老鼠皮肤前体细胞中侧群细胞的分选及特性分析-外科学(手外科)专业论文.docxVIP

华中科技大学硕士学位论文 华 中 科 技 大 学 硕 士 学 位 论 文 4 4 Isolation and Characterization of side population cells from Skin-Derived Precursors Abstract Objective investigate and isolate the SP cell from the SKPs，analyse the ability of its proliferation and differentiation. Method the cultured SKPs population were harvested and stained by fluorochrome Hoechst3334 and PI，Controlled with the Presence of verapami , the SP cells were analysed and isolated by fluorescence actived cell sorting (FACS).The harvested SP cells and MP cells were cultured in the medium with growth factor.the SP cells at passage 2 were harvested ，resuspended and divided into 24 well plates. then differentiate SP cells into neuron-like cells and schwann cells.SP cells and Differentiated cells were analyzed by means of reverse transcriptase-polymerase chain reaction (RT-PCR), immunofluorescence to identify the expression of neural specific markers. Result SP cells(termed skin-derived precursors-side population[SKP-SP])were isolated from SKPs by FACS with a proportion of 0.1%.Compared to the MP cells,the SP cells have greater capacity of proliferation,could be expanded for long periods of time in culture. The result of RT-PCR and immunofluorescence showed that SP cells expressed the SKP specific markers. nestin and fibronectin.Differentiated cells were positive the neural specific markers NeuN，NF-M,and the schwann cells specific markers GFAP，S100. Conclusion SP cells which were isolated from SKPs are multipotent stem cells，and represent the real SKPs.SKP-SP cells provide an solution to get the highly purified cells quickly for the people who need stem cell therapy. 【Key words】 Side population SKPs FACS Neurons Schwann cells PAGE 1 PAGE 1 中英文缩略词表 缩写 英文全称 中文全称 SP Side population 侧群细胞 MP Main population 主群细胞 SKPs Skin-Derived precursors cells 皮肤来源前体细胞 SKP-SP Skin-Derived precursor cells-Side Population SKP-MP Skin-Derived precursor cells-Main Population 皮肤前体细胞的侧群细胞 皮肤前体细胞的主群细胞 pSSCs