冻存复苏人脐带间充质干细胞体外扩增和向脂肪细胞分化的研究

目的：分离培养人脐带间充质干细胞（human Umbilical Cord Mesenchymal Stem Cells hUCMSCs），观察其冻存复苏后向脂肪细胞定向分化的能力。方法：将剔除动静脉的新鲜人脐带组织切成小块培养，得到贴壁细胞，观察细胞生长及检测其表面抗原。将第1代的hUCMSCs采用梯度冷冻技术冻存5个月，复苏后培养至第12代时，加入成脂诱导剂培养。当诱导至21天时行油红O染色，7天及14天时用RT—PCR技术分别检测成脂转化基因过氧化物酶增殖剂受体（PPAR γ-2）和脂蛋白酯酶（LPL）。结果：组织块培养法收获的单个核细胞培养传代后，能获得均一贴壁的间充质干细胞，hUCMSCs冻存复苏后，活细胞约为86％，流式细胞仪分析这些细胞表达CDI3、CD44和CD71等MSCs标志物，不表达CD14、CD34和HLA—DR表面抗原。复苏细胞在成脂诱导剂的作用下，细胞中有脂滴产生，通过油红0染色显示细胞核周围有脂滴聚集，通过RT—PCR检测有LPL及PPARγ2 mRNA的表达。结论：采用组织块贴壁培养法分离获得的人脐带间充质干细胞可冷冻保存。复苏后培养至第12代仍具有向脂肪细胞分化的潜能，可作为脂肪组织工程种子细胞来源。

Differentiation of cryopreserved human Umbilical Cord Mesenchymal Stem Cells into adipogenic cells

Objective To isolate and culture the human Umbilical Cord Mesenchymal Stem Cells(hUCMSCs) and study the potential to differentiate into adipogenic cells after they were frozen and thawed.Methods After stripping off arteries and veins,the remaining parts of umbilical cord were cut into small sections and plated in 100mm culture capsule.Adherent cells coming out of fragments and the cell growth was observed and its surface antigen was detected.The one-passage hUCMSCs were preserved in liquid nitrogen by reduc...