Antioxidant treatment ameliorates prefrontal hypomyelination and cognitive deficits in a rat model of schizophrenia.

Cognitive dysfunction in schizophrenia (SZ) is thought to arise from neurodevelopmental abnormalities that include interneuron hypomyelination in the prefrontal cortex (PFC). Here we report that RNA-sequencing of the medial (m)PFC of the APO-SUS rat model with SZ-relevant cognitive inflexibility revealed antioxidant metabolism as the most-enriched differentially expressed pathway. Antioxidant-related gene expression was altered throughout postnatal development and preceded hypomyelination. Furthermore, reduced glutathione levels and increased mitochondria numbers were observed in the mPFC. Strikingly, chronic treatment with the glutathione precursor N-acetylcysteine (NAC) from postnatal days 5-90 restored not only antioxidant-related mRNA expression and mitochondria numbers, but also myelin-related mRNA expression and mPFC-dependent cognitive dysfunction, while blood glutathione levels remained unaffected. The promyelinating effect of NAC was at least partly due to a positive effect on oligodendrocyte lineage progression. Together, our findings highlight that oxidative stress may contribute to cognitive symptoms in the APO-SUS rat model of SZ and encourage antioxidant therapy in early phases of SZ.

SOX17 is expressed in regenerating oligodendrocytes in experimental models of demyelination and in multiple sclerosis.

We have previously demonstrated that Sox17 expression is prominent at developmental stages corresponding to oligodendrocyte progenitor cell (OPC) cycle exit and onset of differentiation, and that Sox17 promotes initiation of OPC differentiation. In this study, we examined Sox17 expression and regulation under pathological conditions, particularly in two animal models of demyelination/remyelination and in post-mortem multiple sclerosis (MS) brain lesions. We found that the number of Sox17 expressing cells was significantly increased in lysolecithin (LPC)-induced lesions of the mouse spinal cord between 7 and 30 days post-injection, as compared with controls. Sox17 immunoreactivity was predominantly detected in Olig2(+) and CC1(+) oligodendrocytes and rarely in NG2(+) OPCs. The highest density of Sox17(+) oligodendrocytes was observed at 2 weeks after LPC injection, coinciding with OPC differentiation. Consistent with these findings, in cuprizone-treated mice, Sox17 expression was highest in newly generated and in maturing CC1(+) oligodendrocytes, but low in NG2(+) OPCs during the demyelination and remyelination phases. In MS tissue, Sox17 was primarily detected in actively demyelinating lesions and periplaque white matter. Sox17 immunoreactivity was co-localized with NOGO-A+ post-mitotic oligodendrocytes both in active MS lesions and periplaque white matter. Taken together, our data: (i) demonstrate that Sox17 expression is highest in newly generated oligodendrocytes under pathological conditions and could be used as a marker of oligodendrocyte regeneration, and (ii) are suggestive of Sox17 playing a critical role in oligodendrocyte differentiation and lesion repair.

The role of SVZ-derived neural precursors in demyelinating diseases: from animal models to multiple sclerosis.

We will review the role of endogenous neural stem cells in myelin repair both in animal models of demyelination and multiple sclerosis. The mammalian sub-ventricular zone (SVZ) is the largest germinative zone of the adult brain, which contains a well characterized stem cell niche. While most studies highlight the neurogenic potential of SVZ progenitors, recent data indicate that SVZ cells become reactived in response to different pathological cues, like trauma, ischemia, neurodegeneration, inflammation and demyelination. Experimental models of demyelination in rodent demonstrate enhanced proliferation and recruitment of SVZ progenitors into myelin lesions, in response to demyelination. Moreover, cell lineage tracing experiments showed that SVZ progenitor cells can give rise to oligodendrocytes in demyelinated lesions, that could potentially contribute to remyelination. To examine the relevance of these studies in myelin diseases, we recently examined the human SVZ in post-mortem MS brains. The human SVZ is characterized by a ribbon of SVZ astrocytes lining the ependymal border of the lateral ventricles and which behave as multipotential progenitors in vitro. We showed that cellular density and proliferation were enhanced in MS SVZ compared to non-neurological controls. This high cellular density was correlated with the increased number of progenitor cells in MS SVZ, as well as in sub-ventricular lesions. Interestingly, some of these progenitors expressed transcription factors involved in oligodendrogenesis, such as Sox9, Olig2 and Sox10. These data indicate that gliogenesis occurs also in MS SVZ and suggest the recruitment of SVZ-derived oligodendrocyte precursors to peri-ventricular demyelinated lesions. Further investigation of adult neural stem cells and their progenitors in the brain of rodents and non-human primates should help to gain insights in their process of activation in response to demyelination and their role in myelin repair.

[Promoting myelin repair in disorders such as multiple sclerosis and some types of leukodystrophy: current studies]. / Approche expérimentale des stratégies réparatrices des maladies demyélinisantes du système nerveux central.

Several ways of promoting myelin repair in myelin disorders such as multiple sclerosis and certain types of leukodystrophies are currently being investigated. Numerous studies suggest that it is possible to repair the central nervous system (CNS) by cell transplantation or by enhancing endogenous remyelination. Investigations in animal models indicate that cell therapy results in robust anatomical and functional recovery of acute myelin lesions. These models are also used to explore and validate the role of candidate molecules to stimulate endogenous remyelination by activating the myelin competent population or providing neuroprotection. However, in view of the heterogeneity of the lesion environment in MS, it seems more likely that cell therapy alone will not be able to contribute efficiently to the repair of the lesion. Further developments should indicate whether combining multiple approaches will be more powerful to achieve global myelin repair in the CNS than applying these strategies alone.

Complex patterns of GCM1 mRNA and protein in villous and extravillous trophoblast cells of the human placenta.

The Gcm1 gene encodes a transcription factor that is essential for both syncytiotrophoblast differentiation and formation of chorionic villi in mice. Its early expression is very unusual in that it defines a subset of trophoblast cells in the chorion, a layer that otherwise contains trophoblast stem cells. While Gcm1 mRNA expression initiates independently within the chorion, the subsequent maintenance of mRNA expression as well as the onset of protein accumulation is dependent on contact with allantoic mesoderm. Previous studies have shown that human GCM1 mRNA and protein are detectable in the placenta, but their patterns have not been compared nor precisely localized. We, therefore, conducted the present study to determine if the human mRNA and protein are subject to the same complexities of regulation as the mouse. In situ hybridization studies showed that the GCM1 mRNA was expressed in villous cytotrophoblast cells, but only a subset and never within cells immediately at the base of columns. Interestingly, the mRNA was detected throughout the cytotrophoblast columns. GCM1 protein expression studies demonstrated that the transcription factor was present mainly within the nuclei of a subset of cytotrophoblast cells, consistent with its role as a transcription factor. Feint cytoplasmic staining of the transcription factor was found in the syncytiotrophoblast but not in aggregated syncytial nuclei. Nuclear immuno-reactivity for the GCM1 protein was detected in occasional nuclei in the distal part of the column. Therefore, GCM1 expression is regulated both at the transcriptional and translational level. Overall, these studies show that the general features of GCM1 mRNA and protein expression in the human placenta are conserved with the mouse. They also highlight the fact that villous cytotrophoblast cells are extremely heterogeneous with respect to GCM1 expression, a factor that should be considered when using isolated cytotrophoblast cells for culture studies.

Oligodendrocyte precursor migration and differentiation: combined effects of PSA residues, growth factors, and substrates.

Using the oligosphere strategy (V. Avellana-Adalid et al., 1996, J. Neurosci. Res. 45, 558-570), we compared the migratory behavior of oligodendrocyte preprogenitors (OPP) that expressed the polysialylated form of the neural cell adhesion molecule (PSA-NCAM) and of GD3-positive oligodendrocyte progenitors (OP). To study the role of PSA in OPP migration, we used endoneuraminidase-N, which specifically cleaves PSA from NCAM. Kinetic data showed that (i) migration velocity decreased with time and was favored on polyornithine compared to Matrigel; (ii) cells emerging from spheres enriched in PSA-NCAM+ OPP migrated farther than those from spheres enriched in GD3+ OP, their migration being enhanced by the addition of growth factors; (iii) removal of PSA from NCAM moderately reduced OPP migration and induced their differentiation in GD3+ OP and GFAP+ astrocytes; (iv) blocking integrins reduced their migration, suggesting an alternative mechanism of migration. Altogether these data illustrate that motility and differentiation of OPP involve the combinatorial action of PSA-NCAM, molecules of the ECM and their receptors, and growth factors.

Placental expression and chromosomal localization of the human Gcm 1 gene.

Although gcm was first recognized for its role in specifying glial cell fate in Drosophila melanogaster, its mammalian counterparts are expressed predominantly in non-neural tissues. Here we demonstrate expression of the mouse and human GCM 1 proteins in placenta. We have prepared a highly specific antibody that recognizes the GCM 1 protein and have used it to assess the temporal and spatial expression profile of the protein. In both mouse and human placenta, the protein is associated with cells that are involved with exchange between maternal and fetal blood supplies: the labyrinthine cells of the mouse placenta and the syncytio- and cytotrophoblasts of the human placenta. Using the full-length hGcm 1 cDNA as a probe, we have mapped the gene on human chromosome 6p12 by fluorescent in situ hybridization.

Do central nervous system axons remyelinate?

In multiple sclerosis (MS), one of the most frequent demyelinating diseases in man, remyelination of demyelinating lesions exists but is often incomplete. Also reported in experimental models of demyelination, this phenomenom confirms the regenerating potential of the demyelinated central nervous system (CNS) and, in particular, the existence of an endogenous mechanism of oligodendrocyte renewal. Failure in efficient remyelination could result from exhaustion of the pool of remyelinating cells, loss of axons and absence of a permissive environment for remyelination. Identifying the nature and the origin of the cells capable of generating new oligodendrocytes for remyelination could contribute to strategies to activate these cells, and thereby enhance their potential for myelin repair. Within the adult CNS, several cell types are capable of generating new oligodendrocytes following myelin damage: post-mitotic oligodendrocytes frequently found at the lesion site, oligodendrocyte progenitors whose existence has been confirmed both in vitro and in vivo, and multipotent cells localized in the germinative areas of the brain and the spinal cord. Although restricted to particular sites of the CNS, these multipotent cells, which maintain the capacity to self-renew and to migrate throughout adulthood, could constitute a powerful source of remyelinating cells. The study of the mechanisms of proliferation, migration and differentiation of these cells in response to demyelination should allow the definition of new strategies to promote endogenous remyelination and develop therapeutic approaches for demyelinating diseases such as MS. This goal is an appealing alternative to the transplantation of myelin-forming cells and should efficiently complement strategies aimed at reducing neuronal loss and inflammation.

Progenitor cells of the adult mouse subventricular zone proliferate, migrate and differentiate into oligodendrocytes after demyelination.

Identifying a source of cells with the capacity to generate oligodendrocytes in the adult CNS would help in the development of strategies to promote remyelination. In the present study, we examined the ability of the precursor cells of the adult mouse subventricular zone (SVZ) to differentiate into remyelinating oligodendrocytes. After lysolecithin-induced demyelination of the corpus callosum, progenitors of the rostral SVZ (SVZa) and the rostral migratory pathway (RMS), expressing the embryonic polysialylated form of the neural cell adhesion molecule (PSA-NCAM), increased progressively with a maximal expansion occurring after 2 weeks. This observation correlated with an increase in the proliferation activity of the neural progenitors located in the SVZa and RMS. Moreover, polysialic acid (PSA)-NCAM-immunoreactive cells arizing from the SVZa were detected in the lesioned corpus callosum and within the lesion. Tracing of the constitutively cycling cells of the adult SVZ and RMS with 3H-thymidine labelling showed their migration toward the lesion and their differentiation into oligodendrocytes and astrocytes but not neurons. These data indicate that, in addition to the resident population of quiescent oligodendrocyte progenitors of the adult CNS, neural precursors from the adult SVZ constitute a source of oligodendrocytes for myelin repair.

Murine Gcm1 gene is expressed in a subset of placental trophoblast cells.

The gcm gene of Drosophila melanogaster encodes a transcription factor that is an important component in cell fate specification within the nervous system. In the absence of a functional gcm gene, progenitor cells differentiate into neurons, whereas when the gene is ectopically expressed the cells produce excess glial cells at the expense of neuronal differentiation. Recent searches of databases have uncovered high sequence similarity between the Drosophila gem gene and an anonymous human placental cDNA clone (Altschuller et al., 1996; this communication). Here we report the molecular organization of the murine Gcm1, its spatio-temporal pattern of expression in developing placenta, and its map position at E1-E3 on murine chromosome 9. The murine gene is composed of at least 6 exons. The promoter region contains an "initiation sequence" and is GC rich, characteristics of the promoters of several transcription factors. The mRNA has a modest 5'UTR (ca. 200 bases) but an extensive 3' UTR (ca. 2 kb). Northern blot and mRNA in situ hybridization studies showed that Gcm1 expression was readily detectable only in the placenta. It began at embryonic day 7.5 within trophoblast cells of the chorion and continued to about embryonic day 17.5 within a subset of labyrinthine trophoblast cells. Comparison with other transcription factors revealed that Gcm1 expression defines a unique subset of trophoblast cells.

Developmental expression of platelet-derived growth factor alpha-receptor in neurons and glial cells of the mouse CNS.

The synthesis of platelet-derived growth factor-alpha receptor (PDGF-alphaR) is commonly attributed to oligodendrocyte progenitors during late embryonic and postnatal development. However, we recently demonstrated that mature neurons could also synthesize PDGF-alphaR, emphasizing a larger role for this receptor than previously described. In the present study, to analyze the pattern of PDGF-alphaR expression during postnatal development of the mouse CNS, we used in situ hybridization and immunohistochemistry on brain and spinal cord tissue sections. We found that, in addition to immature cells of the oligodendrocyte lineage, neurons of various CNS regions express PDGF-alphaR transcripts and protein as early as postnatal day 1 (P1). Whereas neuronal expression was maintained at all ages, the oligodendroglial expression strongly decreased after P21. In the adult, PDGF-alphaR was detected in very few oligodendrocyte progenitors scattered in the cerebral cortex or in white matter tracts, thus suggesting the presence of PDGF-alphaR on O-2Aadult progenitors. In the mature CNS, PDGF-alphaR transcripts and protein were mainly localized in neurons of numerous structures, such as the olfactory bulb, cerebral cortex, hippocampus, and brainstem nuclei and in motor neurons of the ventral horn of the spinal cord. The differential expression of PDGF-alphaR in oligodendroglia and neurons argues in favor of several roles of PDGF during development.

Expansion of rat oligodendrocyte progenitors into proliferative "oligospheres" that retain differentiation potential.

The limited availability of enriched populations of oligodendroglial progenitors has impeded the exploration of the complex spatio-temporal mechanisms which dictate the chemical "language" of their biology. We have developed a technique to prepare homotypic aggregates of oligodendrocyte progenitors called "oligospheres." These were obtained using various approaches (sieving, Percoll gradient separation and differential adhesion) to purify oligodendroglial progenitors from newborn rat brain. Culturing cells in a mixture of N1 defined medium and conditioned medium from the B104 neuronal cell line in the absence of adhesive substrate allowed to expand routinely and extensively for several months, the oligodendrocyte progenitor population. Under these conditions, the resulting population consisted of 98% GD3-positive/GFAP-negative cells. After dissociation and plating on polyornithine coated substrates, in the presence of low (2%) or high (20%) serum, oligosphere-derived oligodendrocyte progenitors were induced to differentiate into GalC-positive oligodendrocytes or GFAP-positive astrocytes, respectively. When transplanted into the newborn shiverer mouse brain, oligospheres were able to provide a focal reservoir of migrating and myelinating cells. Oligospheres are thus ideal tools for exploring the biological and molecular events of the oligodendrocyte lineage both in vitro and in vivo.

Cell-cell interactions during the migration of myelin-forming cells transplanted in the demyelinated spinal cord.

In the present paper, Dil-labeled myelin-forming cells were traced after their transplantation at a distance from a lysolecithin induced lesion in the adult wild-type and shiverer mouse spinal cord. Optical and ultrastructural observations indicate that after their transplantation, Dil-labeled Schwann cells and oligodendrocyte progenitors were found at the level of the graft as well as at the level of the lesion thus confirming that myelin-forming cells were able to migrate in the adult lesioned CNS (Gout et al., Neurosci Lett 87:195-199, 1988). Between the graft and the lesion, labeled Schwann cells and oligodendrocyte progenitors were absent in the gray matter, but were found as previously described, in specific locations (Baron-Van Evercooren et al., J Neurosci Res 35:428-438, 1993; Vignais et al., J Dev Neurosci 11:603-612, 1993). Both cell types were found along blood vessel walls and more precisely in the Virchow-Robin perivascular spaces. They were identified in the meninges among meningeal cells, collagen fibers, or occasionally in direct contact with the basement membrane forming the glia limitans. In addition to these findings, three major observations were made. In the ependymal region, myelin-forming cells were localized between or at the basal pole of ependymocytes. While Dil-labeled oligodendrocyte progenitors were noted to migrate along the outer surface of myelin sheats in CNS wild-type and shiverer white matter, Schwann cells were excluded from this structure in the wild-type mouse spinal cord. Moreover, in the shiverer mouse, migrating Schwann cells did not seem to interact directly with myelin sheats nor with mature oligodendrocytes. Finally, both cell types were seen to invade extensively the spinal peripheral roots. Our ultrastructural observations clearly suggest that multiple cell-cell and cell-substrate interactions rule the migration of myelin-forming cells in the adult CNS infering that multiple mechanisms are involved in this process.

PDGF-alpha receptor is expressed by mature neurones of the central nervous system.

The platelet-derived growth factors (PDGF) are constitutively expressed by neurones in the central nervous system (CNS). The synthesis of the PDGF-alpha-receptor (PDGF-alpha R) is commonly attributed to oligodendrocyte precursors during late embryonic and early postnatal development, suggesting communication between neurones and glia which orchestrates amplification and final targeting of the myelinating cells. However PDGF A production persists when central myelination is achieved, which suggests that PDGF-alpha R are present in the adult CNS. In this study, we demonstrate the production of PDGF-alpha R transcripts and protein by various neuronal populations of the adult CNS. We propose a developmental shift, where glial cells and neurones are consecutive targets of PDGF A and neuromodulatory effects of PDGF, exerted on mature neurones via the expression of the PDGF-alpha R.

Expression of the highly polysialylated neural cell adhesion molecule during postnatal myelination and following chemically induced demyelination of the adult mouse spinal cord.

We have investigated the expression of the highly polysialylated neural cell adhesion molecule in the mouse spinal cord during postnatal myelination and in the adult after chemically induced demyelination. By double immunohistochemistry, using a monoclonal antibody (anti-Men B) which specifically recognizes polysialic acid (PSA) units on neural cell adhesion molecule (N-CAM), and an anti-myelin basic protein, a caudorostral gradient of expression of PSA-NCAM was observed at postnatal day 1 (P1), which was inversely related to the gradient of myelination. At P7, PSA-NCAM labelling decreased relative to P1. In white matter, this decrease was correlated with the progression of myelination. PSA-NCAM immunoreactivity persisted in as yet unmyelinated structures, i.e. the corticospinal tract, the dorsomedial part of the ventral funiculus and the lateral funiculi, and decreased with the onset of myelination of these structures at P15. In the adult, PSA-NCAM expression remained in discrete structures, i.e. laminae I and II of the dorsal horn and lamina X around the central canal. The ependymal cells and the astrocyte endfeet under the meninges were also labelled. In addition, PSA-NCAM expression was reinduced on various cells and structures after lysolecithin-induced demyelination of the adult mouse spinal cord. At early times after demyelination, PSA-NCAM was expressed on glial cells around the lesion but also at a distance from this zone. Seven days after injection, cellular PSA-NCAM expression was found around but also within the lesion. This expression was totally abolished 15 days after injection. Double immunohistochemistry for PSA and cell-specific markers showed that the cells which expressed PSA-NCAM after demyelination were oligodendrocyte precursors, reactive astrocytes and Schwann cells. PSA-NCAM re-expression on all cell types was transient and ceased when myelin repair was accomplished. The spatial and temporal regulation of PSA-NCAM expression during development and after demyelination suggests a role for PSA-NCAM in glial plasticity during the myelination and remyelination processes.

Transplantation of oligodendrocyte precursors in the adult demyelinated spinal cord: migration and remyelination.

A demyelinating lesion induced by an injection of lysolecithin into the spinal cord can be partly repaired by oligodendrocyte precursors transplanted at a distance of 6-8 mm from the lesion. Using a non-toxic fluorescent dye (Hoechst 33342) as a cell marker, we demonstrate that transplanted oligodendrocyte precursors from different origins (periventricular zone fragments from newborn mouse and cultured rat oligodendrocyte progenitor cells) can migrate along specific pathways (i.e. white matter fasciculi, ependymal wall, meninges and blood vessels). These cells can be attracted when passing at the vicinity of the lesion as well as differentiate and remyelinate axons with the lesion. Myelin repair thus appears to be the result of distinct successive events: migration, specific attraction, differentiation and myelination. This can occur in both shiverer and normal adult hosts.