3,5-二咖啡酰奎宁酸通过激活PI3K/Akt信号通路保护H9C2细胞免受氧化应激诱导的凋亡。
3,5-Dicaffeoylquinic acid protects H9C2 cells against oxidative stress-induced apoptosis via activation of the PI3K/Akt signaling pathway.
作者信息
Bi Yi-Ming, Wu Yu-Ting, Chen Ling, Tan Zhang-Bin, Fan Hui-Jie, Xie Ling-Peng, Zhang Wen-Tong, Chen Hong-Mei, Li Jun, Liu Bin, Zhou Ying-Chun
机构信息
School of Traditional Chinese Medicine, Southern Medical University, Guangzhou, China.
Department of Traditional Chinese Medicine, Nanfang Hospital, Southern Medical University, Guangzhou, China.
出版信息
Food Nutr Res. 2018 Oct 12;62. doi: 10.29219/fnr.v62.1423. eCollection 2018.
BACKGROUND
Oxidative stress-induced apoptosis plays an important role in the development of heart failure. 3,5-Dicaffeoylquinic acid (3,5-diCQA), a phenolic compound, has shown protective effects against oxidative stress in many diseases.
OBJECTIVE
The objective of this study was to investigate the anti-apoptosis potential of 3,5-diCQA in cardiomyocyte cells under oxidative stress and explore its underlying mechanisms.
DESIGN
A model of tert-butyl hydroperoxide (TBHP)-induced apoptosis in a cardiomyocyte cell line (H9C2) was established. Cell viabilities on cell lines were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium (MTT) assay. The apoptosis was measured by hoechst33342 and propidium iodide (PI) fluorescent staining. PI (in red) stained the regions of cell apoptosis; Hoechet33342 (in blue) stained the nuclei. The Western blot was used to determine the expressions of related proteins such as p-PI3K: phosphorylated phosphatidylinositol-3-kinase (p-PI3K), phosphorylated Serine and Threonine kinase AKT (p-AKT), p-PTEN, Bcl-2, Bax, and caspase-3. Afterward, a PI3K inhibitor, LY294002, was applied to confirm the influence of the PI3K/Akt pathway on TBHP-treated cells of 3,5-diCQA. Then, H9C2 cells were pre-incubated with 3,5-diCQA alone to determine if the expression of activated PI3K/Akt signaling was mediated by 3,5-diCQA in H9C2 cells.
RESULTS
The results showed that TBHP resulted in an increase in cardiomyocyte apoptosis, whereas 3,5-diCQA treatment protected cells from TBHP-induced apoptosis in a dose-dependent manner. Moreover, 3,5-diCQA decreased expressions of Bax and caspase-3 but increased the phosphorylation levels of PI3K and Akt in TBHP-treated cells, which are the key molecules mediating cell survival, whereas phosphatase and tensin homologue deleted on chromosome 10 (PTEN) phosphorylation was unchanged. Importantly, pre-incubation with a PI3K inhibitor (LY294002) partly abolished the anti-apoptosis effects of 3,5-diCQA. Further, 3,5-diCQA enhanced the phosphorylation levels of PI3K and Akt in H9C2 cells directly, while LY294002 attenuated the effects of 3,5-diCQA on PI3K and Akt.
CONCLUSION
This study suggested that 3,5-diCQA rescued myocardium from apoptosis by increasing the activation of the PI3K/Akt signaling pathway.
背景
氧化应激诱导的细胞凋亡在心力衰竭的发生发展中起重要作用。3,5 - 二咖啡酰奎宁酸（3,5 - diCQA）是一种酚类化合物，在多种疾病中已显示出对氧化应激的保护作用。
目的
本研究旨在探讨3,5 - diCQA在氧化应激下心肌细胞中的抗凋亡潜力，并探索其潜在机制。
设计
建立叔丁基过氧化氢（TBHP）诱导心肌细胞系（H9C2）凋亡的模型。通过3 -（4,5 - 二甲基噻唑 - 2 - 基）- 2,5 - 二苯基四氮唑溴盐（MTT）法测定细胞系的细胞活力。通过Hoechst33342和碘化丙啶（PI）荧光染色检测细胞凋亡。PI（红色）染色细胞凋亡区域；Hoechet33342（蓝色）染色细胞核。采用蛋白质免疫印迹法检测相关蛋白如磷酸化磷脂酰肌醇 - 3 - 激酶（p - PI3K）、磷酸化丝氨酸和苏氨酸激酶AKT（p - AKT）、p - PTEN、Bcl - 2、Bax和半胱天冬酶 - 3的表达。随后，应用PI3K抑制剂LY294002来确认PI3K/Akt信号通路对3,5 - diCQA处理的TBHP诱导细胞的影响。然后，单独用3,5 - diCQA预孵育H9C2细胞，以确定H9C2细胞中活化的PI3K/Akt信号表达是否由3,5 - diCQA介导。
结果
结果表明，TBHP导致心肌细胞凋亡增加，而3,5 - diCQA处理以剂量依赖方式保护细胞免受TBHP诱导的凋亡。此外，3,5 - diCQA降低了TBHP处理细胞中Bax和半胱天冬酶 - 3的表达，但增加了PI3K和Akt的磷酸化水平，PI3K和Akt是介导细胞存活的关键分子，而10号染色体上缺失的磷酸酶和张力蛋白同源物（PTEN）的磷酸化未改变。重要的是，用PI3K抑制剂（LY294002）预孵育部分消除了3,5 - diCQA的抗凋亡作用。此外，3,5 - diCQA直接增强了H9C2细胞中PI3K和Akt的磷酸化水平，而LY294002减弱了3,5 - diCQA对PI3K和Akt的作用。
结论
本研究表明，3,5 - diCQA通过增加PI3K/Akt信号通路的激活来挽救心肌细胞免于凋亡。
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