Abstract
Endothelial cell damage caused by oxidative stress is widely considered to be a triggering event in atherosclerosis (AS). However, the specific effect elicited by autophagy in endothelial cells undergoing oxidative stress remains controversial, especially during end-stage autophagy. The inhibition of end-stage autophagy has been reported to increase cell pyroptosis and contribute to endothelial damage. Several studies have shown that microRNA-103 is involved in end-stage autophagy; however, its specific mechanism of action is not yet characterized. In this study, we addressed the regulatory role of miR-103 in autophagy during oxidative stress of endothelial cells. Hydrogen peroxide (H2O2) treatment was used as an in vitro model of oxidative stress. MTS and ROS levels were measured to evaluate cell activity. qRT-PCR was used to detect the expression of miR-103. Autophagy was examined using western blot, immunofluorescence staining, and electron microscopy, while western blot analysis detected pyroptosis-related proteins. Results show that miR-103 expression decreased under oxidative stress. Further, miR-103 repressed transcription of Bcl-2/adenovirus E1B 19 kDa interacting protein (BNIP3). The oxidative stress caused by H2O2 caused cell damage from 2 hours (P < 0.05) and increased the level of intracellular reactive oxygen species (P < 0.05); at the same time, the damage could be further aggravated by the stimulation of bafA1 (P < 0.05). Under the stimulation of H2O2, the expression of miR-103 decreased (P < 0.05). However, high expression of miR-103 could reduce the accumulation of LC3II and P62 (P < 0.05) by inhibiting the downstream target gene Bcl-2/adenovirus E1B 19 kDa interacting protein (BNIP3), thus reducing the occurrence of cell pyroptosis (P < 0.05). This process could be blocked by end-stage autophagy inhibitor bafA1 (P < 0.05), which further indicated that miR-103 affected cell injury by autophagy. On the contrary, the low expression of miR-103 promoted the accumulation of autophagy protein and increased the occurrence of pyroptosis (P < 0.05). In conclusion, inhibition of miR-103 restrained end-stage of autophagy by regulating BNIP3, thus changing the occurrence of cell pyroptosis.
Highlights
Atherosclerosis (AS) is a chronic inflammatory response accompanied by a myriad of serious complications, representing an important cause of mortality and disability worldwide [1, 2]
We have described a signaling pathway associated with oxidative stress in coronary atherosclerosis
Our findings demonstrate that H2O2 decreases the expression of miR-103, resulting in the selective upregulation of BNIP3 in human coronary artery endothelial cells and, inhibits end-stage autophagy, in the overactivation of cell pyroptosis, with consequent endothelium injury
Summary
Atherosclerosis (AS) is a chronic inflammatory response accompanied by a myriad of serious complications, representing an important cause of mortality and disability worldwide [1, 2]. The role of autophagy in oxidative stress has been widely examined in recent years, few studies have addressed the role of end-stage autophagy, and those that have, yielded controversial results [7, 8]. MiRs play an important role in regulating autophagy in many diseases, especially cardiovascular diseases [14,15,16]; further studies are needed to investigate the role of miRs in the response to endothelial cell inflammatory injury through end-stage autophagy. Other studies have found that low miR-103 expression results in enhanced expression of the downstream target gene, phosphatase, and tensin homolog deleted on chromosome 10 (PTEN), which inhibits the p38 mitogen-activated protein kinase (MAPK) signaling pathway and reduces the inflammatory response and ER stress response in endothelial cells exposed to oxidized low-density lipoprotein (ox-LDL) [21]. Our results demonstrate that miR-103 protects endothelial cells from oxidative stress by suppressing BNIP3
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