(±) Bay K8644提高衰老大鼠骨髓间充质干细胞成骨分化能力的研究

Study on the promotion effects of ( ± ) Bay K8644 on osteogenic differentiation ability in aging bone marrow stromal cells of rats

目的探讨(±)Bay K8644对衰老的大鼠(24 m)来源的骨髓间充质干细胞(Aging bone marrow stromal cells,Aging-BMSCs)的骨向分化能力的影响。方法通过全骨髓贴壁培养法获得大鼠BMSCs,使用β-半乳糖苷酶染色方法鉴定细胞的衰老状态,随后应用(±)Bay K8644对细胞进行干预。以干性标志物Nanog为靶点,通过实时定量PCR筛选出(±)Bay K8644作用的最佳浓度,随后对细胞进行成骨分化诱导,并检测细胞在药物干预下的骨向分化能力的改变。在诱导第14天后对成骨分化标志酶碱性磷酸酶进行染色及活性测定、第21天后进行茜素红染色。结果(±)Bay K8644明显的降低了Aging-BMSCs细胞内β-半乳糖苷酶的含量(P<0.05),同时干细胞的干性标记物Nanog的表达明显提升(P<0.05)。对细胞进行成骨分化诱导后,成骨分化标志基因表达上调,ALP、茜素红染色的结果提示实验组骨向分化水平明显较对照组提升(P<0.05)。结论 (±)Bay K8644能够明显改善衰老骨髓间充质干细胞的衰老状态,并使其骨向分化的能力得到提升。

Objective To investigate the effects of（ ±） Bay K8644 on osteogenic differentiation ability in aging bone marrow stromal cells（ Aging-BMSCs） of rats. Methods The BMSCs of rats were obtained by whole bone marrow adherent culture method,and the aging condition of cells was identified by β-galactosidase staining. The cells were intervented by（ ±）Bay K8644. PT-Real time PCR was used to screen the optimal concentration by taking Nanog as target. Then the osteogenic ability was examined. RT-PCR and Western Blot were used to detect the changes of the key targets of osteogenesis. ALP staining and ALP activity testing were performed on 14 days after osteogenic induction,and Alizarin red staining was performed after 21 days. Results The concentration of β-galactosidase in aging BMSCs was decreased by the intervention of（ ±） Bay K8644. At the same time,the expression of Nanog was obviously increased. After osteogenic induction,the expression of osteogenesis-associated gene was up-regulated. The ALP and Alizarin red staining showed that the osteogenic differentiation levels in trial group were significantly higher than those in control group. Conclusion The（ ±） Bay K8644 can obviously improve the aging condition of Aging-BMSCs of rats,moreover,which can promote the ability of osteogenesis.