三维培养诱导人脂肪间充质干细胞微球的成软骨分化能力

背景：关节软骨损伤后修复结果不满意，需要新的手段，而脂肪间充质干细胞较适宜做种子细胞诱导软骨，然而怎么能够使诱导的软骨具有功能需要研究。目的：采用三维培养体系诱导人脂肪间充质干细胞微球向软骨分化。方法：无菌切取吸脂术后脂肪组织，分离培养人脂肪间充质干细胞，传至第3代进行流式细胞术分析，成骨成脂肪诱导等鉴定，同时也给予合适的培养条件用三维培养的方式向软骨细胞诱导，并行阿利辛蓝染色鉴定糖胺多糖的合成，苏木精-伊红染色进行组织学分析，免疫荧光检测Ⅱ型胶原表达，称质量测量软骨硬度。结果与结论：分离的人脂肪间充质干细胞CD105，CD44，CD29均高表达，而 CD45，CD34低表达，并且成骨成脂诱导后细胞茜素红染色和油红O染色均为阳性。三维培养法诱导的软骨细胞可表达大量糖胺多糖及Ⅱ型胶原。结果证实，三维培养法诱导人脂肪间充质细胞向软骨分化后，具有软骨细胞的特性。中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程全文链接：

Three-dimensional culture method induces the chondrogenic differentiation of human adipose-derived mesenchymal stem cell microspheres

BACKGROUND:Adipose-derived mesenchymal stem cells are more appropriate as seed cells for cartilage induction in patients who have unsatisfied repair after articular cartilage injuries, but how to induce the cartilage function need to be studied.OBJECTIVE:To induce the chondrogenic differentiation of human adipose-derived mesenchymal stem cell microspheres using three-dimensional culture system.METHODS:Adipose tissues were extracted after liposuction under sterile, to isolate and culture human adipose-derived mesenchymal stem cells. The third passage cells were analyzed by flow cytometry and identified by osteogenic induction and adipogenic induction, and meanwhile, cells were cultured under three-dimensional culture system for chondrogenic differentiation. Then, cells were observed by Alcian blue staining for identification of glycosaminoglycan synthesis as well as hematoxylin-eosin staining for histological analysis. Expression of collagen type II was detected by immunofluorescence method. Cartilage stiffness was determined by mass measurement. RESULTS AND CONCLUSION:Isolated human adipose-derived mesenchymal stem cells highly expressed CD105, CD44, CD29, but showed low expressions of CD45, CD34. After osteogenic and adipogenic induction, cells were positive for alizarin red staining and oil red O staining. Under the three-dimensional culture, chondrocytes could greatly express glycosaminoglycans and collagen type II. The results suggest that under the three-dimensional culture, human adipose-derived mesenchymal cells differentiate into chondrocytes and have the characteristics of chondrocytes.