丹酚酸B对骨髓间充质干细胞存活的影响及其作用机制研究

Effect and mechanism of Salvianolic acid B on survival of bone marrow mesenchymal stem cell

目的探讨丹酚酸B(SalB)对骨髓间充质干细胞(MSC)存活的影响及其作用机制。方法体外培养MSC,建立体外缺血缺氧(OGD)模型。实验分MSC组、MSC+OGD组和MSC+OGD+不同浓度丹酚酸B组,MSC+OGD+丹酚酸B组在进行OGD处理前30 min,加入相应浓度的SalB进行预处理。通过Hochest33342染色和Annexin-V/PI双重染色流式细胞仪检测细胞凋亡和存活,经Rhodamine 123染色观察细胞线粒体膜电位的变化,通过Western blot检测Bcl-2和Bax的表达。结果 Hochest33342染色和Annexin-V/PI流式细胞仪检测显示,MSC经OGD处理后,出现明显的凋亡现象(P<0.01)。OGD处理前加入SalB与单纯OGD处理比较,MSC的凋亡明显减少,存活细胞数目显著增多(P<0.05,P<0.01)。OGD处理可以降低Rhodamine 123的表达,减少Bcl-2水平、增加Bax水平(P<0.01);与OGD组比较,10μmol/L丹酚酸B预处理可增加Rhodamine 123的表达,并可增加Bcl-2水平、减少Bax水平(P<0.05)。结论 SalB预处理可阻止线粒体凋亡途径,从而减少MSC凋亡,提高干细胞的存活率。

Objective To discuss the effect and mechanism of Salvianolic acid B on survival of bone marrow mesenchymal stem cells （MSC）. Methods After culture of bone marrow mesenehymal stem cells MSC in vitro, oxygen-glucose deprivation （OGD） model was established in vitro. Experiments included MSC group, MSC ＋ OGD group and MSC ＋ OGD ＋ different concentrations of Sal B groups, the appropriate concentrations of SalB were added in the group of MSC ＋ OGD ＋ Sal B 30 min before OGD treatment. Apoptosis and survival were determined through Hochest33342 staining and Annexin-V/PI double staining flow cytometry. Changes of mitochondrial membrane potential were evaluated by Rhodamine 123 staining. Expression of Bcl-2 and Bax was detected by Western blot. Results MSC presented significant apoptosis after treatment of OGF according to Hochest33342 staining and Annexin-V/PI double staining flow cytometry （ P 〈 O. 01 ）. Compared with the OGD treatment only, MSC apoptosis was significantly reduced, and the number of survival cells was significantly increased in the group of adding SalB before OGD treatment （ P 〈 O. 05, P 〈 0.01 ）. Treatment of OGD could reduce the Rhodamine 123 expression, decrease the Bcl-2 expression and increase the Bax expression （ P 〈 O. O1 ）. Compared with the OGD treatment only, 10 μmol/L SalB pretreatment could elevate the Rhodamine 123 expression, increase the Bcl-2 expression and decrease the Bax expression （ P 〈 O. 05 ）. Conclusion SalB pretreatment could prevent mitochondrial apoptosis, thereby reduce MSC apoptosis, and increase the survival rate of stem cells.