兔脂肪垫滑膜间充质干细胞分离、纯化及诱导定向分化研究

Separation,Purification and Induction of Directional Differentiation of Rabbit Synovial Mesenchymal Stem Cells

目的：探讨兔脂肪垫滑膜间充质干细胞（SMSCs）的分离、培养、生物学特性及其定向分化的能力。方法采用兔脂肪垫组织块培养法分离原代 SMSCs，有限稀释法纯化和扩增 SMSCs，观察 SMSCs 形态学特点；CCK8法测定第2代及第4代 SMSCs 增殖能力；Alcian Blue、碱性磷酸酶，以及油红“O”染色法观察第4代 SMSCs 在定向诱导液中向成软骨、成骨、成脂肪细胞的分化能力。结果脂肪垫滑膜组织经组织块培养2～3 d 见 SMSCs 生长，12～14 d 可获大量 SMSCs，纯化传代后细胞形态均一；第2、4代 SMSCs 细胞增殖能力比较差异无统计学意义（P ＞0．05）；SMSCs 经诱导可向软骨细胞、成骨细胞、脂肪细胞定向分化。结论关节内脂肪垫滑膜组织采用组织块培养法能有效地分离出 SMSCs，其间充质干细胞具有多向诱导分化潜能。

Objective To observe the separation,culture,biological characteristics and ability to directional differentiation of rabbit synovial mesenchymal stem cells （SMSCs）.Methods Rabbit fat pad tissues were cultured to obtain the primary SMSCs,which were purified and amplified u-sing the limiting dilution method.The morphologic characteristics of SMSCs were observed.The proliferation of passage 2 and 4 cells was determined by CCK8 analysis.The differentiation of pas-sage 4 cells towards chondrocytes,osteoblasts and lipoblasts was detected using alcian blue,alka-line phosphatase and oil red O staining,respectively.Results The SMSCs were found after tissue culture for 2-3 days,and a large number of SMSCs were obtained after tissue culture for 12-14 days.After purification,cells showed uniform morphology.There were no significant difference in proliferation ability between passage 2 cells and passage 4 cells （P 〉0.05）.The SMSCs could be induced to differentiate into chondrocytes,osteoblasts and lipoblasts.Conclusion The SMSCs can be effectively separated from intra-articular fat pad synovial tissue by tissue culture.The separa-ted SMSCs have multi-directional differentiation potential.