人脐带间充质干细胞分离培养鉴定及诱导分化实验  

Isolation,Culture,Identification and Induced Differentiation of Mesenchymal Stem Cells from Human Umbilical Cord

作　　者：谢娜[1] 娄鸣[1] 饶国洲[2] 朱勇[1] 唐成芳[1] 王琳[1] 

机构地区：[1]西安医学院口腔医学系,西安710021 [2]西安交通大学口腔医院,西安710004

出　　处：《现代检验医学杂志》2015年第3期67-70,共4页Journal of Modern Laboratory Medicine

基　　金：陕西省卫生厅卫生科研项目（2014D19）;陕西省教育厅2012年科学研究项目（12JK0757）;陕西省教育厅专项科研计划项目（2013JK0784）.

摘　　要：目的：建立人脐带间充质干细胞（MSCs）分离培养方法，并进行生物学鉴定及定向诱导分化研究。方法采用酶消化 Wharton 胶法体外分离培养人脐带干细胞，通过流式细胞仪分析其表面标记分子，并向成骨、成脂方向诱导分化，钙钴法染色检测 ALP，茜素红染色检测矿化结节，油红“O”染色检测脂肪滴，进行干细胞的成骨成脂活性鉴定。结果分离培养的脐带间充质干细胞 CD73，CD90和 CD105均表达阳性，阳性率为92.45％，95.45％和96.45％，CD34表达阴性，阳性率仅为1.07％。成骨诱导3周后 ALP 染色胞质内可见黑色颗粒，4周后可见大量矿化结节。成脂诱导2周后细胞内出现大量脂肪小滴，多数细胞形态变圆，油红“O”染色阳性。结论脐带来源的间充质干细胞具有成骨、成脂多向分化潜能，为临床干细胞治疗研究的种子细胞来源奠定了基础。Objective To establish the method of isolation,culture and identify biological characterization of mesenchymal stem cells from human umbilical cord （hUCMSCs）;and study their multiple differentiation potency.Methods Stem cells from human umbilical cord were cultured by enzyme Wharton jelly method in vitro.The surface markers were identified by flow cytometry.Multi-differentiation capacity was identified by osteogenic and adipogenic differentiation.ALP was detected with Calcium cobalt staining.The mineralized ability in vitro was measured with Alizarin red staining.Theadipocyte differen-tiation ability was measured with oil red-O staining.Results Flow cytometry analysis revealed that CD73 （92.45%）,CD90 （95.45%）and CD105 （96.45%）were highly expressed on these cells’surface,while CD34 （1.07%）were negative ex-pressed.Cells were cultured with induced-osteogenic medium after 3 weeks,ALP staining in the cytoplasm of black parti-cles,and a large amount of mineralized nodules within cells was observed after 4 weeks.Cells were cultured with induced-adi-pogenic medium after 2 weeks,the majority of these cells were round,oil red O staining of lipid droplets generated within cells was observed.Conclusion Mesenchymal stem cells from human umbilical cord have the potential of multi-directional differentiation.These cells could be induced to differentiate into adipocytes and osteoblasts,which laid the foundation for clinical stem cell therapy research source of seed cells.

关 键 词：脐带 WHARTON 胶 干细胞 诱导分化 

分 类 号：R457.7[医药卫生—治疗学]