| The treatment of spinal cord injury(SCI) has been a major neurological problems in the world. Because spinal cord derived neural progenitor/stem cells (NPCs) possess the ability to self-renew and are multipotential for both neuron and glia, transplanting these stem cells to repair SCI has become the focus of the research in recent years. A series of physiological and pathological events happening after acute SCI, under the circumstance, endogenous NPCs could reactively proliferate and migrate toward the damaged areas and then differentiate into neurons and glia cells. But the details remain unclear. In previous study, we isolated and identified 39 proteins which differentially expressed at ifferent time points after SCI using the two dimensional electrophoresis and MS. Galectin-3 was one of the most interesting proteins, however, the details of its effect on the repairing after SCI were rarely reported. The purpose of this study is to investigate the tissue expression of Gal-3 after acute SCI at different time points and to further explore the effect of Gal-3 on the proliferation, apoptosis and neuronal differentiation of NPCs. Part 1: To investigate the tissue expression of Gal-3 after acute SCI at different time points. We employed a generally accepted and reproducible MASCIS impactor to establish the rat SCI model. Through real time quantitation PCR(qPCR) and western-blot, we analyzed the protein and mRNA expression levels of Gal-3 at 8h, 1d, 3d and 5d after SCI. Result: Gal-3 mRNA expression became weaker at 8h after injury and reverted to a higher level than the contral group at 1d post-injury. The expression increased over the next several days and peaked at 5d after injury. Compared with the control group, Gal-3 mRNA expression increased after injury and peaked at 3d after injury.Part 2: To purify and identify rats embryonic spinal cord-derived NPCs .By using neurosphere culture method, NPCs were isolated from E14 fetus rat's spinal cord and purified after sveral passages. Both differentiation assay and Immuno- histochemistry were investigated to identify NPCs. Result: After 3 passages, more than 98% cells of the neurosphere were nestin postive, and the cells can differentiate into neurons(Tuj1+) and astrocytes(GFAP+). All the result showed that these cells were NPCs.Part 3: To investigate the effect of galectin-3 on rats embryonic spinal cord-derived NPCs in vitro. After three passages, the effects of Gal-3 on the NPCs survival and apoptosis were determined by WST-8 assay and Hoechst 33258 stain. Also, anti-βtubulinⅢ(Tuj1) immunostaining was used to label the neurons differentiated from NPCs, and the percentage of Tuj1 positive cells among hoechst positive cells in the Gal-3 and control group were compared. Result: Compared with the control group, no significant effect was observed on the growth and apoptosis of NPCs under the presence of different concentration of Gal-3(1μg/ml, 4μg/ml and 8μg/ml) in experimental groups(P>0.05). However, Gal-3 could induce the differentiation of NPCs into neuron(P﹤0.01), the proportion of neurons in experimental groups and control group was 7.23%±1.07%, 15.1%±1.56%, 14.57%±2.01% and 15.3%±2.3%, respectively. But on significant differences were observed among the three concentrations (0.5μg/ml, 1μg/mland 4μg/ml) (P﹥0.05). Conclusion: Different concentration of Gal-3 could not affect the proliferation and apoptosis of NPCs, but have the effect of promoting the differentiation of NPCs into neurons.
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