2-APB通过PKCα/HIF-1α信号通路抑制H2O2诱导的软骨细胞凋亡
2-APB inhibits H2O2-induced chondrocyte apoptosis through PKCα/HIF-1α signaling pathway
欧阳紫微 1董雷 2王琰 1程远志 1朱仁弟 1周仁鹏 1赵英杰 1胡伟1
作者信息
- 1. 安徽医科大学药学院,合肥 230032;安徽医科大学第二附属医院药物临床试验研究中心,合肥 230601
- 2. 安徽医科大学第二附属医院药物临床试验研究中心,合肥 230601
- 折叠
摘要
目的 探讨2-氨基乙氧基苯硼酸(2-APB)对H2O2诱导的软骨细胞凋亡的作用及其机制.方法 实验分为Con-trol 组、H2O2 组、2-APB 组和 H2O2+2-APB 组.CCK-8 法检测各组细胞活力;显微镜下观察2-APB对H2O2诱导的软骨细胞形态变化的影响;TUNEL法和流式细胞术检测软骨细胞凋亡情况;流式细胞术检测脂质活性氧(ROS)水平;West-ern blot法检测2-APB对H2O2诱导的各组细胞中Cleaved-PARP、p-PKCα和HIF-1α蛋白的表达情况;免疫荧光法检测PKCα抑制剂BIM-Ⅰ对H2O2诱导的各组细胞中HIF-1α的荧光表达情况.结果 2-APB对H2O2诱导的软骨细胞凋亡具有抑制作用,且当2-APB浓度为100 μmol/L时抑制效果最为显著(F=235.80,P＜0.01);2-APB能够抑制H2O2所致软骨细胞凋亡阳性率(F=114.80,P＜0.01)以及ROS的水平(F=52.99,P＜0.01),并且抑制 Cleaved-PARP(F=10.10,P＜0.05)、p-PKCα(F=24.56,P＜0.05)和 HIF-1α(F=6.85,P＜0.05)蛋白的表达;PKCα抑制剂BIM-Ⅰ能够抑制H2O2所致HIF-1α荧光强度的增加.结论 2-APB可以通过抑制PKCα/HIF-1α通路减少H2O2诱导的软骨细胞凋亡,进而保护软骨细胞.
Abstract
Objective To explore the effect of 2-aminoethoxy-diphenyl borate(2-APB)on H2O2-induced chondro-cyte apoptosis and its mechanism.Methods The experiment was divided into control group,H2O2 group,2-APB group and H2O2+2-APB group.CCK-8 method was used to detect the cell viability of each group;The effect of 2-APB on the morphological changes of chondrocytes induced by H2O2 was observed under microscopy;TUNEL meth-od and flow cytometry were used to detect chondrocyte apoptosis;Flow cytometry was used to detect Lipid reactive oxygen species(ROS);Western blot was used to detect the protein expressions of Cleaved-PARP,p-PKCα and HIF-1α in H2O2-induced cells by 2-APB;Immunofluorescence was used to detect the fluorescent expression of HIF-1α in cells induced by H2O2 by PKCα inhibitor BIM-1.Results 2-APB inhibited H2O2-induced apoptosis in chon-drocytes,and the inhibitory effect was the most significant when the concentration of 2-APB was 100 pmol/L(F=235.80,P＜0.01);22-APB could inhibit the positive rate of H2O2-induced apoptosis of chondrocytes(F=114.80,P＜0.01)and the level of ROS(F=52.99,P＜0.01).and inhibited the expression of Cleaved-PARP(F=10.10,P＜0.05),p-PKCα(F=24.56,P＜0.05)and HIF-1α proteins(F=6.85,P＜0.05).The PKCα in-hibitor BIM-Ⅰ could inhibit the increase in HIF-1α fluorescence intensity caused by H2O2.Conclusion 2-APB can inhibit chondrocytes apoptosis induced by H2O2 through the PKCα/HIF-1α pathway and thus protect chondro-cytes.
关键词
2-氨基乙氧基苯硼酸/软骨细胞/细胞凋亡/H2O2/PKCα/HIF-1α/通路Key words
2-aminoethoxy-diphenyl borate/chondrocytes/apoptosis/H2O2/PKCα/HIF-1α signaling pathway引用本文复制引用
基金项目
国家自然科学基金(82071591)
国家自然科学基金(82371575)
国家自然科学基金(82204405)
安徽省自然科学基金(2108085QH383)
出版年
2024
国家科技期刊平台
NSTL
万方数据