Biological Characteristics Of Oligodendrocyte Lineage Cells And Their Neuroprotective Effects

Oligodendrocyte lineage cells including both oligodendrocytes and oligodendrocyte precursor cells, are the important cellular parts of CNS, especially in the white matter. A lot of reports confirmed that the myelin of oligodendrocyte could inhibit the regeneration of injured axons in the CNS, but less attention on the trophic and protective roles of oligodendrocyte lineage cells in the CNS, especially in the mature CNS. On the other hand, recent experiments indicated perinatal OPCs have the ability to proliferate indefinitely in appropriate culture conditions, reprogrammed to neural stem cells, and the adult OPCs in the hippocampus make synaptic junctions with the axons terminals of neurons. These phenomena indicated that the oligodendrocyte lineage cells maybe act as a more important and complicated role in vivo. Taking advantage of 'cell culture and transplantation, retrograde fluorescent labeling the superior colliculus, optic nerve transection and myelinformation in the retina, we studied the development, differentiation and neuroprotective effects of oligodendrocyte lineage cells in vitro and in vivo, to further clarify the relationship between oligodendrocyte lineage cells and neurons, and seek for a new approach for the therapy of correlated diseases.We introduced improved primary mixed glial culture and different-attachment method to isolate and purify the OPCs, the cells were proliferated in serum-free medium, flow cytometry and immunohischemistry methods were employed to estimate the purity of cultured OPCs. Their abilities of differentiation and expression of trophic factors were identified by RT-PCR and immunostaining. Several methods including TUNEL and MTT were adopted to estimate the protective effects of conditioned culture medium from oligodendrocyte lineage cells on the primary cultured cerebellar granule neurons. Intravitreal transplant of OPCs, combined with retrograde fluorescent labeling the superior colliculus (SC) and intraorbital optic nerve transection, were used to investigate the protective effects of OPCs on the axotomized RGCs in vivo. Intravitreal transplantof OPCs or NSCs on the newborn rats, and retinal transplant of OPCs on the young rats were performed, to observe the myelin formation in the retina at different stages after cellular transplantation. Optic nerve transection was carried out on some rats with myelinated retinae, to study the influence of myelination on the injuried RGCs. The results and conclusions are as follows:1. Using the modified mixed glial culture and different-attachment method, we harvested OPCs with purity more than 93%. The cultured OPCs could express Nestin and Gap-43. OPCs automatically differentiated into mature oligodendrocytes after PDGF withdrawn. Short-term exposed to serum could stimulated the proliferation of OPCs. Both OPCs and OLGs could express BDNF and IGF-1 at mRNA and protein levels. The conditioned culture medium promoted the survival of primary cultured cerebellar granule neurons.2. The intravitreal transplanted OPCs could survive for a long time, the morphology of partial cells became multi-polar. The density of survival RGCs in the OPCs transplantation group is higher than in the control groups within 2 weeks after optic nerve transection, indicating that the oligodendrocyte lineage cells have the ability to save the injured neurons.3. 4 weeks after OPCs transplanted into the vitreous space of newborn rats, bundles of myelinated axons were observable, indicating that OPCs could differentiate and mature in the homological retina. The myelination limited within the NFL but the distributed areas of myelinated bundles in the retina is expanded with prolonged survival time after cellular transplantation. Striatal neural precursor cells could differentiate into oligodendrocytes and produce myelin after transplanted into the vitreous space.4. Bundles of myelinated axons were found in nearly half of the retinae of young rats received retinal OPCs transplantation 7 weeks later. Most of the bundles distributed within the superonasal quarter of retina where transplantation were performed. Electron microscope and immunohistochemistry inspection confirmed no obvious RGCs degeneration within the retina. The myelin sheaths revealed normal CNS morphology and the thickness were diverse. The myelinated axons had larger calibers comparing with unmyelinated axons. The amount of survival RGCs within the myelinated sectors were higher than the control groups within 10 daysafter optic transection. At the same time, the myelinated axon bundles suffered gradual disintegration. The results indicated that mature oligodendrocytes with their myelin sheats maybe play protective roles in vivo.