人骨髓间质干细胞向脂肪细胞诱导分化过程中端粒酶活性的变化

目的观察体外培养人骨髓间质干细胞(mesenchymal stem cells,MSCs)成脂分化过程中端粒酶活性的变化规律,并探讨其可能的机制。方法通过贴壁培养法从人骨髓中分离MSCs,利用造血干细胞相关表面抗体对MSCs进行表型鉴定,利用相应诱导分化剂,诱导MSCs向成骨细胞、脂肪细胞和神经元样细胞分化,分析其多向分化潜能。以端粒重复序列扩增法(TRAP)检测MSCs及MSCs诱导成脂分化过程中端粒酶活性;以Western印迹法检测MSCs及MSCs诱导成脂分化过程中端粒酶反转录酶(hTERT)及端粒相关蛋白端粒重复序列结合因子1(TRF1)、端锚聚合酶1(Tankyrase 1)]的表达水平。结果 TRAP法检测传代培养的MSCs端粒酶呈阴性表达,而MSCs诱导成脂分化过程中,端粒酶活性明显升高,以第7天最高(P<0.05),其后呈下降趋势;Western印迹法检测显示,hTERT在MSCs中有微弱表达,在MSCs诱导成脂分化过程中,hTERT和Tankyrase 1表达水平升高,以第7天最高,此后逐渐下降,而TRF1表达水平保持不变。结论体外培养的MSCs端粒酶呈微弱表达,MSCs在成脂诱导分化过程中,端粒酶活性先升高,其后逐渐下降,而Tankyrase 1可能在其中发挥重要作用。

Changes of telomerase activity during differentiation of human mesenchymal stem cells into adipocytes

ABSTRACT] Objective: To study the telomerase activity changes and mechanism during the differentiation of human mesenchymal stem cells (MSCs) into adipocytes. Methods: Human MSCs were isolated from healthy human bone marrow by plastic adhesion and characterized by immune phenotype. TRAP was performed to measure the telomerase activity in MSCs and during the differentiation of MSCs into adipocytes. Western blotting was applied to detect the protein expression levels of hTERT, TRF1, and Tankyrase1 in MSCs and during adipogenic differentiation of MSCs. Results: By the Telomeric Repeat Amplification Protocol (TRAP) assay, telomerase was found negative in MSCs, up-regulated when MSCs was adipogenic differentiated, and decreased 7 days later. MSCs expressed very low level of hTERT detected by Western blotting. The expression of hTERT and Tankyrase1 protein paralleled with the changes of telomerase activity during the differentiation of MSCs into adipocytes, but the expression of TRF1 protein kept unchanged. Conclusion: Telomerase activity was very low in MSCs, up-regulated and then down-regulated during MSCs adipogenic differentiation, and Tankyrase1 may play a role in it.