On the presence of Ia-positive endothelial cells and astrocytes in multiple sclerosis lesions and its relevance to antigen presentation

Using a monoclonal antibody in combination with the 4-step modified peroxidase-antiperoxidase (PAP) technique, Ia expression was demonstrated on endothelial cells and astrocytes in MS lesions of different ages. On endothelial cells, Ia antigen was found most frequently in grey and white matter parenchyma of acute MS brain and showed lower, comparable numbers in active and silent chronic MS brain tissue. Ia+ astrocytes were most numerous in acute MS lesions. In active chronic MS, Ia+ astrocytes predominated at the lesion edge, where they frequently displayed an atypical, rounded configuration. Positive astrocytes showed somewhat lower numbers within the lesion center and in normal white matter close to the lesion edge. Their frequency was significantly lower in normal white matter remote from the lesion. In silent chronic MS, Ia+ astrocyte processes were detectable only within the lesion center. Grey matter astrocytes displayed no staining with anti-Ia antibody. In normal brain tissue, no Ia antigen could be detected. The presence of Ia molecules on some endothelial cells and astrocytes in MS brain tissue suggests a role in antigen presentation perhaps relevant to the initiation and perpetuation, respectively, of the inflammatory process.     

Monoclonal antibody, KK1, recognizes human retinal astrocytes and distinguishes a subtype of astrocytes in mouse brain

Astrocytes exhibit a diverse morphology and numerous functions in the central nervous system as well as in the retina. In order to obtain markers for the analysis of astrocytes, we prepared monoclonal antibodies that recognized antigens specific to astrocytes. Monoclonal antibody (mAb), designated KK1, reacted with the processes of astrocytes in the nerve fiber layer and the ganglion cell layer in the human retina as detected by indirect immunofluorescence. Normal Müller cells, whose processes are localized vertically in retina, were not labeled by KK1 mAb. In mouse brain, KK1 mAb reacted specifically with astrocytes in the white matter, but not with those in the gray matter. Studies employing a high-resolution confocal laser scanning microscope and double-labeling with KK1 mAb and commercially available anti-glial fibrillary acidic protein (GFAP) mAb (GA5) revealed that KK1 mAb visualized the processes that were not recognized by anti-GFAP rnAb (GA5) in both human retina and mouse brain. In cultured mouse astrocytes. KKI mAb reacted only with anti-GFAP mAb (GA5)-positive cells, but a small percentage of anti-GFAP mAb (GA5)-positive cells were labeled with KK1 mAb. In addition, the subcellular distribution of the KK1 antigen in cultured astrocytes apparently differed from that of GFAP labeled by anti-GFAP mAb (GA5). The antigen that was purified from the normal mouse brain by KK1 mAb-conjugated beads reacted with anti-GFAP mAb(GA5) in immunoblotting. No reactivity of KK1 mAb was observed in immunohistochemical analysis in GFAP − / − mutant mouse brain. These results demonstrate that KK1 mAb specifically recognized an epitope of GFAP that did not react with other anti-GFAP mAb (GA5). Retinal astrocytes and a subtype of astrocytes in the white matter of mouse brain shared the epitope that was recognized by KKI mAb. KKI mAb might be a powerful tool to investigate a subtype of astrocytes.     

A monoclonal antibody that binds to both astrocytes and myelin sheaths

A monoclonal antibody designated III 5H8 was shown to bind both to astrocytes and to myelin sheaths as studied with immunocytochemical techniques on brain sections and cell cultures. Binding to astrocytes was confirmed by double immunofluorescent labelling of frozen sections and cell cultures with anti-GFAP, and appeared to be sensitive to formalin treatment. Binding to myelin sheaths was confirmed by comparing sections labelled with III 5H8 with sections labelled with antibodies against axons and myelin basic protein as well as by staining of sections of hypomyelinated spinal cord with III 5H8. On immunoblots of separated white matter III 5H8 revealed two bands, while on immunoblots of purified myelin only one band was seen. The findings are discussed with respect to the function of astrocytes in white matter and shared antigenic determinants between astrocytes and oligodendrocytes.     

Enhanced expression of a protein antigen (J1-31 antigen, 30 kilodaltons) by reactive astrocytes in lacerated spinal cord

A mouse monoclonal antibody (MAb J1-31, isotype IgG 2b) was raised against an autopsy sample of cerebral white matter from a multiple sclerosis (MS) patient. MAb J1-31 recognizes a protein (J1-31 antigen) in human brain which has a molecular weight of approximately 30,000 daltons (30 kD) as determined by immunoprecipitation followed by SDS-gel electrophoresis (reducing conditions) and autoradiography (Singh et al.: Biosci Rep 6:73-79, 1986). By immunofluorescence microscopy, MAb J1-31 stains glial fibrillary acidic protein (GFAP)-positive cells, namely astrocytes, of both human and rat. Yet J1-31 antigen is distinct from GFAP (Predy et al.: Biosci Rep 7:491-502, 1987). In this paper we report that greatly enhanced staining for J1-31 antigen is exhibited by reactive astrocytes which arise following CNS injury. (Laceration-type surgical lesion of the rat spinal cord served as the experimental model). Enhanced expression of J1-31 antigen reveals some new aspect of the astrocyte response to injury, involving transformation to the reactive state. Consequently, MAb J1-31 may be a useful marker for studies on reactive astrocytes.     

Novel astrocytic protein in multiple sclerosis plaques

Monoclonal antibody J1-31 (MAb J1-31, isotype IgG 2b) was raised against crude homogenate of brain tissue from a multiple sclerosis (MS) patient (autopsy sample; Malhotra et al.: Microbios Letters 26:151-157, 1984). In human brain, MAb J1-31 recognizes an intracellular protein antigen (J1-31 antigen), which bands at approximately 30,000 daltons under reducing conditions for sodium dodecyl sulfate gel electrophoresis (Singh et al.: Bioscience Reports 6:73-79, 1986). By immunofluorescence microscopy, MAb J1-31 stains those cells that are also stained by antiserum to glial fibrillary acidic protein (GFAP), namely astrocytes, retinal Müller cells, and tanycytes in the ependyma (Predy et al.: Bioscience Reports 7:491-502, 1987). In addition, MAb J1-31 stains ciliated ependymal cells that do not express GFAP. Using a model system for gliosis (laceration-type injury of rat spinal cord), we were able to show that astrocytes responding to central nervous system injury exhibit greatly enhanced staining for J1-31 antigen (Predy et al.: Journal of Neuroscience Research 19:397-404, 1988; Predy and Malhotra: Brain Research Bulletin in press, 1989). In this article, we demonstrate that immunofluorescence staining owing to MAb J1-31 is greatly enhanced in MS plaques, as compared to adjacent "apparently normal" white matter. (This is consistent with previous results as MS plaques characteristically show an astroglial response [reactive gliosis] leading to the formation of a glial scar [McKhann: Annual Review of Neuroscience 5:219-239, 1982].) In addition, we present further evidence that J1-31 antigen is distinct from GFAP, although these two proteins may be associated spatially with one another.     

Localization of CD44 (P80) on the external surface of a human astrocytoma cell

A brain antigen, originally identified by a MAb 44D10, has been shown to be a glycoprotein with an Mr of 80 kDa. Cellular localization studies of sections of brain showed that the antigen was associated with the membrane of astrocytes. In the present study we demonstrate its localization to the membrane of an astrocytoma cell line by fluorescence and electron microscopic immunogold methods. Heavy labelling with immunogold was found along cellular processes. Labelling of the smooth surfaces of cells and the microvilli was also observed.     

Peroxiredoxin V in multiple sclerosis lesions: predominant expression by astrocytes

Oxidative stress is implicated in the pathogenesis of multiple sclerosis (MS). Defence against oxidative damage is mediated by antioxidants. Peroxiredoxin V (PRDX V) is an intracellular anti-oxidant enzyme with peroxynitrite reductase activity. It is increased during inflammation, when free radical production intensifies, and is protective in an animal model of brain injury. However, little is known about PRDX V expression in the human brain. We investigated PRDX V expression in white matter from normal human brain (n = 5) and MS patients (n = 18), using immunohistochemistry and immunoblotting. A global increase in PRDX V was evident in MS normal-appearing white matter (NAWM) but the most striking increase was in astrocytes in MS lesions. PRDX V- positive hypertrophic reactive astrocytes were seen in acute lesions where inflammation was present. Yet surprisingly, in chronic lesions (CL), where inflammation has abated and a glial scar formed, there was strong PRDX V staining of post-reactive, scar astrocytes. Furthermore, immunoblotting analysis of tissue from two MS cases confirmed a substantial increase in PRDX V expression in CL compared with NAWM from the same individual. This might indicate ongoing oxidative stress despite the absence of histologically defined inflammation. Further investigations of this phenomenon will be of interest for therapeutic targeting.     

Disappearance of beta2-adrenergic receptors on astrocytes in canine distemper encephalitis: possible implications for the pathogenesis of multiple sclerosis

It has been reported that astrocytes in the white matter of patients with multiple sclerosis (MS) lack beta2-adrenergic receptors. This abnormality might explain why astrocytes in active MS plaques aberrantly express major histocompatibility (MHC) class II molecules, which play an important role in the immunological cascade leading to myelin destruction. Canine distemper (CD) virus primarily infects astrocytes and causes a demyelinating disease in dogs that closely resembles MS. In control dogs, including three dogs with another inflammatory disease, beta2-adrenergic receptor immunoreactivity was observed on both neurons and astrocytes. In dogs with CD encephalitis, beta2-adrenergic receptors were present on neurons, but were absent on astrocytes in acute lesions, demyelinated lesions, and normal-appearing white matter. Similar to MS, several astrocytes in demyelinated lesions expressed MHC class II. These findings suggest that MS and the demyelinating stages of CD encephalitis have a common pathogenetic factor, and that the loss of astrocytic beta2-adrenergic receptors in MS might be induced by a viral infection of astrocytes.     

Angiotensin receptor-like immunoreactivity in adult brain white matter astrocytes and oligodendrocytes.

Most of the physiological effects of brain angiotensins are currently believed to be mediated by angiotensin receptors located principally on neurons. However, numerous studies in vitro have demonstrated the presence of functional angiotensin receptors on brain astrocytes, raising the possibility that glial cells may also participate in mediating the effects of the central renin-angiotensin system. Nevertheless, it is uncertain whether these cells in situ express angiotensin receptors, raising questions about the physiological significance of results observed in cell cultures. We have examined the distribution of angiotensin receptor-like immunoreactivity in glial cells in white matter tracts in the adult CNS, using a panel of antisera to the AT1 and AT2 angiotensin receptors. Antiserum preadsorption and/or Western blot demonstrated the specificity of the antisera in brain tissue. In immunohistochemical experiments, the AT1 antisera selectively labeled AT1-expressing neurons in the piriform cortex, whereas the AT2 antiserum stained cells in the trigeminal motor nucleus, these being nuclei known to express AT1 and AT2 receptors, respectively. Using double-label immunohistochemistry, we observed AT1- and AT2-immunoreactive astrocytes and oligodendrocytes in white matter tracts, which include the rat cerebellar white matter, periventricular white matter, and optic nerve, in addition to the bovine corpus callosum and human subcortical white matter. In contrast, astrocytes in the gray matter region of the cerebral cortex were not found to be angiotensin receptor-like immunoreactive. These results demonstrate the presence of AT1 and/or AT2 angiotensin receptor-like immunoreactivity in brain white matter macroglial cells in situ and support the idea that glial cells may play a more important role in the central renin-angiotensin system than previously thought.     

Lipocalin-type prostaglandin D synthase (beta-trace) is upregulated in the alphaB-crystallin-positive oligodendrocytes and astrocytes in the chronic multiple sclerosis

Lipocalin-type prostaglandin D synthase (L-PGDS), which is mainly synthesized in leptomeningeal cells and oligodendrocytes (OLs) in rodents and humans, is secreted into the human cerebrospinal fluid (CSF) as beta-trace. L-PGDS protects OLs and neurones against apoptosis in twitcher mice, a murine model of Krabbe's disease, and is the second only to a stress protein, alphaB-crystallin, as the most abundant gene product upregulated in the demyelinating focus of multiple sclerosis (MS). Here we report that although the CSF level of L-PGDS is not increased in MS patients, L-PGDS is increased in the white matter of MS patients, especially in the shadow plaque as compared with the normal white matter. L-PGDS immunoreactivity was intensely expressed in OLs within the shadow plaques and in hypertrophied astrocytes within the chronic plaques of MS patients. Both L-PGDS-positive OLs and astrocytes expressed a stress protein, alphaB-crystallin. These results suggest that the upregulation of L-PGDS occurs in OLs and astrocytes as a stress reaction.     

Histological localization of nervous-system antigens in the cerebellum by immunoperoxidase labeling

The indirect immunoperoxidase method was used to localize histologically on sagittal sections of mouse cerebellum antigenic determinants detected by the following antisera: anti-NS-2, anti-NS-3, anti-NS-4, rabbit anti-bovine corpus callosum, rabbit anti-mouse brain, rabbit anti-glial fibrillary acidic protein, and rabbit anti-neurofilament protein. Anti-α-bungarotoxin serum and normal rabbit serum were used as negative controls. The various sera showed similarities in staining pattern as well as differences. Anti-NS-2 antiserum labeled the somata of interneurons in the molecular layer, granule cell bodies, glial cells in the white matter, and along the surfaces of blood vessels. A similar pattern of staining is produced by the anti-NS-3 antiserum except that glial cells are less prominent in the white matter and the blood vessels are not visible at all. Anti-NS-4 antiserum does not label interneurons but does label glomeruli and, less intensely, granule cell bodies in the granular layer. Rabbit anti-mouse brain antiserum is similar to anti-NS-4 antiserum except that fiber tracts in the white matter are stained more intensely. Rabbit anti-bovine corpus callosum labels only white matter. Antisera to neurofilament and glial fibrillary acidic proteins label Bergmann glia and fibrous astrocytes.     

Antibody against mouse liver 5'-nucleotidase immunostains white matter in the adult mouse central nervous system

An antiserum against rat liver 5'-nucleotidase has been shown to immunostain myelinated fibers and oligodendrocytes in the rat CNS, consistent with evidence for 5'-nucleotidase activity in rat brain myelin and oligodendrocytes (Cammer, Sacchi and Kahn, Devel. Brain Res., 1985, 20: 89-96). However, in the mouse CNS, in which myelin also has 5'-nucleotidase activity, that antiserum stained only blood vessels. To obtain an antibody against the mouse enzyme, 5'-nucleotidase was partly purified from mouse liver membranes by detergent extraction, heat treatment, affinity chromatography, acidification, and ammonium sulfate fractionation. The preparation, which was enriched about 110-fold in 5'-nucleotidase specific activity, compared to the starting extract, was electrophoresed on a preparative slab gel containing Triton X-100, a strip was stained histochemically for 5'-nucleotidase, and the material corresponding to the stained band was used to immunize a rabbit. The immune IgG fraction, but not the preimmune IgG, reacted with mouse brain homogenates. The immune serum gave consistently greater inhibition of 5'-nucleotidase activity in mouse liver homogenates, mouse brain myelin and mouse brain homogenates, but not rat brain or liver homogenates, compared to the preimmune serum. The immune serum, but not the preimmune serum, immunostained white matter in the normal adult mouse brain and spinal cord. The findings suggest that the mouse may have one isozyme of 5'-nucleotidase similar to that in rat with respect to subunit sizes but differing in primary structure at one or more antigenic sites and support previous observations of 5'-nucleotidase activity in myelin from mouse brains and spinal cords.     

Monoclonal antibody to intermediate filament proteins in astrocytes

A monoclonal antibody was developed using rat astrocytes purified in vitro as the starting antigenic material. Selection of the monoclonal was on the basis of astrocyte binding specificity in brain sections using indirect immunofluorescence techniques. The antibody (RBA2) that was chosen was specific for astrocytes in that it did not stain neurons or oligodendrocytes in frozen brain sections. It did, however, show binding to vascular smooth muscle and meningeal cells. The antigenic determinant(s) was determined to be on filaments of the intermediate-size class in cultured astrocytes and fibroblasts. From analysis of binding patterns in various tissues and in immunoblots, it was found that RBA2 cross-reacted strongly with glial fibrillary acidic protein (GFAP) and desmin. There was a weaker cross-reactivity to a vimentin-associated component. It is proposed that this antibody can be used as an astrocyte and blood vessel marker in brain sections, a vimentin marker in cultures and as a probe of intermediate filament composition and distribution.     

The human blood-brain barrier glucose transporter (GLUT1) is a glucose transporter of gray matter astrocytes

Human and monkey brain sections were examined by immunohistochemical light and electron microscopy to determine the distribution of GLUT1, a glucose transporter isoform associated with erythrocytes and endothelial cells of the human blood-brain barrier. Protein immunoblotting of fractionated human brain membranes was performed to determine the distribution of molecular forms of the transporter. GLUT1 staining was abundant in erythrocytes and cerebral endothelium of gray and white matter but was also present diffusely in gray matter neuropil when viewed by light microscopy. Immunoelectron microscopy confirmed the gray matter and vascular localization of GLUT1, with specific GLUT1 staining seen in erythrocytes, gray and white matter endothelial cells, astrocyte foot processes surrounding gray matter blood vessels, and in astrocyte processes adjacent to synaptic contacts. No astrocytic staining was identified in white matter. Astrocyte GLUT1 staining was identified only in mature gray matter regions; undifferentiated regions of preterm (22–23 weeks gestation) cortex had GLUT1 staining only in blood vessels and erythrocytes, as did germinal matrix. Immunoblots of adult human frontal cortex revealed that two forms of GLUT1 (45 and 52 kDa) were present in unfractionated brain homogenates. Immunoblots of vessel-depleted frontal lobe revealed only the 45 kDa form in gray matter fractions, and depleted in membranes prepared from white matter regions. We conclude that the GLUT1 isoform of glucose transporter is present both in endothelium of the blood-brain barrier and in astrocytes surrounding gray matter blood vessels and synapses. Furthermore, the form present in astrocytes is likely to have a lower molecular weight than the form found in cerebral endothelium. The GLUT1 transporter may play an important role not only in astrocyte metabolism, but also in astrocyte-associated pathways supporting neuronal energy metabolism. © 1995 Wiley-Liss, Inc.     

Comparison of immunocytochemical staining of astrocytes, oligodendrocytes, and myelinated fibers in the brains of carbonic anhydrase II-deficient mice and normal littermates.

Immunostaining for carbonic anhydrase (CA) was performed in paraffin sections from the brains of CA II-deficient mutant mice and their normal littermates. Double immunofluorescence staining showed CA in myelinated tracts and oligodendrocytes in the cerebellum of the normal but not the CA II-deficient mice, and also in astrocytes in the cerebral cortex of the normal mice but not the mutants. The data show that the CA in normal oligodendrocytes, astrocytes, and myelin is the II isoenzyme, because these structures in the mutants would be positively stained if the staining normally were due to a contaminant in the antiserum or an antibody against a different isoenzyme. The findings in normal gray matter also suggest that many neuronal cell bodies are surrounded by a network of fine, CA-positive astrocytic processes.     

Three-dimensional distribution of astrocytes in zebrafish spinal cord.

We prepared a monoclonal antibody (A-22) that recognizes a 60-kDa protein in the zebrafish brain. The antigen is distributed throughout the brain but is not found outside it. The antibody recognizes star-shaped cells with long processes in the spinal cord. All A-22-positive cells are also GFAP-immunopositive, but there are GFAP-positive cells that are A-22-negative. The cells are connected to small veins and to the surface of the spinal cord. Immunopositive cells are generally homogeneous in size and shape and are found not only in the spinal cord but also in several areas of the brain. These results indicate that the stained cell is an astrocyte. Most of these cells (88%) are distributed in the gray matter of the spinal cord; the remainder (12%) are found in the white matter. Most of the cells in the gray matter are found in the ventral and dorsal horns, but some are also present in the central area along the ventricle. Glial cell bodies form an array along the longitudinal axis and are connected to each other by thick projections. The cellular array is not visible in coronal sections. In contrast, thin processes from the cells extend to the surfaces of veins, to neurons, and to the periphery of the spinal cord. We estimate that there are about 13,500 A-22-positive astrocytes in the spinal cord; however, this represents only 26% of the total number of astrocytes in the spinal cord (approximately 52,000).     

Glutamine synthetase in the central nervous system is not confined to astrocytes.

Glutamine synthetase (GS) immunoreactivity is frequently used as an astroglial 'marker'. However, when sections of adult rat spinal cords were immunostained with antibodies against sheep glutamine synthetase, intense immunofluorescence was observed in cells resembling oligodendrocytes. In white matter in the rat brain GS immunostaining was also found in ovoid oligodendrocyte-like cells, whereas in gray matter in the same tissue sections GS immunostaining was found in astrocytes. Like the antibodies against sheep GS, antibodies against rat GS also immunostained putative oligodendrocytes, and colocalization with 2',3'-cyclic nucleotide-3'-phosphohydrolase in spinal cord supported the designation of the GS-positive cells as oligodendrocytes.     

Apoptosis of astrocytes with enhanced lysosomal activity and oligodendrocytes in white matter lesions in Alzheimer's disease

Cerebral white matter lesions in Alzheimer's disease (AD) consist of subcortical degeneration and ischaemic-hypoxic changes. Glial changes are intimately associated with the white matter lesions, and regressive changes in astrocytes and loss of oligodendroglial cells have been reported. We quantitatively compared glial changes including apoptosis and enhanced lysosomal activity in the frontal and temporal white matter by using terminal dUTP nick end labelling (TUNEL) and immunohistochemistry for glial markers, lysosomes and apoptosis-regulating proteins in non-familial AD brains. The degree of myelin pallor and axonal loss varied considerably in both the frontal and temporal white matter but fibrillary gliosis in demyelinated lesions tended to be less prominent in the temporal white matter in AD cases. A morphometric study with planimetric methods for cross-sectional areas of frontal and temporal white matter revealed that the white matter of AD cases manifested atrophy with significant reduction in frontal (11.9%) and temporal (29.4%) white matter compared to normal controls. Double immunolabelling for glial fibrillary acidic protein (GFAP) and KP1 (CD68) revealed KP1-positive fragmented structures within the weakly GFAP-labelled astrocytes. These KP1-positive structures correspond to process fragmentation and cytoplasmic vacuoles, which in turn indicate enhanced lysosomal activity during regressive changes in astrocytes. The KP1-modified astrocytes were not found in Pick's disease and corticobasal degeneration. The density of apoptotic glial cells, largely oligodendroglial, was significantly higher in the temporal than in the frontal white matter, and most GFAP-positive astrocytes with regressive changes were apoptotic. GFAP-positive astrocyte density was statistically the same in the frontal and temporal white matter, but the density of KP1-modified astrocytes was higher in the temporal than in the frontal white matter. The rate of white matter shrinkage was significantly correlated with the density of apoptotic glial cells and the density of KP1-modified astrocytes in the temporal lobe in AD cases. An increase in apoptotic glial cell density was found to contribute to GFAP-positive astrocytes with regressive changes in temporal white matter, while apoptosis of vascular smooth muscle cells did not show topographical accentuation. Astrocytes labelled with beta amyloid protein were not apoptotic, and the density of apoptotic cells labelled with CD95 and caspase-3 was too low in both types of white matter to be statistically evaluated. Our results imply that regressive changes in astrocytes and glial apoptosis are, to some extent, associated with white matter lesions, particularly of the temporal lobe in AD brains. The presence of apoptotic astrocytes with evidence of regressive change could therefore be a histological hallmark for white matter degeneration in AD.     

Substance P immunoreactive astrocytes are present in multiple sclerosis plaques

Substance P (SP)-like immunoreactive cells were identified in postmortem white matter tissue from patients with multiple sclerosis (MS). Labelled cells, which by morphologic criteria could be identified as astrocytes, were located at the edge of both active (e.g. inflammatory) and inactive (e.g. non-inflammatory) MS lesions. By contrast, SP-immunoreactive astrocytes were not found in normal controls and were only occasionally present in other conditions associated with astrogliosis. These data suggest that SP, a potent mediator of vasodilatation and local immune responses, may play a role in the genesis of the MS plaque. These results also extend the repertoire of potential interactions which may occur between astrocytes and cells of the immune system.     

Immunocytochemical characterisation of the immune reaction in the central nervous system in multiple sclerosis. Possible role for microglia in lesion growth

As there is evidence that in multiple sclerosis T-cell activation occurs in the central nervous system rather than outside, the inflammatory lesion may be extended through antigen presentation by cells at the edge of the plaque. In this study we present an immunocytochemical report on CNS tissue from an active case of MS, with an analysis of the distribution of CD4 and CD8 binding T cells and the expression of class I and II MHC determinants in plaques and white matter. Perivascular cuffs of early lesions, as judged by hypercellularity and minimal demyelination, contained activated T (Tac+) cells, which reacted with an anti-IL-2 monoclonal antibody. Thus sufficient T-cell growth factor would appear to be present to fuel the immune reaction in a growing lesion. The preponderance of T cells of the cytotoxic/suppressor (CD8) phenotype in the CNS parenchyma was found in conjunction with widespread staining of class I MHC antigen, a prerequisite for activity of cytotoxic T cells. Potential antigen presenting cells were demonstrated in MS plaques with a monoclonal antibody against the cytoplasmic, invariant chain of class II MHC. Macrophages and astrocytes, contributed to the staining in the hypercellular plaque border while the distribution of class II+ microglia in white matter suggest they may also be of importance in local antigen presentation.