人脐带间充质干细胞在壳聚糖支架上向类髓核细胞分化的实验研究-外科学(骨外科学)专业论文.docx

人脐带间充质干细胞在壳聚糖支架上向类髓核细胞分化的实验研究中文摘要【关键词】脐带间充质干细胞；作者：董仁斌 指导老师：唐天驷教授英文摘要人脐带间充质干细胞在壳聚糖支架上向类髓核细胞分化的实验研究Human umbilical cord-derived mesenchymal stem cells differentiation to NP-like cells inchitosan-glycerophosphate hydrogels AbstractChapterⅠ In vitro isolation、culture、identification and differentiation ofhuman umbilical cord-derived mesenchymal stem cellsObjective: To explore the methods of ilsolating、culturing、identification and differentiation of mesenchymal stem cells from human umbilical cord. Methods: Human umbilical cord from 8 neonatus with cesarean section surgery were collected to extract human umbilical cord-derived mesenchymalstem cells (hUC-MSCs) using digestion by 0.075% collagenase I at 37℃ temperature. During the wholeculturing and passaging period, morphologic characteristics of hUC-MSCs were observed under inverted microscope. Flow cytometry assaying was used to detect the expression of cell surface markerslike CD90、CD105、CD44、CD34、CD45 and HLA-DR. After identification, hUC-MSCs were inducedby specific inducing medium to detect their differentiating potentials towards osteocyte、adipose and chondrocyte. After induction for 21 days, Alizarin red S staining、Oil red staining、Alcian blue staining and immunocytochemistry staining of collagen Ⅱ were used to evaluate their differentiation. Results: hUC-MSCs from umbilical cord of neonatus could be extracted by digestion of 0.075% collagenase Ⅰ.It took an average of 3 days for the polygonal primary hUC-MSCs to achieve adherence and about 14 days to reach a confluence of 80%. After 3 passaging, the polygonal cells gradually changed intospindle-like cells, which demonstrated a swirling clustering. They had positive expression of CD90、CD105、CD44 and negative expression of CD34、CD45、HLA-DR surface markers. The hUC-MSCs had positive results of Alizarin red S、Oil red、Alcian blue staining and immunocytochemistry staining of collagen Ⅱ after an induction period of 21 days. Conclusions: hUC-MSCs can be isolated from umbil