人骨髓间充质干细胞端粒酶活性的表达

Telomerase activity of human bone marrow mesenchymal stem cells

目的 :探讨人骨髓间充质干细胞 (human mesenchymal stem cells,h MSCs)端粒酶活性的表达以及深低温冻存和向脂肪细胞诱导分化对基因表达的影响。方法 :联合应用密度梯度离心法和贴壁培养法分离人 h MSCs,传代培养。深低温冻存 h MSCs或诱导其向脂肪细胞分化 ,以 TRAP(Telomerase repeatamplification protocol assay)法分别检测新鲜的不同传代次数的 h MSCs、冻存复苏后培养的 h MSCs和诱导分化为脂肪细胞的 h MSCs的端粒酶活性。结果 :用 TRAP法检测 h MSCs(n=19)端粒酶活性 (RTA)是 (1.4 6± 0 .6 7) % ,分化成脂肪的 h MSCs(n=3)的RTA为 (11.80± 2 .5 2 ) % (P<0 .0 0 1) ;不同传代次数 MSCs端粒酶活性的比较中 ,早期 h MSCs(n=10 )的 RTA为(1.4 6± 0 .83) % ,晚期 h MSCs(n=9)的 RTA为 (1.4 6± 0 .4 7) % (P=0 .99) ;在低温冻存对 h MSCs端粒酶活性的影响中 ,新鲜的 h MSCs(n=13)的 RTA为 (1.4 1± 0 .4 4) % ,低温冻存的 h MSCs(n=6 )的 RTA为 (1.5 7± 1.0 7) %(P=0 .6 4)。结论 :h MSCs的端粒酶呈阴性 ,传代培养和低温冻存不影响基因表达水平。向脂肪细胞分化后 h MSCs端粒酶活性表达水平增高 ,其确切机制尚需进一步研究。

Objective: To investigate the telomerase activity in mesenchymal stem cells (hMSCs) from human bone marrow after their in vitro committed differentiation into adipocytes and cryopreservation. Methods: hMSCs were isolated from human bone marrow. The isolated hMSCs were induced to differentiate into adipocytes in vitro or cryopreserved.TRAP assay (telomerase repeat amplification protocol assay) was employed to detect telomerase activity in those hMSCs. Results: Telomerase activity (RTA) in hMSCs ( n =19) was (1 46±0 67)%,while that in hMSCs derived adipocytes ( n =3) was (11 80±2 52)% ( P <0 001).RTA of hMSCs passage 1 3 ( n =10) was (1 46±0 83)%,and that of hMSCs passage 4 7( n =9) was (1 46±0 47)% ( P =0 99).Cryopreservation did not affect the telomerase activity in hMSCs,RTA of fresh hMSCs( n =13) was (1 41±0 44)%,RTA of frozen hMSCs( n =6) was (1 57±1 07)% ( P =0 64). Conclusion: hMSCs are telomerase negative,but telomerase activity in hMSCs derived adipocytes is upregulated. [