白尚星

【摘要】目的观察不同培养基对人脐带间充质干细胞(hUC-MSCs)生物学特性的影响, 优化培养方法。方法采用贴壁法从正常足月胎儿脐带中分离出间充质干细胞, 细胞贴壁后利用DMEM/F12, Mesen PRORSTM Medium这两种培养体系进行体外扩增, 绘制生长曲线, 流式细胞仪检测其表面标志。结果从人脐带中分离出的间充质干细胞为梭形, 呈平行排列生长或漩涡状生长；细胞表达CD29、CD44, 不表达CD34、CD45和HLA-DR。结论加入Mesen PRORSTM Medium培养的细胞增殖速度快, 更适合于脐带间充质干细胞的体外扩增。

【关键词】间充质干细胞；脐带；生物学特性

Influence of different mediums on biological characteristics of human umbilical cord mesenchymal stem cellBAI Shang-xing.Shenyang Sixth People's Hospital, Shenyang 110000, China

【Abstract】Objective To investigate the differences of biological characteristics of human umbilical cord mesenchymal stem cells(hUC-MSCs) in different mediums, and to optimize the purify methods. Methods Mesenchymal stem cells were separated from healthy full-termed delivery fetus using tissue-adherent culture method. These cells were cultured and amplified in DMEM/F12 and Mesen PRORSTM Medium in vitro. The growth curve was drawn.Their surface markers were detected by flow cytometry. Results Mesenchymal stem cells derived from human umbilical cord were spindle-shaped adherent cells.They have strong proliferation ability. The results of growth curve showed that compared with two medium, hUC-MSCs in Mesen PRORSTM Medium grew quickly at every time point. They were strongly positive expression for CD44 and CD29, but negative expression for CD34, CD45 and HLA-DR. Conclusion Cell doubling time in the Mesen PRORSTM Medium was shorter than in DMEM/F12.

【Key words】Mesenchymal stem cell; Umbilical cord; Biological charateristics间充质干细胞(mesenchymal stem cell, MSC)来源于发育早期的中胚层, 是具有高度自我更新能力, 高度增殖能力和多向分化潜能的多能干细胞［1, 2］, 广泛存在于骨髓、脂肪、脐血、脐带等多种组织中［3］。脐带为分娩废弃物, MSC含量丰富, 无需有创穿刺即可获得, 且较骨髓来源的MSC扩增能力更强, 成为近年来MSC研究的重要来源之一［4］。本研究采用两种不同培养基培养hUC-MSCs, 并对其进行生物学特性的观察、细胞免疫表型的鉴定, 以期建立一种稳定、高效、快速、经济的hUC-MSCs富集方法, 为临床细胞移植提供实验支持。

1材料

1. 1脐带选择足月胎儿的脐带, 产妇身体健康, 肝功能正常, 无传染性疾病, 签署知情同意书。脐带4℃无菌保存, 4 h内完成实验操作。

1. 2主要试剂DMEM/F12, Mesen PRORSTM Medium和胰蛋白酶购自GibCO；胎牛血清(FBS)购自HyClone；鼠抗人抗体CD45-FITC、CD34-FITC、CD29-FITC、CD44-FITC、HLA-DR-FITC购自BD。

2方法

2. 1人脐带MSCs 的分离、培养及扩增 脐带剪成3~5 cm小段取出脐带放入培养皿中, 用0.9%氯化钠注射液冲洗, 洗尽血液。用手术剪剔除血管和外膜……