chemical injury (CdCl2) manifest changes which are characteristic of
an astrocyte reaction (astrogliosis) in the central nervous system.
Such changes include cell hypertrophy and an increase in
immunostaining for the astrocytic marker proteins, glial fibrillary
acidic protein and J1-31 antigen. Mitochondria also increase in size
and number, and the endoplasmic reticulum expands in area. These
mechanical and chemical injuries are coordinated, and act
synergistically to induce a considerably more intense astroglial
reaction by 9L cells than can be elicited with either injurious
agent alone, and this occurs without any interactions with
Microglia,
neurons or oligodendroglia. The phenomenon suggests that more than
one transcriptional mechanism is involved in the activation of
astrocytes, and that mechanical and CdCl2-induced injuries,
respectively, probably affect different receptors and second- and
third-messenger pathways. There are a number of questions concerning
the molecular biology of reactive astrocytes which can be addressed
through the use of the 9L rat glioma cell model. This model offers
certain advantages over primary cultures of astrocytes, namely a low
basal level of reactivity (because the cells are not subjected to
mechanical injury prior to experimentation), an absence of
contaminating Microglial
cells, greater ease of reproducibility of results, lower costs and
avoidance of the use of animals
McDonald DR, Brunden KR, Landreth GE (1997) Amyloid fibrils
activate tyrosine kinase-dependent signaling and superoxide
production in Microglia.
J.Neurosci. 17:2284-2294
Abstract: Alzheimer's disease (AD) is a
devastating neurological disorder characterized by loss of cognitive
skills and progressive dementia. The pathological hallmark of AD is
the presence of numerous senile plaques throughout the hippocampus
and cerebral cortex associated with degenerating axons,
neurofibrillary tangles, and gliosis. The core of the senile plaque
primarily is composed of the 39-43 amino acid beta-amyloid peptide
(Abeta), which forms fibrils of beta-pleated sheets. Although
considerable genetic evidence implicates Abeta in the pathogenesis
of AD, a direct causal link remains to be established. Senile
plaques are foci of local inflammatory processes, as evidenced by
the presence of numerous activated Microglia
and acute phase proteins. Abeta has been shown to elicit
inflammatory responses in Microglia;
however, the intracellular events mediating these effects are
largely unknown. We report that exposure of Microglia
and THP1 monocytes to fibrillar Abeta led to time- and
dose-dependent increases in protein tyrosine phosphorylation of a
population of proteins similar to that elicited by classical immune
stimuli such as immune complexes. The tyrosine kinases Lyn, Syk, and
FAK were activated on exposure of Microglia
and THP1 monocytes to Abeta, resulting in the tyrosine
kinase-dependent generation of superoxide radicals. The present data
support a role for oxidative damage in the pathogenesis of AD,
provide an important mechanistic link between Abeta and the
generation of reactive oxygen intermediates, and identify molecular
targets for therapeutic intervention in AD
Moffett JR, Els T, Espey MG, Walter SA, Streit WJ, Namboodiri
MA (1997) Quinolinate immunoreactivity in experimental rat brain
tumors is present in macrophages but not in astrocytes. Exp.Neurol.
144:287-301
Abstract: Experimental tumors of the central nervous
system were investigated with antibodies to quinolinate to assess
the cellular distribution of this endogenous neurotoxin. In advanced
F98 and RG-2 glioblastomas and E367 neuroblastomas in the striatum
of rats, variable numbers of quinolinate immunoreactive cells were
observed in and around the tumors, with the majority being present
within tumors, rather than brain parenchyma. The stained cells were
morphologically variable, including round, complex, rod-shaped, and
sparsely dendritic cells. Neuroblastoma and glioma cells were
unstained, as were neurons, astrocytes, oligodendrocytes, ependymal
cells, endothelial cells, and cells of the choroid plexus and
leptomeninges. Glial fibrillary acidic protein immunoreactivity was
strongly elevated in astrocytes surrounding the tumors. Dual
labeling immunohistochemistry with antibodies to quinolinate and
glial fibrillary acidic protein demonstrated that astrocytes and the
cells containing quinolinate immunoreactivity were morphologically
disparate and preferentially distributed external and internal to
the tumors, respectively, and no dual labeled cells were observed.
Lectin histochemistry with Griffonia simplicifolia B4 isolectin and
Lycopersicon esculentum lectin demonstrated numerous phagocytic
macrophages and reactive Microglia
in and around the tumors whose distribution was similar to that of
quinolinate immunoreactive cells, albeit much more numerous. Dual
labeling studies with antibodies to quinolinate and the lectins
demonstrated partial codistribution of these markers, with most
double-labeled cells having the morphology of phagocytes. The
present findings suggest the possibility that quinolinate may serve
a functional role in a select population of inflammatory cell
infiltrates during the immune response to brain neoplasms
Papavasiliou AK, Mehler MF, Mabie PC, Marmur R, Qingbin S,
Keating RF, Kessler JA (1997) Paracrine regulation of
colony-stimulating factor-1 in medulloblastoma: implications for
pathogenesis and therapeutic interventions. Neurosurgery
41:916-923
Abstract: OBJECTIVE: Colony-stimulating factor
(CSF)-1, a chemotactic and mitogenic factor for macrophages and
Microglia,
is expressed in a variety of nervous system tumors and when present
in nonneural malignancies, is associated with marked inflammatory
infiltrates, dissemination, and poorer prognosis. This study
investigated the paracrine effects of CSF-1 production by
medulloblastoma cells on the macrophage/Microglial
lineage. METHODS: A recurrent metastatic desmoplastic
medulloblastoma was isolated from a 26-year-old man and propagated
in tissue culture. Cellular phenotype and proliferation were
assessed by immunocytochemical techniques; transcript expression for
CSF-1, granulocyte macrophage-CSF, interleukin-3, and c-fms (the
receptor for CSF-1) was examined with reverse
transcriptase-polymerase chain reaction; and conditioned media and
coculture paradigms were used to study cytokine effects on cellular
proliferation. RESULTS: Serially passaged cells were uniformly
immunoreactive for two lineage-independent neuroepithelial markers,
nestin and vimentin. A subpopulation of cells with morphological
characteristics of early differentiation stained for neurofilament
66 (7%) and microtubule-associated protein (5%) (markers of early
neuronal precursors and postmitotic neurons, respectively) and for
the Yp subunit of glutathione-S-transferase (3%) (a marker of early
oligodendroglial progenitors). tumor
cells expressed transcripts for CSF-1, but not for
granulocyte macrophage-CSF, interleukin-3, or c-fms. Treatment of
Microglia
with serum-free medulloblastoma-conditioned media significantly
increased proliferation (P < 0.001), suggesting the secretion of
CSF-1. Coculture of medulloblastoma cells and Microglia
significantly increased proliferation of both cell types (each
condition, P < 0.01). CONCLUSION: These observations suggest that
CSF-1 mediates important paracrine interactions between transformed
cells and the immune system, resulting in increased growth rate and
metastatic potential. Future therapeutic goals need to include
immunotherapeutic protocols to modulate this interaction
Qiu WQ, Ye Z, Kholodenko D, Seubert P, Selkoe DJ (1997)
Degradation of amyloid beta-protein by a metalloprotease secreted by
Microglia
and other neural and non-neural cells. J.Biol.Chem.
272:6641-6646
Abstract: Amyloid beta-protein (Abeta) is the major
component of neuritic (amyloid) plaques in Alzheimer's disease, and
its deposition is an early and constant event in the complex
pathogenetic cascade of the disease. Although many studies have
focused on the biosynthetic processing of the beta-amyloid precursor
protein and on the production and polymerization of Abeta,
understanding the degradation and clearance of Abeta has received
very little attention. By incubating the conditioned medium of
metabolically labeled Abeta-secreting cells with media of various
cultured cell lines, we observed a time-dependent decrease in the
amount of Abeta in the mixed media. The factor principally
responsible for this decrease was a secreted metalloprotease
released by both neural and non-neural cells. Among the cells
examined, the Microglial
cell line, BV-2, produced the most Abeta-degrading activity. The
protease was completely blocked by the metalloprotease inhibitor,
1,10-phenanthroline, and partially inhibited by EDTA, whereas
inhibitors of other protease classes produced little or no
inhibition. Substrate analysis suggests that the enzyme was a
non-matrix metalloprotease. The protease cleaved both Abeta1-40 and
Abeta1-42 peptides secreted by beta-amyloid precursor
protein-transfected cells but failed to degrade low molecular weight
oligomers of Abeta that form in the culture medium.
Lipopolysaccharide, a stimulator of macrophages/Microglia,
activated BV-2 cells to increase their Abeta-degrading
metalloprotease activity. We conclude that secreted Abeta1-40 and
Abeta1-42 peptides are constitutively degraded by a metalloprotease
released by Microglia
and other neural cells, providing a potential mechanism for the
clearance of Abeta in brain tissue
Robertson SJ, Hasenkrug KJ, Chesebro B, Portis JL (1997)
Neurologic disease induced by polytropic murine retroviruses:
neurovirulence determined by efficiency of spread to Microglial
cells. J.Virol. 71:5287-5294
Abstract: Several murine leukemia
viruses (MuLV) induce neurologic disease in susceptible mice. To
identify features of central nervous system (CNS) infection that
correlate with neurovirulence, we compared two neurovirulent MuLV,
Fr98 and Fr98/SE, with a nonneurovirulent MuLV, Fr54. All three
viruses utilize the polytropic receptor and are coisogenic, each
containing a different envelope gene within a common genetic
background. Both Fr98 and Fr98/SE induce a clinical neurologic
disease characterized by hyperexcitability and ataxia yet differ in
incubation period, 16 to 30 and 30 to 60 days, respectively. Fr54
infects the CNS but fails to induce clinical signs of neurologic
disease. In this study, we compared the histopathology, regional
virus distribution, and cell tropism in the brain, as well as the
relative CNS viral burdens. All three viruses induced similar
histopathologic effects, characterized by intense reactive
astrogliosis and Microglial
activation associated with minimal vacuolar degeneration. The
infected target cells for each virus consisted primarily of
endothelial and Microglial
cells, with rare oligodendrocytes. Infection localized predominantly
in white matter tracts of the cerebellum, internal capsule, and
corpus callosum. The only feature that correlated with relative
neurovirulence was viral burden as measured by both viral CA protein
expression in cerebellar homogenates and quantification of infected
cells. Interestingly, Fr54 (nonneurovirulent) and Fr98/SE (slow
disease) had similar viral burdens at 3 weeks postinoculation,
suggesting that they entered the brain with comparable efficiencies.
However, spread of Fr98/SE within the brain thereafter exceeded that
of Fr54, reaching levels of viral burden comparable to that seen for
Fr98 (rapid disease) at 3 weeks. These results suggest that the
determinants of neurovirulence in the envelope gene may influence
the efficiency of virus spread within the brain and that a critical
number of infected cells may be required for induction of clinical
neurologic disease
Rosales AA, Roque RS (1997) Microglia-derived
cytotoxic factors. Part I: Inhibition of tumor
cell growth in vitro. Brain Res. 748:195-204
Abstract: The
rarity of neoplasms in the adult mammalian retina has led us to
hypothesize the presence or increased expression of
'tumor-inhibitory molecules' in the mature differentiated retina. We
have begun to investigate the source(s) of these molecules, and the
following study describes the inhibitory activity of a soluble
Microglia-derived
cytotoxic factor on the proliferation of C6 cells, a glial tumor
cell line. C6 cells were treated for 24, 48, or 72 h with
basal medium or basal medium conditioned by retina-derived Muller
cells (MCCM) or Microglial
cells (MGCM) and assayed for cell proliferation and/or cell death
using various techniques involving fluorescent probes, lactate
dehydrogenase release, or bromodeoxyuridine uptake. C6 cells
increased in number from 24 to 72 h following incubation in basal
medium or MCCM, but not in MGCM, where the cells rounded up and
retracted their processes. The number of dead cells appeared to be
the same in all groups at each time point. Similar findings were
observed in the presence of 1-10% serum. About 25% of cells treated
with basal medium for 72 h were positive for bromodeoxyuridine as
compared with < 1% in MGCM-treated cultures. Our studies suggest
that retina-derived Microglial
cells secrete soluble product(s) that inhibit the growth of C6 cells
in culture. These molecules may provide protection for the mature
retina against the invasion of tumor
cells and may prove useful in the treatment of cancer
Saida K, Saida T, Kai K, Iwamura K (1997) Central nervous
system lesions in rats infected with Friend murine leukemia
virus-related PVC441: ultrastructural and immunohistochemical
studies. Acta Neuropathol.(Berl) 93:369-378
Abstract: Newborn
F344 rats were injected intraperitoneally with PVC441 virus, a
neuropathogenic variant of Friend murine leukemia virus, and
developed paraparesis of hind limbs 35-40 days after infection.
Immunohistochemical study using monoclonal anti-PVC441 antibody
revealed that in the central nervous system endothelial cells but
not neuronal or glial cells were infected with PVC441 virus. The
major pathological changes were myelin vacuolation and
oligodendrocyte degeneration in the white matter at the white-gray
border zone. Anterior and lateral funiculi and intercalated myelin
of anterior horns were dominantly affected in the spinal cord from
the sacral to cervical level. The midbrain was also vacuolated. An
ultrastructural study demonstrated that many viral particles were
present outside the endothelial cells but only sparsely inside
endothelial cells and pericytes. Endothelial cell membranes and
tight junctions were also disrupted. Immunohistochemical studies
with antibodies against major histo-compatibility complex class Ia,
intercellular adhesion molecule-I, glial fibrillary acidic protein,
neurofilament protein, CD3 and OX42 revealed the presence of
abundant Microglia
but not of lymphocytes or polymorphonuclear cells in the lesions.
Axonal degeneration and astrogliosis were mild in degree. These
pathological changes explain the observed spastic paraparesis in the
rats, and represent a good model of spongiform diseases of the human
central nervous system of retroviral origin, such as human T cell
leukemia virus-associated myelopathy and AIDS
Strizki JM, Turner JD, Collman RG, Hoxie J, Gonzalez-Scarano
F (1997) A monoclonal antibody (12G5) directed against CXCR-4
inhibits infection with the dual-tropic human immunodeficiency virus
type 1 isolate HIV-1(89.6) but not the T-tropic isolate HIV-1(HxB).
J.Virol. 71:5678-5683
Abstract: We used a monoclonal antibody
(12G5) directed against an extracellular domain of CXCR-4 to
investigate the role of this receptor in infection of immortalized
lymphoid cell lines, peripheral blood mononuclear cells (PBMCs), and
primary brain Microglia
with a dual-tropic strain of human immunodeficiency virus
(HIV-1(89.6)) and a T-tropic strain (HIV-1(IIIB)). Addition of
antibody 12G5 to cells prior to and during infection with
HIV-1(89.6) inhibited p24 production 100- to 10,000-fold in CEMx174
and 174-CD4 cells and about 10-fold in PBMC cultures but had no
activity against infection of either monocyte-derived macrophages or
brain Microglia.
In contrast, 12G5 had little or no effect on infection of CEMx174
cells with HIV-1(IIIB) or HIV-1(HxB). To identify the region of the
HIV-1(89.6) envelope that confers sensitivity to 12G5, we used
chimeric molecular clones. Chimeras containing the V3 loop region of
HIV-1(89.6) were inhibited by 12G5 to the same degree as wild-type
HIV-1(89.6) whereas replication of those viruses containing the V3
loop of HIV-1(HxB) was not inhibited by the antibody. A similar
pattern was seen in infections of a U87 glioblastoma line that
coexpresses CD4 and CXCR-4. Antibody 12G5 was also able to block
fusion between HeLa-CD4 cells and CEMx174 cells chronically infected
with HIV-1(89.6) but had no effect on fusion mediated by cells
chronically infected with HIV-1(IIIB). Taken together, these results
suggest that different strains of HIV-1 may interact with different
sites on CXCR-4 or may have different binding affinities for the
coreceptor
Sugawa M, Ikeda S, Kushima Y, Takashima Y, Cynshi O (1997)
Oxidized low density lipoprotein caused CNS neuron cell death. Brain
Res. 761:165-172
Abstract: Death induced by oxidized low density
lipoproteins (oxLDL) to embryonic CNS neuronal and neuroblastoma
cells was investigated. Cell damage and viability were evaluated by
LDH leakage and the MTT method, respectively. Dose- and
time-dependent degeneration of neurons occurred after oxLDL (1-100
microg/ml) treatment but was absent after native low density
lipoproteins (LDL). This degeneration was mediated, in part, by
apoptosis because increased TUNEL and Hoechst dye-positive staining
was observed. These effects occurred in the absence of Microglia.
However, DNA degradation was not detected. The cytotoxicity was
attenuated by pre-treatment with antioxidants. These results suggest
that oxidation by oxLDL may be important in neurocytotoxicity in the
brain
Wilms H, Hartmann D, Sievers J (1997) Ramification of
Microglia,
monocytes and macrophages in vitro: influences of various epithelial
and mesenchymal cells and their conditioned media. Cell Tissue Res.
287:447-458
Abstract: Microglial
cells are able to switch between an "active" amoeboid and
a ramified "resting" morphology during development and
after experiencing lesions. We have previously shown that in vitro
Microglial
morphology is controlled by their cellular environment, i. e. cells
become ramified in astrocyte coculture but amoeboid on monolayers of
fibroblasts. In the present study we have extended the analysis of
the control of macrophage morphology by maintaining macrophages of
different origins in coculture with different epithelial or
mesenchymal cells and their conditioned media. Microglia,
monocytes and spleen macrophages seeded onto monolayers of
astrocytes, kidney epithelia or hepatoma cells developed the
ramified morphology but remained amoeboid in fibroblast coculture.
Ramification was also induced by media conditioned by these cells as
well as by phorbolic esters, i.e. activators of protein kinase C. In
double coculture assays, even small numbers of fibroblasts were able
to override the "epithelial" influence. Likewise,
Microglia
remained amoeboid, when incubated on several constituents of the
extracellular matrix. These results indicate that macrophage
ramification is an active process initiated by diffusible factors
secreted by various epithelial cells, possibly acting upon a
protein-kinase-C-related receptor. We interprete the modification of
macrophage morphology as a functional adaptation to the surrounding
type of tissue that is enforced by its constituent cells. Thus, the
specific morphologies of Microglia,
hepatic von Kupffer's cells or peritubular kidney macrophages could
be explained by similar epithelium-macrophage interaction
Zachary JF, Baszler TV, French RA, Kelley KW (1997) Mouse
Moloney leukemia virus infects Microglia
but not neurons even though it induces motor neuron disease.
Mol.Psychiatry 2:104-106
Abstract: Motor neuron degeneration
caused by ts1 MoMuLV occurs by an indirect mechanism and
hypothetically appears associated with a two-cell or three-cell
pathogenesis hypothesis. The first step in this hypothesis is
associated with a small subset of resident Microglial
cells that serve as the principal target cells for ts1 MoMuLV
infection. The second step is likely linked to trophic events,
probably mediated by cytokines, that lead to hypertrophy and
activation of a substantial number of additional Microglial
cells (autocrine effect) and adjacent astrocytes (paracrine effect).
The third step in this hypothesis appears related to indirect
neuronal degeneration mediated by cytotoxins produced by activated
Microglial
cells and astrocytes. In this last step, motor neurons located
within these foci of activated Microglial
cells and astrocytes are 'innocent bystander cells' and degenerate
and die due to paracrine effects. The mechanism of motor neuron
degeneration is poorly understood but is likely linked to a
sequential cascade of trophic factors and cytokines resulting in a
final common pathway for motor neuron death involving production of
oxidative radicals, excitatory aminoacid neurotransmitter-like
substances, prostaglandins, or nitric oxide
Nam M, Johnston P, Lal B, Indurti R, Wilson MA, Laterra J
(1996) Endothelial cell-based cytokine gene delivery inhibits 9L
glioma growth in vivo. Brain Res. 731:161-170
Abstract: Malignant
brain neoplasms present great therapeutic challenges due to their
extremely aggressive behavior and relative isolation by the
blood-brain and blood-tumor
barriers. Endothelial cells may be versatile platforms for
delivering genes to solid tumors by virtue of their location at
blood-tissue interfaces and their proliferation in response to
endothelial mitogens produced by tumors. Immortalized rat brain
endothelial cells that express the E. coli lacZ reporter gene and
the gene for murine interleukin-2 (RBEZ-IL2) were co-inoculated with
9L glioma cells to Fisher rats to examine the effects of endothelial
cell-based cytokine delivery on glioma growth in vivo. 9L glioma
growth was not affected by the implantation of control RBEZ cells.
The growth of subcutaneous and intracranial 9L gliomas was
significantly inhibited by RBEZ-IL2 cells (P < 0.005 and P <
0.01, respectively) when compared to control transfected RBEZ cells.
Rats receiving intracranial 9L glioma cells with RBEZ-IL2 cells
showed increased survival (P < 0.001). Histologic and
immunohistologic analysis showed enhanced activation of
Microglia/macrophages
and CD8-positive T lymphocytes and/or natural killer cells within
brain at sites of 9L inoculation with RBEZ-IL2 cells. This report
establishes that immortalized endothelial cells can be used for
cytokine gene delivery and to activate anti-tumor
host responses to experimental gliomas within the central
nervous system
Personett DA, Chouinard M, Sugaya K, McKinney M (1996)
Simplified RT/PCR quantitation of gene transcripts in cultured
neuroblastoma (SN49) and Microglial
(BV-2) cells using capillary electrophoresis and laser-induced
fluorescence. J.Neurosci.Methods 65:77-91
Abstract: We developed
a simplified protocol for sensitive quantitation of mRNA using
polymerase chain reaction (PCR) amplification of cDNA made by
reverse transcriptase (RT), as resolved with capillary
electrophoresis (CE) and detected with laser-induced fluorescence
(LIF). The conditions required for adequate accuracy of the
simplified version of the RT/PCR quantitation, in which a single
concentration of external standard and amplification to within or
near the plateau phase are used, were established for assay of mRNAs
expressed at high, moderate, and low abundance. The mRNAs for the
cytosolic enzyme, glyceraldehyde phosphate dehydrogenase (GAPDH) and
the growth-associated protein GAP-43 in cultured SN49 neuroblastoma
cells were used as target genes for high and moderate levels of
expression, respectively. Using cultured mouse Microglial
cells (BV-2), we demonstrated the utility of this RT/PCR/CE/LIF
protocol to quantitate a low-abundance mRNA, encoding a form of
nitric oxide synthase (i-NOS) induced by treatment with endotoxin.
The appearance of i-NOS mRNA after endotoxin treatment of BV-2 cells
was confirmed by Northern blot analysis and in situ hybridization
histochemistry, and functional enzyme activity was followed by
release of nitric oxide (as nitrite) into the medium. The many
advantages of the 'single-point' RT/PCR/CE/LIF protocol for
quantitating mRNAs of interest include: simplified protocol,
elimination of the use of radiotracers, high sensitivity and
precision, and semi-automation of the quantitation phase of analysis
Petanceska S, Canoll P, Devi LA (1996) Expression of rat
cathepsin S in phagocytic cells. J.Biol.Chem.
271:4403-4409
Abstract: Cysteine lysosomal proteases are
essential for turnover of intracellular and extracellular proteins.
These enzymes are strongly implicated in normal and pathological
processes involving tissue remodeling. Among the cysteine proteases,
cathepsin S seems to be best suited for such a process since it
retains most of its enzymatic activity at neutral pH. In situ
hybridization analyses of the adult rat brain, spleen, and lung
reveal that cathepsin S mRNA is preferentially expressed in cells of
mononuclear-phagocytic origin. After entorhinal cortex lesion of
adult rat brain (a paradigm for neuronal degeneration and reactive
synaptogenesis), cathepsin S mRNA is dramatically increased in
activated Microglia
in the deafferented dentate gyrus and in macrophages at the wound
site, suggesting a role in lesion-induced tissue remodeling. This
possibility is further supported by the finding that cathepsin S
degrades a number of extracellular matrix molecules at neutral pH
and by the finding that inflammatory mediators stimulate its
secretion from the Microglia
and macrophages. These data suggest that cathepsin S is an important
player in degenerative disorders associated with the cells of the
mononuclear phagocytic system
Shrikant P, Benos DJ, Tang LP, Benveniste EN (1996) HIV
glycoprotein 120 enhances intercellular adhesion molecule-1 gene
expression in glial cells. Involvement of Janus kinase/signal
transducer and activator of transcription and protein kinase C
signaling pathways. J.Immunol. 156:1307-1314
Abstract: It is well
established that the two major glial cells in the central nervous
system (CNS), astrocytes and Microglia,
are key participants in mediating the neurologic dysfunction
associated with HIV infection of the CNS. In this study, we
investigated the ability of the major envelope glycoprotein of HIV,
glycoprotein 120 (gp120), to regulate intercellular adhesion
molecule-1 (ICAM-1) expression in glial cells, because ICAM-1 is
important in mediating immune responsiveness in the CNS,
facilitating entry of HIV-infected cells into the CNS, and promoting
syncytia formation. Our results indicate that gp120 enhances ICAM-1
gene expression in primary rat astrocytes, primary human astrocytes,
a human astroglioma cell line CRT, and primary rat Microglia.
The signal transduction events involved in gp120-mediated
enhancement of ICAM-1 appear to involve activation of both protein
kinase C and tyrosine kinase, because inhibitors of protein kinase C
and tyrosine kinase abrogate gp120-mediated ICAM-1 expression in
both astrocytes and Microglia.
Moreover, gp120 induces tyrosine phosphorylation of signal
transducer and activator of transcription (STAT-1 alpha) as well as
the Janus kinase (JAK2) in glial cells. We also demonstrate that
gp120-mediated ICAM-1 expression has functional significance, as it
enhances the ability of monocytic cells to bind to gp120-stimulated
human astrocytes in an ICAM-1/beta 2 integrin-dependent fashion.
These results provide new insights into how gp120 can influence the
involvement of glial cells in the pathogenesis of AIDS dementia
complex
Tanaka M, Sato A, Okada Y, Makino M, Tabira T (1996) Evidence
that brain capillary endothelial cells can be infected with murine
leukaemia retrovirus, LP-BM5. Neurodegeneration. 5:287-291
Abstract:
Some mice infected with murine leukaemia retrovirus, LP-BM5
including ecotropic, mink cell focus-inducing murine leukaemia
virus, and a replication-defective genome, have been reported to
show weakness, ataxia, or selective deficits in spatial learning
after developing an immunodeficiency syndrome similar to human AIDS.
In the central nervous system, astrocytes and Microglial
cells have been shown to be infected by this virus. We present here
findings that the ecotropic virus and defective genome can infect
murine brain capillary endothelial cells, and infected endothelial
cells show an impaired function as target cells against myelin basic
protein (MBP) specific T cell clone
Czub M, Czub S, Rappold M, Mazgareanu S, Schwender S, Demuth
M, Hein A, Dorries R (1995) Murine leukemia virus-induced
neurodegeneration of rats: enhancement of neuropathogenicity
correlates with enhanced viral tropism for macrophages, Microglia,
and brain vascular cells. Virology 214:239-244
Abstract: A highly
neuropathogenic retrovirus, NT40, was generated by serially
passaging an infectious molecular clone of Friend murine leukemia
virus, FB29, through F344 Fisher rats. NT40 induced severe
neurological signs such as reflex abnormalities and ataxia within
4-6 weeks following neonatal inoculation. FB29 led to only very mild
neurological dysfunctions with longer incubation periods.
Pathological alterations were characterized by mild (FB29) to
extensive (NT40) noninflammatory spongiform degeneration, mainly of
brain-stem areas. Infectious center assays revealed that viral
titers in brain tissues of NT40-infected rats were 100-fold higher
than those of FB29-infected animals. Employing immunohistochemistry,
in situ hybridization, and flow cytometry, NT40 was found to infect
many endothelial cells of brain blood vessels and Microglia,
whereas FB29 infected only Microglia
and those to a lower extent. However, when isolated from adult
diseased rats, Microglial
cells turned out in both cases to be nonproductively infected with
either FB29 or NT40. Of peripheral organs, we found enhanced levels
of NT40 in peritoneal macrophages but not in spleen, thymus, or
serum when compared to FB29. Altogether these data suggest that an
expanded cellular tropism within the CNS and elevated viral titers
in macrophages and Microglia
correlated with enhancement of neuropathogenicity
Dobrenis K, Makman MH, Stefano GB (1995) Occurrence of the
opiate alkaloid-selective mu3 receptor in mammalian Microglia,
astrocytes and Kupffer cells. Brain Res. 686:239-248
Abstract:
Evidence is presented for occurrence of opiate alkaloid-selective,
opioid-peptide-insensitive receptor binding sites, labeled with
[3H]morphine, in primary cultures of cat Microglia
and cat astrocytes, as well as on highly purified preparations of
rat Kupffer cells. These receptors have been designated mu3 on the
basis of their close similarity to receptors first found to be
present on human peripheral blood monocytes. Exposure of the
Microglia
to morphine and etorphine caused marked quantifiable changes in
cellular morphology, including assumption of a more rounded shape
and retraction of cytoplasmic processes; in contrast, several opioid
peptides were without effect on morphology. The effects of morphine
on Microglial
morphology were blocked by the opiate antagonist naloxone. These
effects of drugs on morphology were as predicted for action via the
mu3 receptor. Opiate alkaloid binding sites previously detected on
the rat C6 glioma cell line were also characterized here as of the
mu3 receptor subtype. It is proposed that mu3 receptors have broad
distribution in different macrophage cell types of bone marrow
lineage, including Microglia
and Kupffer cells. Furthermore, these receptors are not restricted
to cells of bone marrow lineage, since they are also present on
astrocytes
Dyck JR, Fliegel L (1995) Specific activation of the Na+/H+
exchanger gene during neuronal differentiation of embryonal
carcinoma cells. J.Biol.Chem. 270:10420-10427
Abstract: We
examined the regulation of the Na+/H+ exchanger gene during
differentiation of the P19 mouse embryonal carcinoma cells.
Treatment of P19 cells with retinoic acid induces the development of
neurons, astroglia, and Microglia
cells. Upon retinoic acid-induced differentiation of P19 cells,
there was an early and rapid 10-fold increase in NHE1 transcription.
A proximal cis-acting AP-2 site of the NHE1 promoter was sufficient
for stimulation of transcription of the gene by differentiation.
Bandshift experiments demonstrated that in retinoic acid-treated
cells there was an elevated level of AP-2 transcription factor
binding to the AP-2 consensus site of the Na+/H+ exchanger gene. In
the differentiation defective mutant RAC65, the effect of
differentiation on Na+/H+ exchanger gene expression was reduced by
60%. Examination of Na+/H+ exchanger activity showed that retinoic
acid-treated P19 cells recovered from an acid load at a rate
approximately three times greater than untreated cells. The
increases in gene expression and protein activity preceded major
changes in cell morphology, suggesting that the initiation of
differentiation is linked to NHE1 gene expression. Our findings show
for the first time that the NHE1 gene is activated early in cell
differentiation and that this activation may play an important role
in the process of neuronal cell differentiation
Gocht A, Lohler J, Scheidel P, Stegner HE, Saeger W (1995)
Gliomatosis peritonei combined with mature ovarian teratoma:
immunohistochemical observations. Pathol.Res.Pract.
191:1029-1035
Abstract: Gliomatosis peritonei (GP) is the
metastatic implantation of glial cells within the peritoneal cavity
of patients with ovarian teratomas. The case of a young woman is
presented, who initially developed a mature teratoma in the left
ovary that was surgically removed. Nine years later a mature
teratoma in the right ovary was excised, upon which GP was found in
the greater omentum. To identify the cellular composition of the
ovarian teratoma and of the omental implants, immunostainings were
performed using antibodies against glial and neuronal antigens as
well as against determinants of hematopoietic cells. In the teratoma
the neuroectodermal part was strongly HNK-1-positive and contained
Janabi N, Peudenier S, Heron B, Ng KH, Tardieu M (1995)
Establishment of human Microglial
cell lines after transfection of primary cultures of embryonic
Microglial
cells with the SV40 large T antigen. Neurosci.Lett.
195:105-108
Abstract: Four continuous cell lines of human
Microglial
cells were obtained by transfection of enriched cultures of human
embryonic brain-derived macrophages with a plasmid encoding for the
large T antigen of SV40. The transformed cells had the macrophagic
characteristics of adherence and intra-cytoplasmic non-specific
esterase activity. They could phagocytize zymosan particles but the
phagocytic activity remained low. They expressed several macrophagic
antigens but not the monocytic markers CD14, CD4, CD68/Ki-M6 and
CD11c. The cells could be activated to express class II major
histocompatibility complex antigens after interferon-gamma
activation. Finally, interleukin-6 was produced spontaneously by the
cells and this production was further increased after interleukin-1
alpha stimulation
Puy L, MacLusky NJ, Becker L, Karsan N, Trachtenberg J, Brown
TJ (1995) Immunocytochemical detection of androgen receptor in human
temporal cortex characterization and application of polyclonal
androgen receptor antibodies in frozen and paraffin-embedded
tissues. J.Steroid Biochem.Mol.Biol. 55:197-209
Abstract:
Immunocytochemical and biochemical studies have demonstrated the
presence of androgen receptor protein in various regions of the
rodent and non-human primate cortex. Localization of androgen
receptor in the human brain has, however, not been studied as
extensively, because of difficulties in obtaining suitable tissue
samples. In the present study, we have localized androgen receptors
in both frozen and paraffin-embedded temporal cortex from epileptic
patients undergoing resection. Polyclonal antibodies were raised
against fusion proteins containing fragments of the human androgen
receptor protein. The antibodies were affinity-purified against the
corresponding fusion protein. Immunoprecipitation and Western
blotting using extracts from human cell lines demonstrated the
specificity of the antibodies for the human androgen receptor and
lack of cross-reactivity with other steroid hormone receptors.
Immunocytochemistry was performed on frozen and paraffin sections of
human temporal cortex and in paraffin-embedded benign hyperplastic
prostates (BPH), as well as prostate and breast carcinomas, by the
streptavidin-biotin-peroxidase method. Antigen-retrieval was
performed in paraffin-embedded sections using microwave irradiation.
Specific nuclear and cytoplasmic immunoreactivity for androgen
receptor was detected in neurons, astrocytes, oligodendrocytes, and
Microglia
cells of the temporal cortex. In contrast, only nuclear staining was
observed in BPH, prostate and breast carcinomas. Immunoprecipitation
of human temporal cortex lysate and subsequent Western blot analysis
demonstrated the expression of a 98 kDa immunoreactive protein,
slightly smaller than the reported molecular weight of the wild-type
androgen receptor. These results provide further evidence for the
expression of androgen receptor in the human temporal cortex. The
use of these immunocytochemical techniques should enable the
retrospective determination of possible changes in androgen receptor
expression in a variety of archival paraffin-embedded tissues,
including samples of the human central nervous system
Viola JJ, Ram Z, Walbridge S, Oshiro EM, Trapnell B,
Tao-Cheng JH, Oldfield EH (1995) Adenovirally mediated gene transfer
into experimental solid brain tumors and leptomeningeal cancer
cells. J.Neurosurg. 82:70-76
Abstract: Among the appealing
features of adenoviruses as vectors for transfer of genes into the
central nervous system (CNS) are that they are not neurotoxic, they
can accommodate the insertion of several large genes, they are not
associated with the hazards of insertional mutagenesis, and they can
be concentrated to a high-titer preparation. The authors evaluated
the feasibility of using adenovirally mediated gene transfer into
cultured human glioma cells and in rat models of solid brain tumors
and meningeal cancer. Replication-deficient adenoviral vector
particles carrying a nuclear-localizing lacZ gene were injected into
established 9L cerebral gliomas in Fischer rats. In addition, the
adenoviral vector was injected into the subarachnoid space, either
simultaneously with intrathecal tumor
inoculation or after establishing leptomeningeal cancer. The
brains and spinal cords were removed at various intervals for
histochemical evaluation for beta-galactosidase activity using X-Gal
staining. Additional rats received a stereotactic intracerebral
injection of the vector into normal brain. No clinical abnormalities
were observed in the injected rats. Injection of the adenoviral
vector into normal brain resulted in diffuse transduction of
astrocytes, Microglia,
neurons, and endothelial cells at the injection site. Injection of a
high-concentration vector preparation into cerebral gliomas resulted
in effective tumor transduction.
Intrathecal injection of the vector in rats with meningeal cancer
resulted in transduction of the infiltrating tumor
in the subarachnoid space when injections were given
simultaneously with, or 7 days after, tumor
inoculation. Transduction rates of both solid and
leptomeningeal tumors correlated with the number of injected
particles. These results suggest that adenoviral vectors can
efficiently transduce solid brain tumors and that the vectors
survive in the cerebrospinal fluid for a sufficient period of time
to allow leptomeningeal tumor
transduction. Adenoviral vector should be evaluated for its
potential use in therapeutic gene transfer approaches in
malignancies of the CNS
Frei K, Malipiero U, Piani D, Fontana A (1994) Microglia and tumor rejection. Neuropathol.Appl.Neurobiol. 20:206-208
Kida S, Ellison DW, Steart PV, Iannotti F, Weller RO (1994)
Perivascular edema fluid pathway in astrocytic tumors. Acta
Neurochir.Suppl (Wien.) 60:384-386
Abstract: Perivascular spaces
are anatomical routes for the bulk flow drainage of fluid from the
gray matter to the subarachnoid space in normal rat brain.
Perivascular cells are the resident scavengers in perivascular
spaces. Following focal brain damage, perivascular cells upregulate
MHC Class II antigens associated with uptake of edema fluid. Similar
cells can be defined in damaged human brain. In the present
investigation, the distribution of MHC Class II upregulated
perivascular cells was measured in 30 astrocytic tumors and adjacent
edematous tissues by immunocytochemistry using the following
antibodies: HLA-DR (MHC Class II), PGM1 and MAC387 (macrophages).
Perivascular cells were PGM1+/MA
Kurpad SN, Wikstrand CJ, Bigner DD (1994) Immunobiology of malignant astrocytomas. Semin.Oncol. 21:149-161
Otero GC, Merrill JE (1994) Cytokine receptors on glial
cells. Glia 11:117-128
Abstract: Given what evidence there is for
the molecular and functional nature of cytokines and their cognate
binding proteins in the immune system and the emerging similarities
or even identities for these ligands and receptors in the nervous
system, two general models may be relevant. The first emerging
pattern is that receptors for related but distinct trophic factors
in the CNS are in many instances multichain complexes with one or
more shared components. The shared components of the receptor
complex may be either signal- or nonsignal-transducing chains. A
second emerging motif is that related ligands and related receptors
fall into gene families. Undoubtedly, these models will facilitate
the cloning of novel members of these families whose function is
quite specific to the nervous system and in particular to glial
cells. This article will review the function of the receptors for
cytokines and families of differentiation/survival/growth factors as
they operate on astrocytes, Microglia,
and oligodendrocytes in development, health, and disease
Vasilakos JP, Carroll RT, Emmerling MR, Doyle PD, Davis RE,
Kim KS, Shivers BD (1994) Interleukin-1 beta dissociates
beta-amyloid precursor protein and beta-amyloid peptide secretion.
FEBS Lett. 354:289-292
Abstract: A heightened production of
interleukin 1 beta (IL-1 beta) has been reported in
Microglial-associated
amyloid deposits in Alzheimer's disease (AD) brains. These plaques
are composed predominantly of beta/A4 peptide derived from
beta-amyloid precursor protein (beta APP). We demonstrate that
short-term (1 h) IL-1 beta-treatment increases beta APPs secretion
and concomitantly decreases cell-associated beta APP in human H4
neuroglioma cells. Long-term (5 h) IL-1 beta treatment did not alter
secreted or cell-associated beta APP content. In contrast, the
secretion of beta/A4-containing epitope was not affected by
short-term IL-1 beta stimulation; however, long-term IL-1 beta
treatment decreased the amount of beta/A4-containing epitope
secreted from the cells. These results show that IL-1 beta modifies
the processing and secretion of beta APP to exacerbate perhaps the
neuropathology of AD
Cupp C, Taylor JP, Khalili K, Amini S (1993) Evidence for
stimulation of the transforming growth factor beta 1 promoter by
HIV-1 Tat in cells derived from CNS. Oncogene 8:2231-2236
Abstract:
Infection by human immunodeficiency virus type 1 (HIV-1), the
etiologic agent of the acquired immunodeficiency syndrome (AIDS), is
often complicated with a high incidence of neurologic disorders. It
is believed that HIV-1, in addition to infecting both macroglial and
Microglial
cells, may influence the expression of several strategic genes of
uninfected neighboring or latently infected brain cells. It is
suspected that the viral-encoded transregulatory protein, Tat,
facilitates cross-communications between these cells. In support of
this concept, earlier studies demonstrated that Tat is released from
the infected cells, and has the capacity to be taken up by the
uninfected cells and exert its biological activity on the responsive
gene. Recent studies in several laboratories suggest the involvement
of Tat in altering the expression of a limited number of cellular
regulatory factors which, in turn, may mediate the altered
physiology of the cells. In this communication, we demonstrate the
ability of the HIV-1 Tat protein to increase expression of
transforming growth factor beta 1 (TGF-beta 1), a cytokine with
potent immunosuppressive activity, in human astrocytic glial cells.
Implications of the Tat-mediated induction of TGF-beta 1 expression
and cytokine involvement in the regulation of immune response and
central nervous system (CNS) pathology are discussed
Kuchelmeister K, Bergmann M, Gullotta F (1993) Cellular
changes in the cerebellar granular layer in AIDS-associated PML.
Neuropathol.Appl.Neurobiol. 19:398-401
Abstract: Six cases of
AIDS-associated progressive multifocal leukoencephalopathy (PML)
exhibited peculiar cellular changes in the cerebellar granular
layer. These cells without discernible cytoplasm showed
hypochromatic nuclei about twice as large as those of normal granule
cells. They were restricted exclusively to the granular layer and
always surrounded PML foci. An astrocytic, leukocytic or
macrophage/Microglial
nature was largely excluded by immunocytochemistry. Human
immunodeficiency virus (HIV) antigen p 24 could not be found in
these cells and there was no unequivocal detection of JC virus (JCV)
DNA and no ultrastructural evidence of papovavirus particles in
them. They possibly represent altered cerebellar granule cells
abortively or latently infected with JCV
Nagra RM, Wong PK, Wiley CA (1993) Expression of major
histocompatibility complex antigens and serum neutralizing antibody
in murine retroviral encephalitis. J.Neuropathol.Exp.Neurol.
52:163-173
Abstract: Murine leukemia virus infection serves as a
model for noninflammatory degeneration of the central nervous system
(CNS). During the course of infection with either of the molecularly
cloned viruses pNE-8 or ts-1, we observed that ts-1 spread twice as
rapidly as pNE-8, and ascended higher in the neuraxis. Endothelial
cells were infected first, followed by oligodendrocytes and neurons,
while astrocytes containing glial fibrillary acidic protein were not
infected. Additionally, ts-1 also infected macrophages/Microglia.
Major histocompatibility complex (MHC) class I beta 2-microglobulin
expression was minimal in pNE-8 infected mice, while it was elevated
in endothelial cells of early ts-1 lesions, and in
macrophages/Microglia
during later stages. Occasional infected cells expressed beta
2-microglobulin while rare endothelial and parenchymal cells
expressed MHC class II in both viral infections. Limited intra-CNS
MHC expression may be one of the mechanisms of viral persistence and
will present a barrier to developing immunotherapy for CNS
retroviral infections. The few mice that escaped lethal infection
had higher serum titers of neutralizing antibodies and showed no
neuropathologic changes or detectable virus in the CNS. Higher
titers of neutralizing antibodies may protect the CNS from infection
Constam DB, Philipp J, Malipiero UV, ten Dijke P, Schachner
M, Fontana A (1992) Differential expression of transforming growth
factor-beta 1, -beta 2, and -beta 3 by glioblastoma cells,
astrocytes, and Microglia.
J.Immunol. 148:1404-1410
Abstract: The type beta transforming
growth factors (TGF) are potent regulators of the growth and
functions of lymphocytes and macrophages. Recently the human
glioblastoma cell line 308 was shown to produce TGF-beta 2. The
relevance of this finding was evaluated further by comparing human
glioblastoma cells with their nontransformed animal counterpart,
astrocytes, with regard to the production of the three TGF-beta
isoforms observed so far in mammals. In this report astrocytes are
demonstrated to secrete also TGF-beta 2 and to express TGF-beta 1,
-beta 2, and -beta 3 mRNA in vitro. In contrast, cultured murine
brain macrophages release TGF-beta 1 and are positive for TGF-beta 1
mRNA only. Glia cell-derived TGF-beta 1 and -beta 2 are detected in
latent form whereas both latent and active TGF-beta are identified
in the supernatant of three human glioblastoma cell lines tested.
These cell lines, however, show heterogeneity in regard to the
isoform of TGF-beta expressed but share with astrocytes the
inability to release TGF-beta 3. Provided production and activation
of latent TGF-beta occur in vivo, astrocytes and Microglia
may then be expected to exert regulatory influences on immune
mediated diseases of the central nervous system
Frei K, Piani D, Malipiero UV, Van Meir E, de Tribolet N,
Fontana A (1992) Granulocyte-macrophage colony-stimulating factor
(GM-CSF) production by glioblastoma cells. Despite the presence of
inducing signals GM-CSF is not expressed in vivo. J.Immunol.
148:3140-3146
Abstract: One of the morphologic hallmarks of human
gliomas are inflammatory infiltrates with accumulation of
macrophages in the tumor site.
The signals leading to the macrophage response are only at the
beginning of being understood. Novel chemotactic factors that have
recently been characterized as secretory products of glioblastoma
cells may attract mononuclear cells from the blood. Within the tumor
tissue blood-derived monocytes and macrophages of the brain
tissue, the Microglial
cells, may increase in cell numbers due to tumor-derived growth
factors. Both astrocytoma cell lines and cultured astrocytes have
been shown recently to produce granulocyte-macrophage (GM)-CSF. We
show that in vitro not only astrocytoma but also glioblastoma cell
lines secrete GM-CSF when stimulated with TNF-alpha or IL-1.
However, there is no evidence for GM-CSF production by glioblastoma
cells in vivo: fresh tumor samples
lack the mRNA for GM-CSF and the protein is not detectable in the
tumor cyst fluids or the
cerebrospinal fluids of glioblastoma patients. This contrasts IL-1
and IL-6 that are detectable in the tumor
cyst fluids and IL-6 also in the cerebrospinal fluids of the
patients. Unlike GM-CSF, transforming growth factor-beta 2 mRNA is
expressed in ex vivo tested glioblastoma tissues. Absence of GM-CSF
in vivo may be explained by the presence of tumor-derived inhibitory
factors, such as transforming growth factor-beta 2 and PGE which
suppress GM-CSF production by glioblastoma cells in vitro. The
accumulation of macrophages at the tumor
site may be due to local elaboration of chemoattractants
and/or not yet defined growth factors rather than due to GM-CSF
production
Simmons ML, Murphy S (1992) Induction of nitric oxide
synthase in glial cells. J.Neurochem. 59:897-905
Abstract:
Primary astrocyte cultures, C6 glioma cells, and N18 neuroblastoma
cells were assayed for nitric oxide synthase (NOS) activity with a
bioassay of cyclic GMP production in RFL-6 fibroblasts. Treatment of
astrocyte cultures for 16-18 h with lipopolysaccharide (LPS) induced
NOS-like activity that was L-arginine and NADPH dependent, Ca2+
independent, and potentiated by superoxide dismutase. Induction was
evident after 4 h, was dependent on the dose of LPS, and required
protein synthesis. Treatment of astrocyte cultures with leucine
methyl ester reduced Microglial
cell contamination from 7 to 1%, with a loss of 44% of NOS-like
activity. C6 cells treated with LPS also showed Ca(2+)-independent
and L-arginine-dependent NOS-like activity. N18 cells demonstrated
constitutive Ca(2+)-dependent NOS-like activity that was not
enhanced by LPS induction. These data indicate that NOS-like
activity can be induced in Microglia,
astrocytes, and a related glioma cell line as it can in numerous
other cell types, but not in neuron-like N18 cells
Achim CL, Schrier RD, Wiley CA (1991) Immunopathogenesis of
HIV encephalitis. Brain Pathol. 1:177-184
Abstract: HIV infection
leads to severe immunosuppression and in a sub-population of
patients, encephalitis. Whether systemic immunosuppression is
required for CNS infection is still unclear. However, latent
infection of monocytes/macrophages is an important mechanism by
which HIV escapes immune surveillance and enters the CNS. Unlike
other viral encephalitides, HIV predominantly infects
macrophages/Microglia
and not neurons and glia. These cells produce retroviral proteins
and cytokines which may be neurotoxic. Despite significant MHC
expression within the CNS, there is a limited infiltration of immune
cells, possibly due to a defect in systemic immunity.
Anti-retroviral therapy by decreasing viral replication and
reversing immunosuppression, may arrest nervous system damage
Jung HW, Berens ME, Krouwer HG, Rosenblum ML (1991) A
three-dimensional micro-organ culture system optimized for in vitro
growth of human malignant brain tumors. Neurosurgery
29:390-398
Abstract: A brain tumor
is composed not only of tumor
cells, but also of normal glial, mesenchymal, endothelial,
and Microglial
cells, as well as lymphocytes and macrophages. Therefore,
homogeneous cultures of tumor
cells, currently used for chemosensitivity testing, do not
accurately model in situ tumors. We have developed an in vitro
growth assay for brain tumors that includes normal host cells and is
potentially useful for studies of chemotherapy and biological
response modifiers. Human glioblastoma xenografts (U251-MG) were
resected from mice, minced, and explanted into agarose-coated
culture wells. After 5 to 7 days, microtumors emerged as expanding
spheroids, which grew most efficiently in minimum essential medium
supplemented with 20% fetal calf serum, 90% of which was replaced on
alternate days. The growth rate and bromodeoxyuridine labeling index
were similar in the microtumors and the xenografts, and light
microscopy revealed highly cellular, pleomorphic tumors with high
mitotic activity in both. Immunohistochemical studies also
demonstrated the persistence of macrophages in both xenografts and
microtumors. Microtumors treated for 2 hours with 75 mumol/L
1,3-bis-(2-chloroethyl)-1-nitrosourea showed a growth delay of 1.5
days; no effects were observed after treatment with lower doses.
This in vitro system for brain tumor
culture may provide a useful technique for the study of new
therapies as an alternative to in vivo xenograft studies using
immunodeficient animals
Sutter A, Hekmat A, Luckenbach GA (1991) Antibody-mediated
tumor cytotoxicity of
Microglia.
Pathobiology 59:254-258
Abstract: The status of Microglial
cells as potent effector cells in antibody-mediated tumor
cytotoxicity (ADCC) could be established. Microglia
(greater than or equal to 99.9% pure) derived from brain cortices of
newborn mice were shown to lyse human tumor
cell lines expressing different levels of epidermal growth
factor (EGF) receptors in the presence of MAb 425, a monoclonal
murine anti-primate EGF receptor antibody. MAb 425 mediates
Microglial
ADCC (MiADCC) at concentrations as low as 10(-11) M. Antibody
ligands binding unilaterally to either EGF receptors on target cells
or Fc receptors on Microglia
have little effect on MiADCC. At 10(-10) M MAb 425, a 10(3)-fold
excess of MAb 425 F(ab')2 fragments or irrelevant antibodies of
identical isotype did not block MAb-425-induced MiADCC. Formation of
effector-target cell contacts seems to be critical for MiADCC and
MiADCC could not be inhibited by anti-tumor
necrosis factor-alpha antibodies. In addition to its
stimulatory effect on MiADCC, MAb 425 bound to EGF receptors exerted
a microgliotrophic effect. Factor(s) derived from astrocytes enhance
MiADCC
Dozic S, Suvakovic V, Cvetkovic D, Jevtovic D, Skender M
(1990) Neoplastic angioendotheliomatosis (NAE) of the CNS in a
patient with AIDS subacute encephalitis, diffuse leukoencephalopathy
and meningo-cerebral cryptococcosis. Clin.Neuropathol.
9:284-289
Abstract: A 12-year-old, hemophilic boy died with
acquired immune deficiency syndrome (AIDS) after a clinical course
characterized by progressive psycho-organic syndrome and
opportunistic infections. Postmortem neuropathological examination
revealed a cerebral form of neoplastic angioendotheliomatosis (NAE),
leukoencephalopathy, giant cell encephalitis and meningo-cerebral
cryptococcosis. The most unusual finding was the presence of
proliferated neoplastic cells within lumina of some blood vessels
throughout the central nervous system (CNS). These cells displayed
cytologic features of malignancy and stained positively for common
leukocyte antigen. Coronal sections showed diffuse cerebral and
cerebellar leukoencephalopathy with most pronounced loss of myelin
and axons in deep white matter, while the subcortical arcuate fibers
and the corpus callosum were partially spared. In these areas
numerous small foci of severe myelin loss were present. Microglial
nodules and distinctive multinucleated giant cells (MGC) were
numerous. Intracytoplasmic and intranuclear acidophilic inclusions
were found in a few multinuclear and mononuclear cells. Close
contact between mononuclear and multinuclear cells suggesting their
fusion was also observed. As far as we know this is the first case
of NAE encountered in AIDS, one of the rare primary cerebral forms
and the youngest reported case of NAE up to now. This case could be
considered as one proof more that NAE is a special form of malignant
lymphoma
Vazeux R, Cumont M, Girard PM, Nassif X, Trotot P, Marche C,
Matthiessen L, Vedrenne C, Mikol J, Henin D, . (1990) Severe
encephalitis resulting from coinfections with HIV and JC virus.
Neurology 40:944-948
Abstract: We observed 3 cases of progressive
multifocal leukoencephalopathy (PML) among frozen CNS samples
obtained at autopsy from 102 adult AIDS patients. In 2 patients, PML
was associated with severe HIV encephalitis. In those 2 cases, the
areas of extensive JC-induced demyelination were massively
infiltrated by HIV infected macrophages/Microglial
cells with evidence for localized increase of HIV encephalitis in
PML lesions. Using immunohistochemistry and in situ hybridization,
we demonstrated that each virus infects, in a latent or productive
fashion, different CNS cell populations. Therefore, the extension of
HIV encephalitis could not be related to an intracellular
transactivation of 1 virus by the other. However, the results are
consistent with dissemination of viral infection by the recruitment
of HIV-infected macrophages to damaged areas of the brain. This
phenomenon might be generalized to other pathogens that are
frequently associated with HIV CNS infection. Early detection and
treatment of opportunistic CNS lesions could be important to prevent
extension of HIV encephalitis