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目的观察脐带间充质干细胞(umbilical cord mesenchymal stem cells,UC-MSCs)体外长期培养的生物学特性及遗传稳定性。方法从剖宫产足月健康新生儿脐带中分离培养UC-MSCs,并连续传代,显微镜下观察细胞形态。分别检测P3和P12代UC-MSCs的增殖能力、免疫表型、向成脂肪细胞和成骨细胞的分化诱导率、衰老情况、成纤维细胞集落形成单位(CFU-F)及染色体核型。结果 P3代UC-MSCs为形态相对均一的梭形贴壁细胞,呈平行排列生长或旋涡状生长;P12代UC-MSCs宽大扁平,贴壁性减退,漂浮细胞增多,胞浆内出现黑色颗粒和空泡;P3和P12代UCMSCs均表达CD73、CD90、CD105,不表达CD34、CD45、HLA-DR;P3较P12代UC-MSCs的细胞增殖能力及向成脂肪细胞分化诱导率均明显升高(P均<0.05);向成骨细胞分化诱导率及细胞衰老率均明显降低(P均<0.05);成纤维细胞集落数及染色体中期分裂相明显增多(P<0.05)。结论长期体外培养的UC-MSCs生物学特性发生改变,遗传学特性稳定,但染色体中期分裂相减少。
Objective To observe the biological characteristics and genetic stability of long-term culture of umbilical cord mesenchymal stem cells (UC-MSCs) in vitro. Methods UC-MSCs were isolated and cultured from umbilical cords of full-term healthy neonates with cesarean section. The cells were passaged continuously and observed under a microscope. Proliferation, immunophenotype, differentiation induction to adipocytes and osteoblasts, aging, fibroblast colony forming units (CFU-F) and chromosome karyotype of P3-P12 UC-MSCs were detected respectively. Results The P3-generation UC-MSCs were spindle-shaped adherent cells with relatively uniform morphology and were arranged in parallel or spirally. The UC-MSCs on P3 generation were flat and adherent, the number of floating cells increased, and black particles appeared in the cytoplasm The expression of CD73, CD90, CD105, CD34, CD45 and HLA-DR in UCMSCs on P3 and P12 generation was significantly higher than that on P12 on UC-MSCs P <0.05). The differentiation into osteoblasts and the cell senescence rate were significantly decreased (P <0.05). The number of fibroblasts and metaphase chromosomes were significantly increased (P <0.05). Conclusion The biological characteristics of long-term cultured UC-MSCs changed, but their genetic characteristics were stable, but the metaphase of chromosomes was reduced.