小檗碱通过ROS/JNK信号通路对AngⅡ诱导的人脐静脉血管内皮细胞凋亡的抑制作用及其机制

doi:10.13481/j.1671-587x.20190422

摘要/Abstract

摘要： 目的：探讨小檗碱对血管紧张素Ⅱ（AngⅡ）诱导的人脐静脉血管内皮细胞（HUVECs）凋亡的影响及其机制，阐明小檗碱抗动脉粥样硬化（AS）的机制。方法：体外培养的HUVECs分为对照组（未处理），AngⅡ组（加入1 μmol·L^-1 AngⅡ处理24 h），低、中和高剂量小檗碱组（1 μmoL·L^-1 AngⅡ预处理3 h后，分别加入25、50和100 μmol·L^-1小檗碱处理24 h）。采用MTT法和流式细胞术分别检测各组细胞的存活率和凋亡率，活性氧（ROS）检测试剂盒检测各组细胞中ROS水平，Western blotting法检测各组细胞中磷酸化c-Jun氨基末端激酶（p-JNK）、B淋巴细胞瘤-2基因（Bcl-2）、Bcl-2相关X蛋白（Bax）和活化的含半胱氨酸的天冬氨酸蛋白水解酶3（cleaved caspase-3）蛋白表达水平。结果：与对照组比较，AngⅡ组细胞存活率和Bcl-2蛋白表达水平降低（P<0.05），细胞凋亡率、ROS水平及p-JNK、Bax和cleaved caspase-3蛋白表达水平均升高（P<0.05）；与AngⅡ组比较，各剂量小檗碱组细胞存活率和Bax蛋白表达水平明显升高（P<0.05），细胞凋亡率、ROS水平及p-JNK、Bax和cleaved caspase-3蛋白表达水平均降低（P<0.05），且呈剂量依赖性。结论：小檗碱可抑制AngⅡ诱导的HUVECs凋亡，其作用机制可能与抑制ROS/JNK信号通路有关。

关键词: 血管紧张素Ⅱ, 人脐静脉血管内皮细胞, 小檗碱, 细胞凋亡

Abstract: Objective:To investigate the effect of berberine on the angiotensin Ⅱ (AngⅡ)-induced apoptosis of human umbilical vein endothelial cells (HUVECs) and its mechanism,and to clarify the anti-AS mechanism of berberine. Methods:The HUVECs cultured in vitro were divided into control group (untreated), AngⅡ group (treated with 1 μmoL·L^-1 Ang Ⅱ for 24 h) and low,medium,and high doses of berberine groups (after pretreated with 1 μmoL·L^-1 Ang Ⅱ for 3 h, treated with 25, 50, and 100 μmoL·L^-1 berberine for 24 h, respectively).The survival rates and the apoptotic rates of cells in each group were detected by MTT method and flow cytometry,the ROS level in each group was detected by ROS kit, and the expression levels of p-JNK, Bcl-2, Bax, and cleaved caspase-3 proteins in each group were detected by Western blotting method. Results:Compared with control group, the survival rate of cells and the Bcl-2 protein expression level in AngⅡgroup were decreased(P<0.05), while the apoptotic rate of cells, the ROS level and the expression levels of p-JNK, Bax, and cleaved caspase-3 proteins were increased(P<0.05); compared with AngⅡgroup, the survival rates of cells and the expression levels of Bax protein in low,medium, and high doses of berberine groups were significantly increased(P<0.05), while the apoptotic rates of cells, the ROS levels and the expression levels of p-JNK, Bax, and cleaved caspase-3 proteins were decreased(P<0.05) in a concentration-dependent manner. Conclusion:Berberine can inhibit the apoptosis of HUVECs induced by AngⅡ, which may be related to the inhibition of ROS/JNK signaling pathway activation.

Key words: angiotensin Ⅱ, human umbilical vein endothelial cells, berberine, apoptosis

中图分类号:

R285.5

岳胜, 乔国华, 岳磊, 朱平. 小檗碱通过ROS/JNK信号通路对AngⅡ诱导的人脐静脉血管内皮细胞凋亡的抑制作用及其机制[J]. 吉林大学学报(医学版), 2019, 45(04): 872-876.

YUE Sheng, QIAO Guohua, YUE Lei, ZHU Ping. Inhibitory effect of berberine on Ang Ⅱ-induced apoptosis of human umbilical vein endothelial cells via ROS/JNK signaling pathway and its mechanism[J]. Journal of Jilin University(Medicine Edition), 2019, 45(04): 872-876.

参考文献

[1] 闫杰,陈静,黄宇玲,等.Let-7d靶向LOX-1逆转ox-LDL诱导血管内皮细胞凋亡观察[J]. 郑州大学学报:医学版,2018,53(5):564-569.
[2] CHEN L, YANG W, GUO Y, et al. Exosomal lncRNA GAS5 regulates the apoptosis of macrophages and vascular endothelial cells in atherosclerosis[J]. PLoS One, 2017, 12(9):e0185406.
[3] 位庚, 李红蓉, 常丽萍, 等. 血管紧张素Ⅱ诱导血管内皮细胞损伤及中药保护作用[J]. 中国老年学杂志, 2017, 37(13):3353-3355.
[4] ZHANG L, WANG J, LIANG J, et al. Propofol prevents human umbilical vein endothelial cell injury from Ang Ⅱ-induced apoptosis by activating the ACE2-(1-7)-Mas axis and eNOS phosphorylation[J]. PLoS One, 2018, 13(7):e0199373.
[5] LI X M, WANG Q Z, GUO L. Beneficial effect of berberine on atherosclerosis based on attenuating vascular inflammation and calcification[J]. Chin J Pharm Toxicol, 2017, 31(10):993-994.
[6] 贾燕珺. 小檗碱的抗动脉粥样硬化作用及其机制[J]. 中国动脉硬化杂志, 2013, 21(4):364-368.
[7] REULE C A, GOYVAERTS B, SCHOEN C. Effects of an L-arginine-based multi ingredient product on endothelial function in subjects with mild to moderate hypertension and hyperhomocysteinemia-a randomized, double-blind, placebo-controlled, cross-over trial[J]. BMC Complement Altern Med, 2017, 17(1):92-100.
[8] 史海霞,杨嘉君,聂绪强, 等.赤芍醇提物对血管内皮细胞一氧化氮合成和分泌的影响[J].郑州大学学报:医学版,2017,52(2):159-163.
[9] 刘永刚, 李芳君. 羟基红花黄色素A对AngⅡ诱导的人脐静脉内皮细胞凋亡的抑制作用[J]. 中药材, 2013, 36(7):1128-1131.
[10] SHAN F L, SHAO Z W, JIANG S H, et al. Erlotinib induces the human non-small-cell lung cancer cells apoptosis via activating ROS-dependent JNK pathways[J]. Cancer Med, 2016, 5(11):3166-3175.
[11] LIN C L, LEE C H, CHEN C M, et al. Protodioscin induces apoptosis through ROS-mediated endoplasmic reticulum stress via the JNK/p38 activation pathways in human cervical cancer cells[J]. Cell Physiol Biochem, 2018, 46(1):322-334.
[12] 李斌, 曾勉, 何婉媚,等. Ghrelin通过JNK信号通路抑制LPS诱导的肺泡巨噬细胞凋亡[J]. 中山大学学报:医学科学版, 2015, 36(2):181-188.
[13] XIANG Y K, YE W, HUANG C H, et al. Brusatol inhibits growth and induces apoptosis in pancreatic cancer cells via JNK/p38 MAPK/NF-κB/Stat3/Bcl-2 signaling pathway[J]. Biochem Biophys Res Commun, 2017, 487(4):820-826.
[14] 张婧. 蓝莓花色苷对血管紧张素Ⅱ致人脐静脉血管内皮细胞凋亡的保护作用[D]. 长春:吉林大学, 2013.
[15] 黄小燕, 王宇光, 王怡,等. 麦冬皂苷D通过上调CYP2J2/EETs抗Ang Ⅱ诱导的内皮细胞凋亡[J]. 中国中药杂志, 2018,43(2):377-384.
[16] JIN Y, KHADKA D B, CHO W J. Pharmacological effects of berberine and its derivatives:a patent update[J]. Expert Opin Ther Pat, 2015, 26(2):229-243.
[17] 黄兴, 李艳芬, 寇冠军,等. 小檗碱抗动脉粥样硬化作用机制研究进展[J]. 药物评价研究, 2016, 39(3):469-473.
[18] XIONG C Y, FU Y H, HU H B, et al. Berberine inhibited apoptosis of human umbilical vein endothelial cells induced by Staphylocoocus aureus:an experimental research[J]. Zhongguo Zhong Xi Yi Jie He Za Zhi, 2014, 34(6):710-713.
[19] 郭君平, 王丽君, 王宝明,等. 小檗碱对脂多糖诱导的人脐静脉血管内皮细胞损伤的影响[J]. 中国急救医学, 2016, 36(9):789-793.
[20] KHODAYAR M J, JAVADIPOUR M, KESHTZAR E, et al. Role of berberine against arsenic induced oxidative damage in isolated rat liver mitochondria[J]. J Environ Biol, 2016, 37(2):285-290.