CD146阳性亚群有作为软骨组织工程种子细胞的应用潜力

Application potential of CD146 positive subpopulation as seed cells for cartilage tissue engineering

背景:再生医学的发展、组织工程技术的出现为软骨缺损重建提供了新的解决思路。在组织工程中,间充质干细胞是应用广泛的种子细胞,然而干细胞作为一个异质性的细胞群体其不同亚群发挥着不同的功能。因此,应用间充质干细胞关键功能亚群进行软骨修复具有广泛应用前景。目的:从人脂肪间充质干细胞中分离出CD146阳性亚群细胞,验证其生物学特性及其作为软骨组织工程种子细胞的潜力。方法:人脂肪间充质干细胞由浙江金时代生物技术有限公司提供,通过流式细胞术对人脂肪间充质干细胞表面标志物进行鉴定,应用免疫磁珠分选方法从人脂肪间充质干细胞中分选出CD146阳性表达的细胞亚群。通过基因芯片检测技术及生物信息学分析技术揭示2种细胞的分子特性;体外诱导2种细胞成软骨分化并进行鉴定;冻存复苏前后检测2种细胞的细胞活性及凋亡情况。结果与结论:①人脂肪间充质干细胞表达高水平干细胞相关标志物CD73、CD90,不表达造血干细胞相关标志物CD34、CD45、HLA-DR;②生物信息分析结果表明CD146阳性亚群细胞与脂肪间充质干细胞相比在炎症通路及骨骼肌肉系统疾病有不同功能;③CD146阳性亚群细胞能够成球软骨分化,并且其成软骨分化能力要优于人脂肪间充质干细胞;④CD146阳性亚群细胞复苏后凋亡情况和活性均要优于人脂肪间充质干细胞;⑤结果表明,CD146阳性亚群细胞具有良好的成软骨分化潜力,是一种具有前景的软骨组织工程种子细胞。

BACKGROUND:The development of regenerative medicine and the appearance of tissue engineering technology provide a new solution for cartilage defect reconstruction.In tissue engineering,mesenchymal stem cells are widely used seed cells.However,as a heterogeneous cell group,stem cells play different roles in different subsets.Therefore,the application of key functional subsets of mesenchymal stem cells in cartilage repair has a broad application prospect.OBJECTIVE:To sort CD146 positive subpopulation cells from human adipose-derived mesenchymal stem cells to verify their biological characteristics and potential as seed cells in cartilage tissue engineering.METHODS:Human adipose-derived mesenchymal stem cells were provided by Zhejiang Jinshidai Biotechnology Co.,Ltd.Surface markers of human adipose-derived mesenchymal stem cells were identified by flow cytometry.CD146 positive subpopulation cells were sorted from human adipose-derived mesenchymal stem cells using magnetic-activated cell sorting.Molecular characteristics of two kinds of cells were analyzed by gene chip detection technology and bioinformatics analysis technology.Two kinds of cells were induced to chondrocytes in vitro and histologically examined.Cell viability and apoptosis of two kinds of cells were detected before and after cryopreservation.RESULTS AND CONCLUSION:Human adipose-derived mesenchymal stem cells highly expressed stem cell-associated markers CD73 and CD90,but did not express hematopoietic stem cell-associated markers CD34,CD45,and HLA-DR.Bioinformatics analysis results showed that CD146 positive subpopulation had different functions in inflammatory pathways and musculoskeletal diseases compared with human adipose-derived mesenchymal stem cells.CD146 positive subpopulation could differentiate into cartilage,and its chondrogenic differentiation ability was better than that of human adipose-derived mesenchymal stem cells.CD146 positive subpopulation had better apoptosis and activity than human adipose-derived mesenchymal stem cells after resuscitation.These results suggest that CD146 positive subpopulation has good chondrogenic differentiation potential and is a promising seed cell for cartilage tissue engineering.