Re-evaluation of nestin as a marker of oligodendrocyte lineage cells

Maturation of oligodendrocyte progenitors (O2A) is characterized by morphological changes and the sequential expression of specific antigens leading to the formation of myelin membrane. Monoclonal antibodies A2B5, A007, anti-vimentin, and anti-galactocerebroside, recognize oligodendroglia at different stages of development. The neuroepithelial precursor marker nestin is also expressed by the oligodendroglial lineage; we have used enriched populations of progenitors isolated from neonatal rat brain cultures to further examine the cellular distribution of this intermediate filament protein. The phenotypic distribution of nestin positive cells among the oligodendrocyte lineage showed that 65% reacted with A2B5, whereas only 5% were A007(+), and 4% galactocerebroside(+). The remaining 25% of the cells were not labeled and had small cellular bodies devoid of processes, characteristic of the pre-O2A progenitor. Further analysis of the nestin(+) population showed that the majority of the cells were also vimentin(+). Antibody-dependent complement mediated cytolysis of A2B5(+) (O2A cells) and galactocerebroside(+) (mature oligodendrocytes) cells left a population of nestin(+) cells that were induced to proliferate in the presence of growth factors and to differentiate into A2B5(+) and galactocerebroside(+) cells. Proliferating cells maintained in the presence of platelet-derived growth factor or basic fibroblast growth factor retained nestin expression along with A2B5. By contrast, in serum-free medium nestin expression decreased while postmitotic cells acquired A007 and galactocerebroside. Our results suggest that nestin expression is a marker of pre-O2A cells that is maintained in proliferating glial progenitors, but is quickly down-regulated in postmitotic oligodendrocytes (A007(+)/galacto-cerebroside(+)) along with A2B5 and vimentin. However, other glial cells including type 2 astrocytes and some amoeboid microglia also share nestin expression.     

Characterization of GFP-MAL expression and incorporation in rafts

During myelin formation, membrane-associated proteins have to be sorted and transported in specified membrane regions such as compact and non-compact myelin membranes. One protein that may be involved in such a process is the Myelin and Lymphocyte protein MAL (VIP17/ MVP17). MAL was identified as a novel myelin membrane component expressed by oligodendrocytes and Schwann cells. Since MAL has been shown to be important in the apical sorting machinery of polarized cells, we have started to investigate the possible functional role of MAL in sorting myelin membrane-associated molecules. In this study, we have generated cDNA constructs with green fluorescent protein (GFP) either at the N- or C-terminus of MAL. Transfection experiments showed that GFP-MAL expression resembles that of normal MAL, whereas the MAL-GFP fusion construct was not properly transported within the cell. Furthermore, we could demonstrate that GFP-MAL is enriched in detergent insoluble glycolipid-enriched microdomains as already seen for untagged MAL. As a prerequisite for the generation of transgenic mice expressing GFP-MAL under the control of its own regulatory elements, we have generated a cDNA construct with an 8-kb MAL promotor fragment fused to GFP-MAL. Transfection experiments of the Oli-neu oligodendrocyte cell line showed that GFP-MAL was expressed, but only in cells, which were stimulated for differentiation with cAMP. In summary, the results confirm that the fusion protein GFP-MAL is incorporated into detergent-insoluble complexes and the 8-kb MAL promotor fragment is sufficient to be activated in oligodendrocytes.     

Calcium signalling in cells of oligodendroglial lineage

Intracellular Ca2+ is the key signal that regulates the efficacy of neurotransmitter release and synaptic plasticity in neurons but is also an important second messenger involved in the signal transduction and modulation of gene expression in both excitable and non-excitable cells. Glial cells, including cells of oligodendroglial (OLG) lineage, are capable of responding to extracellular stimuli via changes in the intracellular Ca2+. This review article focuses on the mechanisms of Ca2+ signalling in cells of OLG lineage with the goal of providing the basis for understanding the relevance of receptor- and non-receptor-mediated signalling to oligodendroglial development, myelination, and demyelination. Conclusions to date indicate that cells of OLG lineage exhibit remarkable plasticity with regard to the expression of ion channels and receptors linked to Ca2+ signalling and that perturbation of [Ca2](i) homeostasis contributes to the pathogenesis of demyelinating diseases.     

Membrane traffic in myelinating oligodendrocytes

In the central nervous system (CNS), the myelin sheath is synthesised by oligodendrocytes as a specialised subdomain of an extended plasma membrane, reminiscent of the segregated membrane domains of polarised cells. Myelination takes place within a relatively short period of time and oligodendrocytes must have adapted membrane sorting and transport mechanisms to achieve such a high rate of myelin synthesis and to maintain the unique organisation of the myelin membrane. In adult life, maintenance of the functional myelin sheath requires a carefully orchestrated balance of myelin synthesis and turnover. Imbalance in these processes may cause dys- or demyelination and disease. This review summarises what is currently known about myelin protein trafficking and mistrafficking in oligodendrocytes. We also present data demonstrating distinct transport pathways for myelin structural proteins and the expression of SNARE proteins in differentiating oligodendrocytes. Myelinating glial cells may well serve as a model system for studying general aspects of membrane trafficking and organisation of membrane domains.     

Cell lineage of the subcommissural organ secretory ependymocytes: differentiating role of the environment

SCO-ependymocytes have a secretory activity and a neural innervation relating them to neurosecretory nerve cells. To elucidate the cell lineage of the SCO-ependymocytes and emphasize the role of the neural innervation in their differentiation, in particular 5-HT innervation, we analyzed the developmental pattern of expression of several glial and neuronal markers: (1) in the SCO of mammals possessing (rat, cat) or devoid (mouse, rabbit) of 5-HT innervation, (2) in rat 5-HT deafferented SCO, and (3) in rat SCO transplanted in a foreign environment, the fourth ventricle. The ability of SCO-ependymocytes to transiently express GFAP during development and express the glial alpha alpha-enolase confirms the glial lineage of the SCO-ependymocytes. Synthesis of vimentin by SCO-ependymocytes relates them to the classical ependymocytes. The ability of mature SCO-ependymocytes to take up GABA only when they are innervated by 5-HT terminal underlines the role of the neural environment on the differentiation of these ependymocytes and suggests that differential maturation of the SCO according to its innervation, may lead to specific functional specialization of this organ in different species.     

Comparative characterization of human fetal neural stem cells and induced neural stem cells from peripheral blood mononuclear cells

Human-induced neural stem cells (iNSCs) transplantation is a potential treatment of neurodegeneration diseases. However, whether the reprogrammed cells have the same characterizations as human fetal neural stem cells needs further exploration. Here we isolated human fetal neural stem cells from aborted 12-week fetal brains and compared with iNSCs reprogrammed from human peripheral blood mononuclear cells in gene expression, proliferation ability, differentiation capacity, and the responses to tumor necrosis factor-α. We found that iNSCs and NSCs both expressed neural stem cell markers Nestin, SOX1, and SOX2. However, only iNSCs can be patterned into dopaminergic neurons and motor neurons. Furthermore, both iNSCs and NSCs can differentiate into oligodendrocyte progenitor cells. In addition, a low dose of tumor necrosis factor-α did not inhibit the proliferation and differentiation of iNSCs and NSCs. In conclusion, iNSCs have properties similar to, and even better than, fetal neural stem cells and may be suitable for disease modeling and transplantation.     

Muscle-specific gene expression during myogensis in the mouse

Over the past decade, significant advances in molecular biological techniques have substantially increased our understanding of in vivo myogenesis, supplementing the information that previously had been obtained from classical embryological and morphological studies of muscle development. In this review, we have attempted to correlate morphogenetic events in developing murine muscle with the expression of genes encoding the MyoD family of myogenic regulatory factors and the contractile proteins. Differences in the pattern of expression of these genes in murine myotomal and limb muscle are discussed in the context of muscle cell lineage and environmetal factors. The differences in gene expression in these two types of muscle suggest that no single coordinated pattern of gene activation is required during the initial formation of the muscles of the mouse. © 1995 Wiley-Liss, Inc.     

Differentiation of Leydig cells in the Mongolian gerbil

Information on postnatal Leydig cell (LC) differentiation in the Mongolian gerbil has been unavailable. Therefore, current investigation was designed to examine LC lineage differentiationin this rodent, from birth to adulthood. Gerbil testes at 1 day, 1–7 weeks (w), 2 and 3 months of age were conventionally processed by light and transmission electron microscopy. Immunocytochemistry for specific markers of steroidogenic enzymes, namely 3β‐hydroxysteroid dehydrogenase (3β‐HSD) and 11β‐hydroxysteroid steroid dehydrogenase 1 (11β‐HSD1) and also for androgen receptor (AR) was performed. The establishment of adult Leydig cell populations (ALC) during testis maturation in the gerbil follows the pattern previously described in other mammalian species, with the four progressive stages of differentiation. The LC progenitors were identified at second w by 3β‐HSD expression; the first newly formed ALC were recognized at fourth w whereas immature ALC appeared at fifth w. The latter were recognized by abundance of cytoplasmic lipid, in addition to expression of 11β‐HSD1 and intense nuclear AR immunoreaction. Mature ALC in gerbil exhibited irregular eccentric nuclei and a cytoplasmic canaliculus in the perinuclear area. Only one third of mature ALC in adult gerbils showed a high expression of 11β‐HSD1 and AR expression was highly variable among them. In conclusion, the process of differentiation of ALC population in gerbil follows the pattern previously established for other rodents. However, the resulting mature ALC are strikingly different due their singular asymmetric morphology and presence of a cytoplasmic canaliculus and as well as their functional heterogeneity. Microsc. Res. Tech., 2010. © 2009 Wiley‐Liss, Inc.     

Stem cells for tissue engineering of articular cartilage

Articular cartilage injuries are one of the most common disorders in the musculo-skeletal system. Injured cartilage tissue cannot spontaneously heal and, if not treated, can lead to osteoarthritis of the affected joints. Although a variety of procedures are being employed to repair cartilage damage, methods that result in consistent durable repair tissue are not yet available. Tissue engineering is a recently developed science that merges the fields of cell biology, engineering, material science, and surgery to regenerate new functional tissue. Three critical components in tissue engineering of cartilage are as follows: first, sufficient cell numbers within the defect, such as chondrocytes or multipotent stem cells capable of differentiating into chondrocytes; second, access to growth and differentiation factors that modulate these cells to differentiate through the chondrogenic lineage; third, a cell carrier or matrix that fills the defect, delivers the appropriate cells, and supports cell proliferation and differentiation. Stem cells that exist in the embyro or in adult somatic tissues are able to renew themselves through cell division without changing their phenotype and are able to differentiate into multiple lineages including the chondrogenic lineage under certain physiological or experimental conditions. Here the application of stem cells as a cell source for cartilage tissue engineering is reviewed.