Myelin Basic Protein Gene Research Articles

Tumor necrosis factor modulates transcription of myelin basic protein gene through nuclear factor kappa B in a human oligodendroglioma cell line

Tumor necrosis factor-α (TNF-α) is a major mediator of inflammation and it is involved in many neurological disorders such as multiple sclerosis. Levels of TNF-α and lymphotoxin-α have been found elevated in plaques, bloods, and cerebral spinal fluids from multiple sclerosis patients. The expression of myelin basic protein (MBP), a major protein of the myelin sheath, is affected by cytokines secreted by activated immune cells. To determine the signal transduction pathway involving tumor necrosis factor’s action in myelination and demyelination, we have cloned and analyzed cis-elements on promoters of the human and mouse MBP genes. There are two putative nuclear factors kappa-B (NF-κB) cis-elements on the human and one on the mouse gene promoter. In an electrophoretic mobility shift assay, all three NF-κB cis-elements showed binding to a protein, which was recognized by an antibody against NF-κB P65 component. The specificity of the binding was demonstrated in a competitive assay using NF-κB consensus oligonucleotides. A two base pair site-directed mutation on the mouse NF-κB cis-element abolished its binding activity. We created a DNA construct by linking the mouse MBP gene promoter containing the NF-κB cis-element to luciferase gene. Transfection of this construct into a human oligodendroglioma cell line showed TNF-α increased the transgene expression. Furthermore the mutation of NF-κB site abolished TNF-α -induction of the transgene. The data demonstrate that NF-κB is the mediator between tumor necrosis factor’s action and MBP gene expression. Elucidating the molecular mechanisms underlying TNF-α regulation of MBP gene expression provides new scientific bases for the development of therapy against oligodendrocyte-specific and myelin-related disorders such as multiple sclerosis.

Cyclic AMP inducibility of the myelin basic protein gene promoter requires the NF1 site

In the central nervous system oligodendrocyte differentiation is accompanied by the activation of a specific transcriptional program responsible for the synthesis of myelin genes. One of the signals leading to the expression of myelin components, such as the myelin basic protein (MBP) gene is cyclic AMP (cAMP). Previous work using a cell line in which the endogenous MBP gene can be induced by increased cAMP levels (D6P2T) showed that the region of the MBP gene that was required for induction of the gene by cAMP lay between −248 and −105 in the 5′ flanking region. This region contains numerous transcription factor binding sites, including sites for NF1, Sp1, and MEBA. In order to determine if the NF1 site itself was specifically responsible for the cAMP responsiveness of the MBP promoter, stably transfected cells carrying MBP promoter deletion constructs were used. Deletion of just the NF1 site caused loss of responsiveness to cAMP levels. Furthermore, site-specific mutations in the NF1 site that interfere with NF1 protein binding, in the context of the full length promoter, abolished cAMP responsiveness and caused derepression of the promoter. Analysis of protein binding to the NF1 site showed that the mutation resulted in loss of binding to the site and that the proteins binding at the site are modified in the presence of cAMP elevating agents. These results demonstrate that the NF1 site is indispensable for cAMP responsiveness of the MBP promoter and, together with other DNA elements, plays a role in controlling MBP gene expression.

Identification of Golli and myelin basic proteins in human brain during early development.

The myelin basic protein gene (Mbp) encodes for the major myelin structural proteins and it is included in the Golli-Mbp gene complex. Previously, we observed MBP-like proteins in the human central nervous system (CNS) at developmental stages preceding myelination. In an effort to distinguish between Golli (HOG5 and HOG7) and MBP mRNAs and to determine their spatiotemporal distribution, we performed in situ hybridization using two human Golli specific probes: one corresponding to exon 5a absent from all MBP transcripts, and the other corresponding to exon 5c specific for HOG5. HOG7 transcript was observed first, in 5 gestational week-old embryos, whereas both Golli transcripts were detected at 6-7 weeks gestation in the proliferative zones of the entire CNS. Golli proteins immunoreactivity was observed in microglia and early neurons of the developing telencephalon. During midgestation (17-22 weeks gestation), at the onset of myelination, MBP and Golli mRNAs were observed in the telencephalic subventricular zone and occasionally in the future cerebral cortex. Developmental expression of the human Golli-Mbp indicates that the two Golli proteins have different onset of expression, distribution and possibly function. These results support the hypothesis that at least one of them, HOG7, may be involved in the regulation of early neurogenesis, while both may have additional, still undefined function at the onset of myelination.

P27Kip1 Enhances myelin basic protein gene promoter activity

The process of oligodendrocyte differentiation is a complex event that requires cell cycle withdrawal, followed by the activation of a specific transcriptional program responsible for the synthesis of myelin genes. Because growth arrest precedes differentiation, we sought to investigate the role of cell cycle molecules in the activation of myelin gene promoters. We hypothesized that the cell cycle inhibitor p27Kip1, which is primarily responsible for arresting proliferating oligodendrocyte progenitors, may be involved in the transcriptional regulation of myelin genes. In agreement with this hypothesis, overexpression of p27Kip1 in the CG4 cell line, but not in 3T3 fibroblasts, enhances the expression of luciferase driven by the myelin basic protein (MBP) promoter. Interestingly, this effect is specific for p27Kip1; overexpression of other cell cycle inhibitors had no effect. Additionally, this effect is independent of halting the cell cycle; treatment with the cell cycle blocker roscovitine did not affect MBP promoter usage. We conclude that p27Kip1 contributes to oligodendrocyte differentiation by regulating transcription of the MBP gene. © 2002 Wiley-Liss, Inc.

P27(Kip1) enhances myelin basic protein gene promoter activity.

The process of oligodendrocyte differentiation is a complex event that requires cell cycle withdrawal, followed by the activation of a specific transcriptional program responsible for the synthesis of myelin genes. Because growth arrest precedes differentiation, we sought to investigate the role of cell cycle molecules in the activation of myelin gene promoters. We hypothesized that the cell cycle inhibitor p27(Kip1), which is primarily responsible for arresting proliferating oligodendrocyte progenitors, may be involved in the transcriptional regulation of myelin genes. In agreement with this hypothesis, overexpression of p27(Kip1) in the CG4 cell line, but not in 3T3 fibroblasts, enhances the expression of luciferase driven by the myelin basic protein (MBP) promoter. Interestingly, this effect is specific for p27(Kip1); overexpression of other cell cycle inhibitors had no effect. Additionally, this effect is independent of halting the cell cycle; treatment with the cell cycle blocker roscovitine did not affect MBP promoter usage. We conclude that p27(Kip1) contributes to oligodendrocyte differentiation by regulating transcription of the MBP gene.

Expression and regulation of golli products of myelin basic protein gene during in vitro development of oligodendrocytes

The myelin basic protein (MBP) gene produces two families of proteins, the classic MBPs, important for myelination of the CNS, and the golli proteins, whose biological role in oligodendrocytes (OLs) is still unknown. The goals of this work were to study the in vitro pattern of expression of the golli products during OL differentiation and to compare it with that of the classic MBP products of the gene. Mouse primary glial cultures were analyzed at the mRNA and protein levels with an array of techniques. We found that OLs express golli mRNA primarily during intermediate stages of differentiation, which was confirmed by immunocytochemical analysis. Golli expression was low in proliferating OL progenitors as well as in terminally mature OLs. Golli proteins were found associated with the OL cell soma and nuclei and, to a lesser extent, with the cellular processes. We also found that golli proteins are not targeted to myelin in vitro and in vivo, in contrast to the classic MBPs. Finally, we found that golli expression is regulated during OL development and can be manipulated by growth factors such as basic fibroblast growth factor, neurotrophin-3, and retinoic acid.

The Specificity of the Myelin Basic Protein Gene Promoter Studied in Transgenic Mice

The myelin basic proteins (MBPs) are a family of polypeptides that are predominantly expressed in the nervous system where they play a major role in myelination. We have generated four lines of transgenic mice carrying a transgene in which 1.34 kb of the 5′-flanking sequence of the mouse MBP gene was fused upstream of the coding region of the Escherichia coli lac Z gene in order to investigate developmental and tissue-specific expression of the MBP gene. Expression of both the lacZ transgene and the endogenous MBP gene followed a common developmental pattern in mouse brain. Transgene expression was detected in primary oligodendrocytes, but not in type 2 astrocytes. In addition, the lacZ gene product was expressed in epithelial cells of certain nonneural tissues, namely kidney, epididymis, ureter, and seminal vesicles. The ectopic expression of the transgene was associated with the development of DNase I hypersensitive sites at the site of insertion which was found to be within the intron 1 region of the endogenous MBP gene. The results reported here strongly suggest that the 1.34-kb 5′-flanking region of the MBP gene contains cis-regulatory elements that confer developmental regulation of the MBP gene, although this region appears to lack elements that restrict its expression to the nervous system.

Expression of Golli mRNA during development in primary immune lymphoid organs of the rat

The gene-of-the-oligodendrocyte lineage (Golli)–MBP transcription unit contains three Golli-specific exons together with eight exons of the “classical” myelin basic protein (MBP) gene, yielding alternatively spliced proteins which share amino acid sequence with MBP. Unlike MBP, a late antigen expressed only in the nervous system, Golli gene products are expressed pre- and post-natally at many sites. In this study, we determined the sequence of Golli in rat by RT-PCR and 5′ RACE and showed that Golli sequences are expressed in primary lymphoid organs as early as e16.5, which could explain the anergic rat T cell response we previously observed in Golli-induced meningitis.

Focal cerebral ischemia induces increased myelin basic protein and growth-associated protein-43 gene transcription in peri-infarct areas in the rat brain.

Although oligodendrocytes are vulnerable to focal cerebral ischemia, remyelination of denuded or regenerating axons in the peri-infarct area has been observed in the central nervous system. We studied the expression of myelin basic protein (MBP), a major component of central nervous system myelin, in peri-infarct areas in adult rat brain after transient middle cerebral artery occlusion (MCAO) and correlated it to the expression of the growth-associated protein-43 (GAP-43), a marker for axonal regeneration and sprouting, using non-radioactive in situ hybridization techniques. Within the infarct, MBP messenger RNA (mRNA) had disappeared by 24 h, whereas myelin protein, identified by MBP and myelin oligodendrocyte glycoprotein (MOG) immunohistochemistry, appeared structurally intact until day 3. Peri-infarct oligodendrocytes increased their expression of MBP mRNA from 24 h to maximal levels at day 7, corresponding to the appearance of process-bearing MBP and occasional MOG-immunoreactive oligodendrocytes in parallel sections. Quantitative analysis revealed significant increases in the density of oligodendrocytes (up to 7.6-fold) and in the level of MBP mRNA expressed by individual cells. Parallel sections showed that increased expression of GAP-43 mRNA in neurons was concomitant to MBP mRNA upregulation in oligodendrocytes. While the mechanisms regulating oligodendrocyte survival and myelination signals are not clear at this point, axonal sprouting could putatively serve as a stimulus for the upregulation of oligodendrocyte cell numbers, differentiation state, and/or active myelination in the peri-infarct areas.

Changes in Microtubule Stability and Density in Myelin-Deficient Shiverer Mouse CNS Axons

Altered axon-Schwann cell interactions in PNS myelin-deficient Trembler mice result in changed axonal transport rates, neurofilament and microtubule-associated protein phosphorylation, neurofilament density, and microtubule stability. To determine whether PNS and CNS myelination have equivalent effects on axons, neurofilaments, and microtubules in CNS, myelin-deficient shiverer axons were examined. The genetic defect in shiverer is a deletion in the myelin basic protein (MBP) gene, an essential component of CNS myelin. As a result, shiverer mice have little or no compact CNS myelin. Slow axonal transport rates in shiverer CNS axons were significantly increased, in contrast to the slowing in demyelinated PNS nerves. Even more striking were substantial changes in the composition and properties of microtubules in shiverer CNS axons. The density of axonal microtubules is increased, reflecting increased expression of tubulin in shiverer, and the stability of microtubules is drastically reduced in shiverer axons. Shiverer transgenic mice with two copies of a wild-type myelin basic protein transgene have an intermediate level of compact myelin, making it possible to determine whether the actual level of compact myelin is an important regulator of axonal microtubules. Both increased microtubule density and reduced microtubule stability were still observed in transgenic mouse nerves, indicating that signals beyond synaptogenesis and the mere presence of compact myelin are required for normal regulation of the axonal microtubule cytoskeleton.

Hypoxic–ischemic injury results in acute disruption of myelin gene expression and death of oligodendroglial precursors in neonatal mice

Studies of ischemic brain injury in neonatal rodents have focused upon the pathophysiology of neuronal damage. Much less consideration has been given to white matter injury, even though it is a major contributor to chronic neurological dysfunction in children. In the human neonate, particularly in those born prematurely, periventricular white matter is highly susceptible to hypoxic–ischemic (H–I) injury. To understand the basis for this selective vulnerability, we examined myelin gene expression and cell death in the subventricular layer and the surrounding white matter of neonatal mice following H–I insult. Using an in situ hybridization technique that gives high resolution and is very sensitive, we examined myelin basic protein and proteolipid protein gene expression three and twenty-four hours after a H–I insult. To elicit unilateral forebrain hypoxic and ischemic injury, 9–10-day-old mice underwent right carotid artery ligation followed by timed (40–70 min) exposure to 10% oxygen. Twenty-four hours following H–I, myelin basic protein and proteolipid protein transcripts were markedly reduced in striatum, external capsule, fornix, and corpus callosum in the injured side. Three hours after lesioning (ligation+70 min hypoxic exposure) myelin basic protein gene transcripts were visibly reduced in the ipsilateral white matter tracts. Interestingly, some cells in the subventricular layer expressed proteolipid protein transcripts, and 3 h after a H–I insult they were degenerating in the injured but not contralateral side. TUNEL staining showed an increase in the number of positive cells in the injured subventricular layer and corpus callosum but the adjacent striatum did not show a corresponding change in the number of TUNEL labeled cells. Ultrastructural studies of the subventricular zone and corpus callosum 3 h after H–I revealed that many subventricular cells, glial cells in the corpus callosum, and callosal axons in the injured side had already degenerated. However, the subventricular cells, glia and axons in the contralateral corpus callosum were spared. Many cells in the injured corpus callosum exhibited a apoptotic morphology; yet more mature oligodendrocytes in this region appeared normal. Our results show that a H–I insult causes a surprisingly swift and dramatic degenerative response in the subventricular layer and adjacent white matter. Within 3 h after H–I, the programmed cell death cascade was initiated; internucleosomal DNA degradation took place in subventricular and glial cells; oligodendrocyte progenitors died and axonal degeneration in the ipsilateral corpus callosum was extensive. The swiftness of the subventricular and glial cell degeneration suggests the H–I insult directly targets glia, as well as neurons, and raises the provocative question of whether glia exert damaging effects upon neurons and axons. Since the severity of the H–I insult can be modulated by varying the duration of hypoxia, the model is ideal to study whether oligodendrocyte progenitors are more susceptible to death than mature oligodendrocytes, whether mature oligodendrocytes de-differentiate and then are induced to remyelinate surviving axons, and/or whether oligodendrocyte progenitors in the subventricular layer can be stimulated to proliferate, migrate, and remyelinate the surviving axons.

Immunohistochemical Analysis of Equine Pulmonary Granular Cell Tumours

Histopathological and immunohistochemical examinations were made on four female horses aged 9–12 years with pulmonary granular cell tumours (GCTs). The tumours, which were multiple, of varying size, firm and off-white in colour, surrounded the bronchi and bronchioles. Metastatic lesions were not detected. The tumour cells had abundant eosinophilic cytoplasm filled with prominent coarse eosinophilic granules. Immunohistochemically, these tumour cells reacted uniformly with vimentin and S100 antibodies. Most were immunolabelled by antibodies against glial fibrillary acidic protein (GFAP), myelin basic protein (MBP) and protein gene product 9·5 (PGP9·5), and a few cells were positive with Leu7 antibody. However, the tumour cells did not react with antibodies against neurofilament protein (NF), cytokeratin (CK), chromogranin, α1 antichymotrypsin (AACT), myoglobin, desmin, α-actin or α-smooth muscle actin (α-SMA). These immunohistochemical properties of tumour cells support the hypothesis that equine pulmonary GCTs are derived from Schwann cells of the peripheral nervous system in peribronchial and peribronchiolar tissues. GFAP, MBP, Leu7 and PGP9·5 antibodies should help to distinguish equine granular cell tumours from other tumours.

A role for an AP-1-like site in the expression of the myelin basic protein gene during differentiation

Differentiation of oligodendrocyte progenitors into mature oligodendrocytes involves the timely, cell-type specific expression of a number of different genes. Among these, the expression of the myelin basic protein (MBP) gene closely parallels the course of oligodendrocyte differentiation. To understand how transcription of the myelin basic protein gene is controlled, binding to the distal end of the 5′ flanking sequence of the MBP gene was investigated. Specific protein-DNA complexes were localized to an AP-1-like element located between −1230 and −1240. The protein–DNA complexes formed at this site were shown to change as the cells differentiated. In undifferentiated cells two complexes were formed but, as the cells differentiated, binding was nearly completely lost. One of the two complexes was shown to contain a member of the fos family of transcription factors but no jun family members were involved. Mutation of the AP-1-like site resulted in loss of the complex and a change in expression of a reporter construct driven by the mutated promoter sequence. These results demonstrate a role for the AP-1-like site in repression of MBP gene expression in oligodendrocyte progenitor cells.

The pathobiology of myelin mutants reveal novel biological functions of the MBP and PLP genes.

Substantial biological data indicate that the myelin basic protein (MBP) and myelin proteolipid protein (PLP/DM20) genes produce products with functions beyond that of serving as myelin structural proteins. Much of this evidence comes from studies on naturally-occurring and man-made mutations of these genes in mice and other species. This review focuses upon recent evidence showing the existence of other products of these genes that may account for some of these other functions, and recent studies providing evidence for alternative biological functions of PLP/DM20. The MBP and PLP/DM20 genes each encode the classic MBP and PLP isoforms, as well as a second family of proteins that are not involved in myelin structure. The biological roles of these other products of the genes are becoming clarified. The non-classic MBP gene products appear to be components of transcriptional complexes in the nucleus, and they also may be involved in signaling pathways in T-cells and in neural cells. The non-classic PLP/DM20 gene products appear to be components of intracellular transport vesicles in oligodendrocytes. There is evidence for other functions of the classic PLP/DM20 proteins, including a role in neural cell death mechanisms, autocrine and paracrine regulation of oligodendrocytes and neurons, intracellular transport and oligodendrocyte migration.

No evidence for transmission disequilibrium between a new marker at the myelin basic protein locus and multiple sclerosis in French patients.

The myelin basic protein (MBP) gene is a candidate locus for susceptibility to multiple sclerosis. Several groups have tested a complex (TGGA)n repeat in the 5' region of this gene for association/linkage with multiple sclerosis, with divergent results. This region of tandem repetitive sequence has been subjected to complex rearrangements, and there is a possibility that alleles of the same size have different internal structures, which reduces the interest of this marker for linkage disequilibrium studies and may at least partly explain the conflicting results obtained so far. To overcome this problem, we isolated a new polymorphic (CA)n repeat within the Golli-MBP locus. The limited number of alleles identified makes this other marker suitable for transmission disequilibrium studies. We tested this marker for linkage with multiple sclerosis, using the transmission-disequilibrium test (TDT) on a sample of 196 nuclear families in which the genotypes of both parents could be unambiguously defined. We found no evidence of transmission disequilibrium between multiple sclerosis and any of the three alleles of this marker, even when the patients were subdivided according to their HLA-DRB1*1501 status. The present data thus provide no evidence for a contribution of the MBP gene to multiple sclerosis susceptibility in French patients.

Thymocytes express the golli products of the myelin basic protein gene and levels of expression are stage dependent.

The golli products of the myelin basic protein gene have been shown to be expressed in mouse thymus and brain. The full repertoire of thymic cell types expressing golli products has not yet been determined, although immunoreactivity has been found in some macrophages. We have analyzed the cellular expression of golli mRNAs and proteins in the thymus. The results showed that MTS5(+) cortical/MTS10(+) medullary epithelial cells and NLDC145(+) dendritic cells did not express golli, while some macrophages did exhibit strong immunoreactivity. GOLLI: mRNAs were not detected in macrophages by in situ hybridization. Thymocytes expressed significant levels of golli mRNAs and proteins by in situ hybridization and immunohistochemistry. Interestingly, golli immunoreactivity varied with thymocyte stage of differentiation. For example, CD4(-)CD8(-) (double-negative) thymocytes expressed relatively high levels of golli. Upon further differentiation into CD4(-)CD8(-) (double-positive) thymocytes, golli protein expression declined dramatically. When thymocytes developed into CD8(-) or CD4(+) (single-positive) thymocytes, golli protein expression increased again, but it never achieved the levels found in double-negative thymocytes. Thus, the altered levels of expression of golli proteins in developing thymocytes correlated with the transitions from double-negative to double-positive and double-positive to single-positive stages. The lack of significant golli expression in thymic stromal cells may offer an alternative explanation for the mechanism of inefficient negative selection of those autoreactive thymocytes with specificity for myelin basic proteins.

A Transgenic Mouse Model for Inducible and Reversible Dysmyelination

Oligodendrocytes are glial cells devoted to the production of myelin sheaths. Myelination of the CNS occurs essentially after birth. To delineate both the times of oligodendrocyte proliferation and myelination, as well as to study the consequence of dysmyelination in vivo, a model of inducible dysmyelination was developed. To achieve oligodendrocyte ablation, transgenic animals were generated that express the herpes virus 1 thymidine kinase (HSV1-TK) gene under the control of the myelin basic protein (MBP) gene promoter. The expression of the MBP-TK transgene in oligodendrocytes is not toxic on its own; however, toxicity can be selectively induced by the systemic injection of animals with nucleoside analogs, such as FIAU [1-(2-deoxy-2-fluoro-beta-delta-arabinofuranosyl)-5-iodouracil]. This system allows us to control the precise duration of the toxic insult and the degree of ablation of oligodendrocytes in vivo. We show that chronic treatment of MBP-TK mice with FIAU during the first 3 postnatal weeks triggers almost a total depletion of oligodendrocytes in the CNS. These effects are accompanied by a behavioral phenotype characterized by tremors, seizures, retarded growth, and premature animal death. We identify the period of highest oligodendrocytes division in the first 9 postnatal days. Delaying the beginning of FIAU treatments results in different degrees of dysmyelination. Dysmyelination in MBP-TK mice is always accompanied by astrocytosis. Thus, this transgenic line provides a model to study the events occurring during dysmyelination of various intensities. It also represents an invaluable tool to investigate remyelination in vivo.

The re-expression of the homeodomain transcription factor Gtx during remyelination of experimentally induced demyelinating lesions in young and old rat brain

Since myelination and remyelination both involve investing an axon with a myelin sheath, a plausible hypothesis is that the two processes involve the expression of similar transcription factors. In this study we have addressed this hypothesis by comparing the expression of messenger RNA of Gtx, a homeodomain transcription factor expressed within oligodendrocytes during myelination, with the expression of messenger RNAs of the major myelin proteins, myelin basic protein and proteolipid protein during remyelination of experimentally induced demyelination in the adult rat brain. We have found a close temporal and spatial association between the expression patterns of the three messenger RNA species during remyelination. By comparing the expression patterns in rapidly remyelinating lesions in young adult rats with slowly remyelinating lesions in old adult rats, we have shown that Gtx messenger RNA expression follows the reappearance of myelin basic protein and proteolipid protein messenger RNAs regardless of the rate of remyelination. This observation demonstrates a clear association between the expression of Gtx messenger RNA and myelin repair. We have also shown that there is a decrease in constitutive levels of expression of myelin basic protein, proteolipid protein and Gtx messenger RNA in old adults compared with young adults. Taken together, our results indicate that Gtx, which has multiple binding sites in the promoter regions of both myelin basic protein and proteolipid protein genes, may have a similar role in the regulation of myelin protein gene expression during remyelination as has been proposed in myelination.

Identification of a novel silencer that regulates the myelin basic protein gene in neural cells

The myelin basic protein gene produces two families of proteins, the golli proteins and the ‘classic’ myelin basic proteins from three transcription start sites (tsp). The golli proteins are expressed from the first tsp, and little is known about genetic elements that control its activity. We have examined elements that may regulate the expression of the golli products produced from this promoter in neural cell lines with constructs containing upstream portions of the first tsp by transient transfection assays. Three putative regulatory elements were identified, among them a 345 bp novel silencer region, termed the golli silencer region (GSR), which was characterized in detail. This silencer was responsible for a significant (approx. 60%) inhibition of luciferase expression in PC12 cells. It was orientation-dependent and a double dose of this GSR completely abolished expression of the luciferase reporter activity. Transfections with deleted constructs identified three critical sites that bind at least two repressor proteins. We postulate that the silencer activity is the result of synergistic interactions between these repressor proteins and might involve the formation of a high-ordered protein–DNA structure.

GFAP-positive and myelin marker-positive glia in normal and pathologic environments.

The data herein demonstrate that in addition to the well-characterized myelin marker-positive, glial fibrillary acidic protein (GFAP)-negative, membrane sheet-bearing oligodendrocytes, another type of myelin marker-positive, process-bearing glia exists in normal and pathologic conditions. This second type of myelin marker-positive glia expresses GFAP, and therefore these cells have been referred to as mixed phenotype glia. Although mixed phenotype glia have been documented previously, their identity and function have remained a mystery. The goal of this immunocytochemical study was to further characterize these cells. Using the MBPlacZ transgenic mouse in which beta-galactosidase is under the control of the myelin basic protein (MBP) gene promoter, GFAP-positive/beta-galactosidase-positive and myelin/oligodendrocyte-specific protein (MOSP)-positive/beta-galactosidase-positive cells were detected in subcortical white matter and in perivascular locations within cerebral white and gray matter. In cultures prepared from highly enriched myelin marker-positive immature glia, mixed phenotype glia were detected that were GFAP-positive and either MOSP-, MBP-, O1-, and O4-positive. The expression of multiple myelin markers by mixed phenotype glia may suggest that these cells are of oligodendrocyte origin. Increased numbers of MOSP-positive/GFAP-positive mixed phenotype glia were detected in sections from adult hypomyelinated brain from shiverer, quaking, and PKU mice compared to myelinated control adult mouse brain. Similarly, cultures from control brain exposed to elevated pH for 2-3 weeks showed dramatically increased numbers of mixed phenotype glia (80%) compared to control (<10%). Increased numbers of mixed phenotype glia also were detected in shiverer cultures (40%). Since increases in the number of mixed phenotype glia occur in shiverer, quaking, and PKU mouse brain, these data suggest that mixed phenotype glia contribute to gliosis in pathologic white matter.