| Repair of injured adult mammalian CNS is one of the most challenging fields in neuroscience research. Tissue and cell transplantations are important methods to enhance CNS regeneration and have great potential for clinical application. The tissue and cells that have been transplanted and have beneficial effects are embryonic nervous tissue, neural stem cells. Schwann cells, olfactory ensheathing cells (OECs). In recent years, transplantation of OEC has been proved to be particularly promising. Furthermore, there is a growing consencus that for successfal clinical therapy combination of various methods is necessary. The present study was first to work out the methods of OEC culture, purification, and characterization for OEC transplantation. On top of that method to produce transgenic NT-3, a member of the neurotrophin family that has been proved to facilitate regeneration of the descending was studied, so that transplantation of NT-3 transgenic OEC can has both the bridging effect of OEC and neurotrophic function. This study includes 4 parts: Part I Isolation, purification and morphology of the OECs The OECs were isolated from the olfactory bulb and purified. A purity of the >90% of OECs was achieved. And their properties were studied. (l).The morphology and viability of the OECs varies with time of culture. Before 6 days the OECs were macrophage-like, large and fusiform, with short processes. As the culture time prolonged the cells became more refractive when examined under phase contrast microscope, more and more cells turned to be tripolar or multipolar. The processes were much more longer. (2). Between the 2th to 20th day, the viability of the OEC remained nonnal. After 20 days the viability was deteriorated, as evidenced MTT assay. Part II Neurotrophic effect of OEC The neruotrophic effects of the OEC on spinal cord GABAergic neurons were studied by co-culture method. Culture of GABAergic neurons without OECs served as control. The number of the GAflAergic neurons in the co-cultured group was significantly higher than that of the control group, 39.7 ?6.3 vs 27.6 + 2.7/0.45 mm2. The length of the neurites was 630 + 112 j.tmin the co-cultured group vs 270 ?97pm in the control. Part III Cytoprotective effect of OEC The potential cytoprotective effects of OECs on apoptosis ofPCl2 cells induced by serum deprivation are of considerable interest of our study. PC 12 cells can be induced to turn to neurons in serum cultured. When deprived of serum, the PC12 cells undergo apoptosis. This character was utilized to study the protective effect of S OEC on PCI2 cells. By adding supernatant of the OEC culture into serum deprived PC12 culture, the survival of the PC12 cells was assessed, as analyzed by MIT assay. The morphological changes of apoptotic PC12 cells were examined with light microscopy, electron microscopy, and fluorescence microscopy. ,At same time, apoptotic DNA fragmentation was determined with DNA electrophoresis.The number of apoptotic cells was detected with flow cytometer in 4 groups of PC12 cultures, namely, PC12 with OEC supematant; Co-culture of PC12 and OEC; Serum deprived PC12; PCI2 with serum. The results are as follows (1) the survival of the PC12 cells was significantly increase with OEC supernatant .(2)the distinct morphological changes showed in different groups. (3) the percentages...
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