虎杖苷调节Sirt1-FoxO1信号通路对高糖诱导的心肌细胞焦亡的影响

户小均; 黄丹; 苏维; 张和平; 张泉

doi:10.12125/j.chj.202304122

虎杖苷调节Sirt1-FoxO1信号通路对高糖诱导的心肌细胞焦亡的影响

Impact of polydatin on myocardial cell pyroptosis induced by high glucose through regulation of Sirt1-FoxO1 signal pathway

摘要

摘要:
目的基于沉默信息调节因子1（silence information regulator1，Sirt1）-叉头状转录因子O1（forkhead transcription factor O1，FoxO1）信号通路研究虎杖苷（polydatin，PD）抑制高糖诱导的心肌细胞焦亡的作用机制。
方法以心肌细胞H9c2为研究对象，使用不同浓度PD处理筛选PD浓度。将细胞分为模型组、PD低、中、高剂量组（PD-L、M、H，20、40、80 μmol/L）、空白对照（control）组、Sirt1抑制剂PD-H+sirtinol （10 μmol/L）组。细胞计数试剂盒（cell counting Kit-8，CCK8）法检测细胞活性；酶联免疫吸附试验（enzyme-linked immunosorbent assay，Elisa）法检测细胞内乳酸脱氢酶（lactate dehydrogenase，LDH）释放量、白细胞介素1β（Interleukin-1β，IL-1β）水平；二氯荧光素双醋酸盐（dichlorodihydrofluorescein diacetate，DCFH-DA）荧光探针检测细胞活性氧（reactive oxygen species，ROS）水平；流式细胞术检测细胞焦亡情况；实时荧光定量PCR检测细胞中Nod样受体蛋白3（Nod-like receptor protein 3，NLRP3） mRNA、消皮素D（gasdermin D，GSDMD） mRNA表达水平；Western blot检测蛋白表达。
结果与control组比较，模型组细胞活性下降（P<0.01），经过PD不同浓度处理后，细胞活性逐渐上升（P<0.01）；与control组比较，模型组LDH释放量、IL-1β水平、ROS水平、细胞焦亡率、NLRP3 mRNA、GSDMD mRNA表达、焦亡蛋白NLRP3、裂解的半胱氨酸天冬氨酸酶（cleaved-caspase-1，c-caspase-1）、GSDMD、GSDMD-N表达均上升，Sirt1-Foxo1信号通路蛋白表达下降（P<0.01）；与模型组比较，PD各组及PD-H+sirtinol组LDH释放量、IL-1β水平、ROS水平、细胞焦亡率、NLRP3 mRNA、GSDMD mRNA表达、焦亡蛋白NLRP3、c-caspase-1、GSDMD、GSDMD-N表达均下降，Sirt1-FoxO1信号通路蛋白表达上升（P<0.05）。sirtinol可以逆转PD对于高糖诱导的心肌细胞焦亡的保护作用。
结论 PD可以通过上调Sirt1-FoxO1信号通路蛋白表达，改善细胞炎症损伤，抑制高糖诱导的心肌细胞焦亡。

Abstract:
AIM To study the mechanism of polydatin (PD) in inhibiting myocardial cell pyroptosis induced by high glucose based on silence information regulator1 (Sirt1)-forkhead transcription factor O1 (FoxO1) signal pathway.
METHODS Cardiac cell H9c2 was taken as the research object and the PD concentration was screened by different concentrations of PD treatment. Cells were grouped into model group, PD low, medium and high dose groups (PD-L, M, H, 20, 40, 80 μmol/L), blank control group (control) and Sirt1 inhibitor group PD-H+sirtinol (10 μmol/L). Cell Counting Kit-8 (CCK8) method was applied to detect the cell activity and enzyme-linked immunosorbent assay (Elisa) method was used to measure the release of lactate dehydrogenase (LDH) and the level of Interleukin-1β (IL-1β). The level of reactive oxygen species (ROS) was detected by Dichlorodihydrofluorescein diacetate (DCFH-DA) fluorescence probe, the charred cells were detected by flow cytometry and the expression levels of Nod-like receptor protein 3 (NLRP3) mRNA and gasdermin D (GSDMD) mRNA in cells were detected by real-time fluorescence quantitative PCR. Western blot was applied to detect the protein expression.
RESULTS Compared with those in control group, the cell activity in model group decreased (P<0.01) and after treatment with different concentrations of PD, the cell activity increased gradually (P<0.01). Compared with those in control group, LDH release, IL-1β level, ROS level, pyroptosis rate, NLRP3 mRNA, GSDMD mRNA expressions, and expressions of pyroptosis protein NLRP3, c-caspase-1, GSDMD and GSDMD-N in model group increased, and the expression of Sirt1-FoxO1 signal pathway protein decreased (P<0.01). Compared with those in model group, LDH release, IL-1β level, ROS level, scorching rate, NLRP3 mRNA, GSDMD mRNA expressions, and expressions of scorching protein NLRP3, c-caspase-1, GSDMD and GSDMD-N in PD group and sirtinol group decreased, and the expression of Sirt1-FoxO1 signal pathway protein increased (P<0.05). Sirtinol reversed the protective effect of PD on myocardial cell pyroptosis induced by high glucose.
CONCLUSION PD can improve the inflammatory injury of cells and inhibit the high glucose-induced myocardial cell pyroptosis by up-regulating the expression of Sirt1-FoxO1 signal pathway protein.

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