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.     

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.     

A new monoclonal antibody, A3B10, specific for astrocyte-lineage cells recognizes calmodulin-regulated spectrin-associated protein 1 (Camsap1)

Recent studies of adult neurogenesis of the mammalian central nervous system have suggested unexpected plasticity and complexity of neural cell ontogenesis. Redefinition and reconstitution of cell classification and lineage relationships, especially between glial and neural precursors, are an urgent and crucial concern. In the present study, we describe a new monoclonal antibody, A3B10, which was produced by immunizing mice with the membrane fraction prepared from astrocyte-enriched primary neural cell cultures. Immunohistochemistry of brain sections, including brains from glial fibrillary acidic protein (GFAP)-deficient mice and primary mixed neural cell cultures, as well as immunoblot analysis and immunoelectron microscopy, have revealed that 1) A3B10 recognizes a majority of cells in ependyma in neonatal and adult rats, 2) A3B10 stains almost all GFAP(+) cells and some S100beta(+) cells in the corpus callosum, 3) A3B10 specifically stains astrocytes in vitro in primary cultures of rat embryonic cerebral hemispheres, 4) A3B10 equally stains ependymal cells of wild-type and GFAP-deficient mice, and 5) A3B10 antigen might construct intermediate filament bundles with GFAP and/or vimentin. These data suggested that the antibody labels a wide array of astorcytic-lineage cells including astrocytes, astrocyte precursors, and neural stem cells. Screening a cDNA library derived from rat embryonic brain has revealed that the antibody recognizes calmodulin-regulated spectrin-associated protein 1 (Camsap1). Thus this antibody may provide not only a new marker to identify astrocyte-lineage cells but also a new target molecule to elucidate the ontogeny, development, and pathophysiological functions of astrocyte-lineage cells.     

Fluoro-Jade, but not Fluoro-Jade B, stains non-degenerating cells in brain and retina of embryonic and neonatal rats

Fluoro-Jade (FJ) and Fluoro-Jade B (FJB) are fluorescein derivatives currently used to stain brain cells under degeneration. In this study, we investigated the FJ staining of nondegenerating cells in embryonic and neonatal rat brain and retina. In embryonic rat brain (embryonic day 15; E15), very intense staining of cells was observed. The number of FJ-stained cells and the intensity of staining decreased with increasing in animal age, being almost absent by postnatal day 16 (P16). Only a few cells in neonatal rat brain were in the process of cell death, as verified by the TUNEL technique. The FJ-stained cells in neonatal brain were positive for the neuronal marker neuronal nuclei antigen (NeuN). In retina, FJ stained mainly cells from the ganglion cell layer at P2 and the neuroblastic layer at P2 and P6. In contrast to FJ, FJB did not stain nondegenerating cells in embryonic and neonatal rats. These results show that in addition to staining degenerating brain cells, FJ also stains nondegenerating central nervous system cells in embryonic and neonatal stages.     

Immunohistochemical characterization of neurofibrillary tangles induced by mitotic spindle inhibitors

Summary Neurofibrillary tangles were induced in the motor neurons of the rabbit spinal cord by the intrathecal injection of colchicine, vinblastine, and vincristine. The tangles stained intensely by immunofluorescence and by the peroxidase-anti-peroxidase procedure using neurofilament antisera raised against chicken brain antigen, as previously reported for aluminum-induced neurofibrillary tangles. No immunohistochemical reactivity could be demonstrated between the tangles and a 150,000 dalton bovine neurofilament antiserum, although the adjacent axons were intensely stained in cryostat sections of the spinal cord.Supported by the Veterans Administration and by a grant of the National Science Foundation (BNS-7912962)     

Noradrenergic sympathetic innervation of the spleen: I. Nerve fibers associate with lymphocytes and macrophages in specific compartments of the splenic white pulp

Sympathetic noradrenergic nerve fibers, stained with antiserum for tyrosine hydroxylase (TH), richly innervate the splenic white pulp. These fibers distribute with the vascular and trabecular systems, and associate mainly with the central artery and its branches, the periarteriolar lymphatic sheath (PALS), the marginal sinus, and the parafollicular zone, with occasional delicate fibers also present in the follicles. Simultaneous staining of TH-positive nerve fibers and markers for specific lymphoid cells has shown several regions of contact between nerves and lymphocytes or macrophages. The TH-positive nerve fibers in the plexuses around the central arterial system and in the PALS are present among T lymphocytes (OX-19-positive cells) including both T helper and T suppressor cells, and interdigitating cells. At the marginal sinus, TH-positive fibers run adjacent to macrophages (ED3-positive cells), B lymphocytes (IgM-positive), and intensely fluorescent IgM-positive cells. Along the parafollicular zone, TH-positive nerve fibers run adjacent to T lymphocytes, peripheral follicular B lymphocytes, and intensely fluorescent IgM-positive cells. Within some follicles, delicate fibers end adjacent to both T and B lymphocytes. These relationships suggest a direct interaction between norepinephrine release from the TH-positive nerve terminals and the lymphocytes and macrophages closely associated with them, and focuses attention on the potential neural modulation of related functions such as T and B lymphocyte entry into the spleen and antigen capture (marginal zone), antigen presentation and T cell activation (PALS), B cell activation (parafollicular zone and marginal zone), and lymphocyte egress (outer marginal zone).     

Nonintegrin laminin receptor precursor protein is expressed on olfactory stem and progenitor cells

The biochemical identification and immunocytochemical characterization of a cell surface antigen, expressed on globose basal cells (GBCs) of the rodent olfactory epithelium (OE), are described. The monoclonal antibody (MAb) GBC-3 recognizes a surface protein, confirmed by both live cell staining and fluorescence-activated cell sorting. Two-dimensional SDS-PAGE/Western blot followed by tandem mass spectrometry demonstrates that the cell surface GBC-3 antigen is the 40 kDa laminin receptor precursor protein. The MAb GBC-3 labels the vast majority of cells among the GBC population and does not stain either sustentacular cells or horizontal basal cells (HBCs) in the normal rat OE. After epithelial lesion by exposure to methyl bromide, the remaining cells, which are mostly GBCs, are heavily stained by GBC-3, and colabeled with GBC-3 and sustentacular cell or HBC markers. GBC-3 will be a potentially useful tool for identifying and characterizing GBCs.     

Transient expression of neurofilament protein without filament formation in purkinje cell development: Immunohistological and electron microscopic study of chicken cerebellum

Purkinje cell bodies and dendrites in the chick embryo stained intensely by immunofluorescence and by the Avidin-Biotin Complex (ABC) method using monoclonal and polyclonal antibodies to the neurofilament (NF) proteins. On day 20 (the day before hatching) NF immunoreactivity markedly decreased concomitant with the first appearance of NF-positive basket axons in the molecular and Purkinje cell layers. On postnatal day 3, as in mature avian cerebellum, NF-negative Purkinje cells were surrounded by NF-positive baskets. Cytoplasmic 10 nm filaments were not observed in immature and mature Purkinje cells by electron microscopy. Basket axons in the hen were packed with 10 nm filaments.     

Serotonin-immunoreactive cells in the superior cervical ganglion of the rat. Evidence for the existence of separate serotonin- and catecholamine-containing small ganglionic cells

Superior cervical ganglia of 8 adult male rats were examined by indirect immunofluorescence microscopy, using an antibody to a conjugate of bovine serum albumin and serotonin, and an antibody to a conjugate of bovine serum albumin and noradrenaline. The fixative used was 4% paraformaldehyde in 0.1 M phosphate buffer, pH 7.3. Consecutive cryostat sections of whole ganglia were alternately stained with these antibodies. Serotonin- as well as catecholamine-immunoreactive, small ganglionic cells were demonstrated, both arranged in clusters. Serotonin-immunoreactive cells were mostly located at the cranial or caudal side of the ganglia. Serotonin-immunoreactive cells provided with processes were easily observed. Only a few mast cells were seen. Catecholamine-immunoreactive cells were rounded without processes. This cell type did not seem to have a preferential position within the ganglia. Intermingling of both immunoreactive, small ganglionic cells was not observed. A considerable variety in the number of both immunoreactive cell types was established. Catecholamine-immunoreactive cells were absent in 3 out of 8 animals. Evidence is provided that the catecholamine-immunoreactive cells are storing noradrenaline. The presented data seem to indicate the presence of 3 different types of small, intensely fluorescent (SIF) cells in the superior cervical ganglion of the rat, viz. a dopamine-storing, a noradrenaline-storing, and a serotonin-storing SIF cell type.     

An ultrastructural and immunocytochemical analysis of leptomeningeal and meningioma cultures

Using immunocytochemical methods, we localized several glycoproteins of the extracellular matrix to leptomeningeal cells and meningiomas in vitro. Three cell lines derived from normal human leptomeninges and seven from meningiomas were studied by indirect immunofluorescence to evaluate the cellular production of fibronectin, laminin, collagen type IV, and procollagen type III. All leptomeningeal cell lines stained intensely and uniformly for all matrix proteins; all meningioma cell cultures stained uniformly, but the intensity of staining varied considerably. After removal of the cells in culture adherent to glass with 25 mM ammonium hydroxide, indirect immunofluorescence demonstrated an exuberant residual extracellular residue enriched with fibronectin, laminin, collagen type IV, and procollagen type III. Electron microscopic examination of all leptomeningeal and meningioma cultures revealed desmosomes and dense tonofilament formation; in addition, granular, filamentous basement membrane-like material was abundant in the extracellular spaces of all cultures. Sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis of the cell layer of two leptomeningeal and four meningioma cultures showed production of interstitial collagen types I and III; diethylaminoethyl (DEAE)-cellulose chromatography of the medium demonstrated preferential production of procollagen type I. Our findings show conclusively that normal arachnoid cells in vitro synthesize several of the collagen subtypes and may be responsible for the "fibrous response" of the leptomeninges to trauma, infection, or infiltration by tumor. The similarities between leptomeningeal cells and meningiomas demonstrated by electron microscopy and by indirect immunofluorescence support the notion that meningiomas are derived from arachnoid cells. The localization of various mesenchymal glycoproteins within the intra- and extracellular spaces and the ubiquity of specialized intercellular junctions suggest that leptomeningeal cells in culture have the potential to behave like both stromal and epithelial cells.     

Neuronal populations stained with the monoclonal antibody Cat-301 in the mammalian cerebral cortex and thalamus

The monoclonal antibody Cat-301 was used to examine neurons in the cerebral cortex and dorsal thalamus of several mammalian species, including Old World monkeys, cats, bush babies, guinea pigs, and rats. In each species, subpopulations of cortical and thalamic neurons are stained along the surfaces of their somata and proximal dendrites. Cat-301-positive cortical neurons include specific groups of pyramidal cells (e.g., corticospinal but not corticobulbar or callosal neurons in the monkey sensory-motor areas) and certain GABA-immunoreactive nonpyramidal cells. In the thalamus, the relay neurons projecting to the cortex and not the intrinsic neurons are stained. The Cat-301-positive neurons are nonhomogeneously distributed in the cat and monkey cortex and thalamus. In the cortex, they are densely packed in 2 bands that in most areas include layers III and V, but that in primary sensory areas include layers IV and VI. Because the density of stained neurons, their distribution, and the intensity of their staining vary among cortical areas, the borders between neighboring areas can often be detected by the differences in Cat-301 staining. Broader, regional differences are also readily apparent, for areas in the parietal and occipital lobes contain large numbers of intensely stained cells, but most areas in the frontal and temporal lobes contain fewer, more lightly stained neurons. The same broad differences are seen within the thalamus: only those nuclei reciprocally connected with intensely stained cortical areas contain large numbers of Cat-301-positive neurons. Differences among species include variations in cell density and distribution when a given cortical area or thalamic nucleus is compared between cats and monkeys. Greater differences are seen among the other species. Immunoreactive neurons in the cerebral cortex are sparse and lightly stained in guinea pigs, are restricted to the hippocampal formation in rats, and are very rare and isolated in bush babies. Similarly, Cat-301-positive thalamic neurons are restricted to only one or 2 nuclei in the guinea pig and rat and are extremely rare in the bush baby. Cat-301 stains organized groups of neurons in the cat and monkey cortex and thalamus. In addition to the laminar organization of stained cells in all cortical areas (see above), the Cat-301-positive neurons of monkey areas 17 and 18 are grouped into radial arrays. In area 17, clusters of stained cells are present in layers above and below layer IVC. These clusters lie at the centers of ocular dominance columns, within patches stained for cytochrome oxidase (CO). Most of these cells are also GABA-immunoreactive.(ABSTRACT TRUNCATED AT 400 WORDS)     

Identification of a synaptic vesicle-specific 38,000-dalton protein by monoclonal antibodies

Synaptic vesicles were purified from the guinea pig cerebrum by sucrose density gradient centrifugation, and monoclonal antibodies (MAbs) were produced against this vesicle fraction. Seven MAbs (171B5, 171E8, 174D12, 174H11, 177A2, 177H11 and 178D4) recognized a novel acidic protein of about 38,000 daltons which was specific to synaptic vesicles. In immunofluorescence microscopy, the staining pattern of these MAbs corresponded to the distribution of the synapses in the guinea pig central nervous system. These MAbs appeared to stain all synaptic regions, irrespective of their synaptic function or type of neurotransmitters. MAb 171B5 and 174H11 stained the rat, rabbit and bovine synapses similarly to the guinea pig. Two other MAbs (171E8 and 177H11) stained other mammals weakly but the remaining 3 MAbs reacted only with the guinea pig. In immunoelectron microscopy of both the cerebellar tissue and isolated vesicle fraction, these MAbs selectively labeled the synaptic vesicles but not other structures. Immunoblot analysis was performed on electrophoretically separated proteins in vesicle fraction and brain homogenate. All of 7 MAbs reacted with a band at a molecular weight of about 38,000 from the guinea pig. Isoelectric focussing disclosed that this protein was acidic (pI 4.5-5).     

Anti-CD11b monoclonal antibody reduces ischemic cell damage after transient focal cerebral ischemia in rat

We investigated the effect of an anti-CD11b monoclonal antibody (1B6c) on ischemic cell damage after transient middle cerebral artery occlusion. We divided animals into three groups: MAb 1 group (n = 5)—rats were subjected to 2 hours of transient occlusion and 1B6c (1 mg/kg) was administered intravenously at 0 and 22 hours of reperfusion; MAb 2 group (n = 5)—same experimental protocol as MAb 1 group, except that the initial dose of 1B6c was increased to 2 mg/kg; and control group (n = 5)—same experimental protocol as MAb 2 group, except that an isotype-matched control antibody was administered. Animals were weighed and tested for neurological function before and after occlusion of the middle cerebral artery. Forty-six hours after reperfusion, brain sections were stained with hematoxylin and eosin for histology evaluation. We observed a significant reduction of weight loss and improvement in neurological function after ischemia in the MAb 2 animals compared to MAb 1 and vehicle-treated animals (p < 0.05). The lesion volume was significantly smaller in the MAb 2 group (19.5 ± 1.9%) compared to MAb1 (29.9 ± 2.6%) and vehicletreated (34.2 ± 5.4%) group (p < 0.01). Tissue polymorphonuclear cell numbers were reduced in both 1B6c-administered groups. Our data demonstrate that administration of anti-CD11b antibody results in a dose-dependent, significant functional improvement and reduction of ischemic cell damage after transient focal cerebral ischemia in the rat.     

The subnuclear organization of the rat interpeduncular nucleus: a light and electron microscopic study

The subnuclear organization of rat interpeduncular nucleus (IPN) has been examined by light microscopy following staining with Nissl and Holmes methods, 3H-leucine autoradiography, acetylcholinesterase (AChE), and cytochrome oxidase histochemistry on plastic sections stained with toluidine blue, and by electron microscopy. Three unpaired and four paired subnuclei are recognized. The rostral subnucleus is heavily stained for AChE, which clearly delineates its borders. It is distinguished ultrastructurally by two types of synapses on dendrites, and two on perikarya. Of the former, one type is formed by presynaptic processes which contain spherical and dense-cored vesicles and make asymmetrical contacts. Dense-cored vesicles are observed in many of the postsynaptic dendrites. A second type has presynaptic processes containing small, pleomorphic vesicles which make symmetrical contacts. Synapses on perikarya are found in the rostral, central, intermediate, lateral, and interstitial subnuclei. The dorsal subnucleus is continuous with the serotonin-containing B8 cells. The central subnucleus is distinguished by longitudinally oriented medial habenular axons separating palisades of cell bodies. These axons, which also traverse the intermediate subnuclei, form en passant S synapses with small dendrites of the central subnucleus. The intermediate subnuclei react faintly for AChE and intensely for cytochrome oxidase. They contain crest synapses formed by two habenular afferents, one from each medial habenula, which contact a narrow dendritic process en passant. The lateral subnuclei react intensely for AChE and have ultrastructural features similar to the rostral subnuclei. The interstitial subnuclei lie within each fasciculus retroflexus as it enters IPN. The small dorsal lateral subnuclei are evident by light microscopy.     

The mechanism of inhibition of experimental allergic encephalomyelitis in the rat by monoclonal antibody against CD4

Lewis rats with actively induced or passively transferred experimental allergic encephalomyelitis (EAE) were treated with a monoclonal antibody (MAb) which binds to the CD4 antigen of rat helper/inducer T cells. Actively immunized animals treated at the first onset of clinical signs experienced only a mild form of the disease and rapidly recovered while the majority of those treated prophylactically never showed clinical signs of EAE. Passively transferred EAE was also completely inhibited with anti-CD4 MAb. In treated animals which exhibited only mild clinical signs of EAE, spinal cord and cerebellar leukocyte infiltrates were quite similar to those in untreated rats but where anti-CD4 MAb treatment completely prevented clinical EAE, histological signs were minimal or absent. Like Lewis rats which have recovered naturally from EAE, those treated with anti-CD4 MAb were both resistant to a secondary challenge with myelin basic protein and harboured potential encephalitogenic cells which were capable of transferring disease to recipient rats. Disease in these recipients was, however, of much greater severity than that experienced by animals receiving cells from naturally recovered (untreated) donors. These data demonstrate that administration of anti-CD4 MAb to rats can prevent EAE by a mechanism which does not ablate the encephalitogenic CD4+ cells or prevent the development of resistance to EAE but which may inhibit the disease by preventing the function of already activated effector cells.     

Hypothalamic agouti-related protein immunoreactivity in food-restricted, obese, and insulin-treated animals: evidence for glia cell localization

Agouti-related protein (AGRP) has been implicated in the regulation of metabolic balance. Overexpression of this peptide leads to obesity. Its activity is mediated via the melanocortin-4 (MC-4) receptor where it acts as an MC-4 receptor antagonist. In this study, we characterized the AGRP brain distribution and cellular localization in control, food-restricted, obese, and insulin-treated rats using immunohistochemistry. AGRP immunostaining was found selectively in regions of the arcuate and ventromedial hypothalamic nuclei. These regions were stained less intensely in food-restricted rats than in controls. AGRP-positive cells in the hypothalamus of obese animals were three times more numerous than in control rats. Also, insulin treatment acted to decrease AGRP immunostaining. Analysis of AGRP cellular localization demonstrated its presence in the cytoplasm of numerous small (7-12 microm) cell bodies of putative protoplasmic astrocytes as well as in nerve fibers. Glia fibrillary acidic protein (GFAP) immunostaining of sections adjacent to those stained for AGRP revealed astrocytes with morphology similar to AGRP-positive cells. A few AGRP-positive nerve cell bodies were also found in the arcuate nucleus of obese rats. We conclude that AGRP hypothalamic content is decreased by fasting and intracerebroventricular (i.c.v.) insulin treatment and increased in obesity. In addition to its presence in nerve fibers, AGRP localization in astroglia-like cells suggests a possible role for these elements in its synthesis or its sequestration from the neuronal compartment.     

Cultured rat Schwann cells express low affinity receptors for nerve growth factor

Schwann cell cultures prepared from postnatal Sprague-Dawley rat sciatic nerves were used to demonstrate the presence of specific receptors for the beta-subunit of nerve growth factor (NGF) on rat Schwann cells. Indirect immunofluorescence microscopy with a monoclonal antineuronal NGF receptor (NGFR) antibody indicated that NGFR antigen was expressed on the surface of Schwann cells but not of endoneurial fibroblasts. Studies with 125I-NGF confirmed this distribution of NGFR in the cultures and showed that the Schwann cell NGFR had a single NGF binding affinity (Kd of 1.8 x 10(-9) M). 125I-NGF binding by the cultured Schwann cells increased with time in vitro, reaching a plateau level on the 4th day, but decreased with increasing age, reaching 40% of the neonatal value in Schwann cells isolated from 12-day-old rats. Treatment of the cultures with NGF did not alter Schwann cell phenotype, survival or proliferation.     

Expression of glutamic acid decarboxylase in nervous tissue structures targeted by autoantibodies in patients with diabetic autonomic neuropathy

We have previously identified an association between symptomatic diabetic autonomic neuropathy (DAN) and autoantibodies to sympathetic and parasympathetic nervous structures. The antigens identified by these autoantibodies are not known, but glutamic acid decarboxylase (GAD) has been suggested as a candidate target, since anti-GAD autoantibodies are present in patients with long-term diabetes and GAD is expressed in a variety of cell types and structures in the nervous system. The aim of this study was to examine GAD expression in sympathetic ganglia and vagus nerve and to compare the distribution of GAD within these tissues with that of anti-sympathetic ganglia and anti-vagus nerve autoantibodies from patients with DAN, using single and double indirect immunofluorescence on tissue sections. The monoclonal antibody GAD-6, specific for GAD₆₅, gave a granular, peripheral, cytoplasmic staining pattern in sympathetic ganglion cells. Dual immunofluorescence demonstrated that serum from a patient with anti-sympathetic ganglion autoantibodies stained the same cells, but homogeneously throughout the cytoplasm. In the vagus nerve, patient's serum stained the fibres only; GAD-6 stained the cytoplasm of parasympathetic ganglion cells but only occasional fibres. In addition, GAD enzymatic activity was detectable in both sympathetic ganglia and vagus nerve. Incubation of sera or GAD-6 overnight with a crude homogenate of human brain as an antigen source abolished staining of the nervous tissues by GAD-6, but not by patients' sera. The different localisation of GAD and the autoantigens targeted by patients' sera indicates that GAD is not the target of the autoantibodies characteristic of DAN. Moreover, absorption studies using human brain homogenate suggest that the targets of anti-sympathetic ganglion and anti-vagus nerve autoantibodies are absent or represented only at low levels in the central nervous system and may be confined to the periphery.     

Fc gamma receptors and HLA-DR antigens on thymus cells in myasthenia gravis

Receptors for the Fc part of IgG (Fc gamma R) and HLA-DR antigens were detected in thymus tissue from patients with myasthenia gravis (MG) using monoclonal antibodies (B1D6 and OKIa1) in indirect immunofluorescence. The amount of Fc gamma R and HLA-DR antigens was increased on epithelial reticular cells and on interdigitating reticular cells in hyperplastic thymus as compared to normal thymus. In thymomas from MG patients the neoplastic epithelial cells expressed Fc gamma R, whereas only a few cells had HLA-DR antigens. Neither thymocytes nor B lymphocytes in the hyperplastic thymuses and in the thymomas were stained by B1D6.