Bax channel blocker逆转衰老骨髓间充质干细胞生物学行为的研究

Bax channel blocker reverses the biological characteristics of aged bone marrow stromal cells

目的:探讨Bax channel blocker(BCB)对自然衰老大鼠骨髓间充质干细胞(aged-BMSCs)生物学功能的影响。方法:全骨髓贴壁培养法获得22月龄(老年)和2周龄(年青)大鼠BMSCs,β-gal染色鉴定aged-BMSCs,用BCB对实验组细胞进行干预,以Nanog和Oct-4为靶标,RT-PCR筛选最佳作用浓度。使用RT-PCR和Western Blot分别检测细胞衰老水平及成骨分化的关键基因及蛋白的表达;对细胞药物干预同时进行成骨分化诱导,培养14 d后进行碱性磷酸酶(ALP)染色,21 d后茜素红染色。结果:BCB对aged-BMSCs最佳实验浓度为10μmol/L。BCB能明显降低aged-BMSCs细胞内β-gal含量,在mRNA水平和蛋白水平降低衰老相关基因p53和p21WAF1/cip1表达;明显提升干细胞标记物Nanog、Oct-4。成骨分化诱导后,成骨分化标志基因表达上调,ALP和茜素红染色水平提升。结论:BCB能够逆转aged-BMSCs的衰老,促进其骨向分化。

Objective： To investigate the effects of Bax channel blocker（BCB） on the biological characteristics of aged bone marrow stromal cells （aged-BMSCs）. Methods： BMSCs from 22 month old （aged） and 2 week old rats （young） were obtained by whole bone marrow adherent culture and identified by β-gal staining. The optimal dose of BCB for the detection of Nanog and Oct-4 expression was determined by Real Time PCR. Then the senescence-associated characteristics at cellular and molecular levels and osteogenic ability were tested by RT-PCR and Western Blot. Osteogenic induction and BCB intervention were proceeded. ALP staining and Alizarin Red staining were conducted to observe the osteogenic potential of the ceils. Results： BCB at 10 ttmol/L（ optional dose） declined β-gal expression in aged-BMSCs, decreased the expression of p53 and p21^WAF1/cipl at mRNA and protein levels, and promoted the expression of Nanog and Oct-4, promoted the expression of osteogenesis-associated genes, increased ALP and Alizarin Red staining levels. Conclusion： BCB can reverse the biological characteristics of aged-BMSCs in vitro and promote the osteogenesis potential.