MicroRNA‑494 suppresses hypoxia/reoxygenation‑induced cardiomyocyte apoptosis and autophagy via the PI3K/AKT/mTOR signaling pathway by targeting SIRT1

Ning,Shuwei; Li,Zhiying; Ji,Zhenyu; Fan,Dandan; Wang,Keke; Wang,Qian; Hua,Lei; Zhang,Junyue; Meng,Xiangguang; Yuan,Yiqiang

doi:10.3892/mmr.2020.11636

MicroRNA‑494 suppresses hypoxia/reoxygenation‑induced cardiomyocyte apoptosis and autophagy via the PI3K/AKT/mTOR signaling pathway by targeting SIRT1

Authors:
- Shuwei Ning
- Zhiying Li
- Zhenyu Ji
- Dandan Fan
- Keke Wang
- Qian Wang
- Lei Hua
- Junyue Zhang
- Xiangguang Meng
- Yiqiang Yuan
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Affiliations: Laboratory of Cardiovascular Disease and Drug Research, Zhengzhou No. 7 People's Hospital, Zhengzhou, Henan 450016, P.R. China, Institute of Medical and Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, Henan 450052, P.R. China, Department of Cardiovascular Internal Medicine, Henan Provincial Chest Hospital, Zhengzhou, Henan 450003, P.R. China
Published online on: October 26, 2020 https://doi.org/10.3892/mmr.2020.11636
Pages: 5231-5242
Copyright: © Ning et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Acute myocardial infarction can be caused by ischemia/reperfusion (I/R) injury; however, the mechanism underlying I/R is not completely understood. The present study investigated the functions and mechanisms underlying microRNA (miR)‑494 in I/R‑induced cardiomyocyte apoptosis and autophagy. Hypoxia/reoxygenation (H/R)‑treated H9c2 rat myocardial cells were used as an in vitro I/R injury model. Apoptosis and autophagy were analyzed by Cell Counting Kit‑8 assay, Lactic dehydrogenase and superoxide dismutase assay, flow cytometry, TUNEL staining and western blotting. Reverse transcription‑quantitative PCR demonstrated that, H9c2 cells treated with 12 h hypoxia and 3 h reoxygenation displayed significantly downregulated miR‑494 expression levels compared with control cells. Compared with the corresponding negative control (NC) groups, miR‑494 mimic reduced H/R‑induced cell apoptosis and autophagy, whereas miR‑494 inhibitor displayed the opposite effects. Silent information regulator 1 (SIRT1) was identified as a target gene of miR‑494. Furthermore, miR‑494 inhibitor‑mediated effects on H/R‑induced cardiomyocyte apoptosis and autophagy were partially reversed by SIRT1 knockdown. Moreover, compared with si‑NC, SIRT1 knockdown significantly increased the phosphorylation levels of PI3K, AKT and mTOR in H/R‑treated and miR‑494 inhibitor‑transfected H9c2 cells. Collectively, the results indicated that miR‑494 served a protective role against H/R‑induced cardiomyocyte apoptosis and autophagy by directly targeting SIRT1, suggesting that miR‑494 may serve as a novel therapeutic target for myocardial I/R injury.

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1	Gupta R and Wood DA: Primary prevention of ischaemic heart disease: Populations, individuals, and health professionals. Lancet. 394:685–696. 2019. View Article : Google Scholar : PubMed/NCBI
2	Nelson T, Garg P, Clayton RH and Lee J: The role of cardiac MRI in the management of ventricular arrhythmias in ischaemic and non-ischaemic dilated cardiomyopathy. Arrhythm Electrophysiol Rev. 8:191–201. 2019. View Article : Google Scholar : PubMed/NCBI
3	Hausenloy DJ and Yellon DM: Myocardial ischemia-reperfusion injury: A neglected therapeutic target. J Clin Invest. 123:92–100. 2013. View Article : Google Scholar : PubMed/NCBI
4	Rader DJ: Lysosomal acid lipase deficiency - A new therapy for a genetic lipid disease. N Engl J Med. 373:1071–1073. 2015. View Article : Google Scholar : PubMed/NCBI
5	Kaczanowski S: Apoptosis: Its origin, history, maintenance and the medical implications for cancer and aging. Phys Biol. 13:0310012016. View Article : Google Scholar : PubMed/NCBI
6	Khalil H, Abd ElHady A, Elawdan KA, Mohamed D, Mohamed DD, Abd El Maksoud AI, El-Chennawi FA, El-Fikiy B and El-Sayed IH: The mechanical autophagy as a part of cellular immunity; facts and features in treating the medical disorders. Immunol Invest. Sep 29–2020.(Epub ahead of print). doi: 10.1080/08820139.2020.1828453. View Article : Google Scholar
7	Dong Y, Chen H, Gao J, Liu Y, Li J and Wang J: Molecular machinery and interplay of apoptosis and autophagy in coronary heart disease. J Mol Cell Cardiol. 136:27–41. 2019. View Article : Google Scholar : PubMed/NCBI
8	Wang A, Zhang H, Liang Z, Xu K, Qiu W, Tian Y, Guo H, Jia J, Xing E, Chen R, et al: U0126 attenuates ischemia/reperfusion-induced apoptosis and autophagy in myocardium through MEK/ERK/EGR-1 pathway. Eur J Pharmacol. 788:280–285. 2016. View Article : Google Scholar : PubMed/NCBI
9	Sun MH, Chen XC, Han M, Yang YN, Gao XM, Ma X, Huang Y, Li XM, Gai MT, Liu F, et al: Cardioprotective effects of constitutively active MEK1 against H₂O₂-induced apoptosis and autophagy in cardiomyocytes via the ERK1/2 signaling pathway. Biochem Biophys Res Commun. 512:125–130. 2019. View Article : Google Scholar : PubMed/NCBI
10	Yuan H, Mischoulon D, Fava M and Otto MW: Circulating microRNAs as biomarkers for depression: Many candidates, few finalists. J Affect Disord. 233:68–78. 2018. View Article : Google Scholar : PubMed/NCBI
11	Liu K, Ma L, Zhou F, Yang Y, Hu HB, Wang L and Zhong L: Identification of microRNAs related to myocardial ischemic reperfusion injury. J Cell Physiol. 234:11380–11390. 2019. View Article : Google Scholar : PubMed/NCBI
12	Huang Z, Wu S, Kong F, Cai X, Ye B, Shan P and Huang W: MicroRNA-21 protects against cardiac hypoxia/reoxygenation injury by inhibiting excessive autophagy in H9c2 cells via the Akt/mTOR pathway. J Cell Mol Med. 21:467–474. 2017. View Article : Google Scholar : PubMed/NCBI
13	Lu TX and Rothenberg ME: MicroRNA. J Allergy Clin Immunol. 141:1202–1207. 2018. View Article : Google Scholar : PubMed/NCBI
14	Jin Y and Ni S: miR-496 remedies hypoxia reoxygenation-induced H9c2 cardiomyocyte apoptosis via Hook3-targeted PI3k/Akt/mTOR signaling pathway activation. J Cell Biochem. 121:698–712. 2020. View Article : Google Scholar : PubMed/NCBI
15	Huang ZQ, Xu W, Wu JL, Lu X and Chen XM: MicroRNA-374a protects against myocardial ischemia-reperfusion injury in mice by targeting the MAPK6 pathway. Life Sci. 232:1166192019. View Article : Google Scholar : PubMed/NCBI
16	Tan H, Qi J, Fan BY, Zhang J, Su FF and Wang HT: MicroRNA-24-3p Attenuates myocardial ischemia/reperfusion injury by suppressing RIPK1 expression in mice. Cell Physiol Biochem. 51:46–62. 2018. View Article : Google Scholar : PubMed/NCBI
17	Wu A, Lou L, Zhai J, Zhang D, Chai L, Nie B, Zhu H, Gao Y, Shang H and Zhao M: miRNA expression profile and effect of wenxin granule in rats with ligation-induced myocardial infarction. Int J Genomics. 2017:21758712017. View Article : Google Scholar : PubMed/NCBI
18	Zhai F, Zhang X, Guan Y, Yang X, Li Y, Song G and Guan L: Expression profiles of microRNAs after focal cerebral ischemia/reperfusion injury in rats. Neural Regen Res. 7:917–923. 2012.PubMed/NCBI
19	Sun G, Zhou Y, Li H, Guo Y, Shan J, Xia M, Li Y, Li S, Long D and Feng L: Over-expression of microRNA-494 up-regulates hypoxia-inducible factor-1 alpha expression via PI3K/Akt pathway and protects against hypoxia-induced apoptosis. J Biomed Sci. 20:1002013. View Article : Google Scholar : PubMed/NCBI
20	Wang X, Zhang X, Ren XP, Chen J, Liu H, Yang J, Medvedovic M, Hu Z and Fan GC: MicroRNA-494 targeting both proapoptotic and antiapoptotic proteins protects against ischemia/reperfusion-induced cardiac injury. Circulation. 122:1308–1318. 2010. View Article : Google Scholar : PubMed/NCBI
21	Meng X, Tan J, Li M, Song S, Miao Y and Zhang Q: Sirt1: Role under the condition of ischemia/hypoxia. Cell Mol Neurobiol. 37:17–28. 2017. View Article : Google Scholar : PubMed/NCBI
22	Yuan Y, Cruzat VF, Newsholme P, Cheng J, Chen Y and Lu Y: Regulation of SIRT1 in aging: Roles in mitochondrial function and biogenesis. Mech Ageing Dev. 155:10–21. 2016. View Article : Google Scholar : PubMed/NCBI
23	Zhang W, Huang Q, Zeng Z, Wu J, Zhang Y and Chen Z: Sirt1 inhibits oxidative stress in vascular endothelial cells. Oxid Med Cell Longev. 2017:75439732017. View Article : Google Scholar : PubMed/NCBI
24	Han Y, Luo H, Wang H, Cai J and Zhang Y: SIRT1 induces resistance to apoptosis in human granulosa cells by activating the ERK pathway and inhibiting NF-kappaB signaling with anti-inflammatory functions. Apoptosis. 22:1260–1272. 2017. View Article : Google Scholar : PubMed/NCBI
25	Li Y, Wang P, Yang X, Wang W, Zhang J, He Y, Zhang W, Jing T, Wang B and Lin R: SIRT1 inhibits inflammatory response partly through regulation of NLRP3 inflammasome in vascular endothelial cells. Mol Immunol. 77:148–156. 2016. View Article : Google Scholar : PubMed/NCBI
26	Liu Y, Li X, Zhu S, Zhang JG, Yang M, Qin Q, Deng SC, Wang B, Tian K, Liu L, et al: Ectopic expression of miR-494 inhibited the proliferation, invasion and chemoresistance of pancreatic cancer by regulating SIRT1 and c-Myc. Gene Ther. 22:729–738. 2015. View Article : Google Scholar : PubMed/NCBI
27	Li H, He C, Wang X, Wang H, Nan G and Fang L: MicroRNA-183 affects the development of gastric cancer by regulating autophagy via MALAT1-miR-183-SIRT1 axis and PI3K/AKT/mTOR signals. Artif Cells Nanomed Biotechnol. 47:3163–3171. 2019. View Article : Google Scholar : PubMed/NCBI
28	Alves-Fernandes DK and Jasiulionis MG: The role of SIRT1 on DNA damage response and epigenetic alterations in cancer. Int J Mol Sci. 20:31532019. View Article : Google Scholar
29	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
30	Mao GX, Xu XG, Wang SY, Li HF, Zhang J, Zhang ZS, Su HL, Chen SS, Xing WM, Wang YZ, et al: Salidroside delays cellular senescence by stimulating mitochondrial biogenesis partly through a miR-22/SIRT-1 pathway. Oxid Med Cell Longev. 2019:52760962019. View Article : Google Scholar : PubMed/NCBI
31	Zeyad A, Hamad M, Amor H and Hammadeh ME: Relationships between bacteriospermia, DNA integrity, nuclear protamine alteration, sperm quality and ICSI outcome. Reprod Biol. 18:115–121. 2018. View Article : Google Scholar : PubMed/NCBI
32	Xu Z, Han X, Ou D, Liu T, Li Z, Jiang G, Liu J and Zhang J: Targeting PI3K/AKT/mTOR-mediated autophagy for tumor therapy. Appl Microbiol Biotechnol. 104:575–587. 2020. View Article : Google Scholar : PubMed/NCBI
33	Diez ER, Altamirano LB, García IM, Mazzei L, Prado NJ, Fornes MW, Carrión FD, Zumino AZ, Ferder L and Manucha W: Heart remodeling and ischemia-reperfusion arrhythmias linked to myocardial vitamin d receptors deficiency in obstructive nephropathy are reversed by paricalcitol. J Cardiovasc Pharmacol Ther. 20:211–220. 2015. View Article : Google Scholar : PubMed/NCBI
34	Yao L, Chen H, Wu Q and Xie K: Hydrogen-rich saline alleviates inflammation and apoptosis in myocardial I/R injury via PINK-mediated autophagy. Int J Mol Med. 44:1048–1062. 2019.PubMed/NCBI
35	Majtnerová P and Roušar T: An overview of apoptosis assays detecting DNA fragmentation. Mol Biol Rep. 45:1469–1478. 2018. View Article : Google Scholar : PubMed/NCBI
36	D'Arcy MS: Cell death: A review of the major forms of apoptosis, necrosis and autophagy. Cell Biol Int. 43:582–592. 2019. View Article : Google Scholar : PubMed/NCBI
37	Kasprowska-Liśkiewicz D: The cell on the edge of life and death: Crosstalk between autophagy and apoptosis. Postepy Hig Med Dosw. 71:825–841. 2017. View Article : Google Scholar
38	Doherty J and Baehrecke EH: Life, death and autophagy. Nat Cell Biol. 20:1110–1117. 2018. View Article : Google Scholar : PubMed/NCBI
39	Zhai C, Tang G, Peng L, Hu H, Qian G, Wang S, Yao J, Zhang X, Fang Y, Yang S, et al: Inhibition of microRNA-1 attenuates hypoxia/re-oxygenation-induced apoptosis of cardiomyocytes by directly targeting Bcl-2 but not GADD45Beta. Am J Transl Res. 7:1952–1962. 2015.PubMed/NCBI
40	Gao CK, Liu H, Cui CJ, Liang ZG, Yao H and Tian Y: Roles of MicroRNA-195 in cardiomyocyte apoptosis induced by myocardial ischemia-reperfusion injury. J Genet. 95:99–108. 2016. View Article : Google Scholar : PubMed/NCBI
41	Tian ZQ, Jiang H and Lu ZB: MiR-320 regulates cardiomyocyte apoptosis induced by ischemia-reperfusion injury by targeting AKIP1. Cell Mol Biol Lett. 23:412018. View Article : Google Scholar : PubMed/NCBI
42	Fan ZX and Yang J: The role of microRNAs in regulating myocardial ischemia reperfusion injury. Saudi Med J. 36:787–793. 2015. View Article : Google Scholar : PubMed/NCBI
43	Yu X, Zhang S, Zhao D, Zhang X, Xia C, Wang T, Zhang M, Liu T, Huang W and Wu B: SIRT1 inhibits apoptosis in in vivo and in vitro models of spinal cord injury via microRNA-494. Int J Mol Med. 43:1758–1768. 2019.PubMed/NCBI
44	Tang Q, Len Q, Liu Z and Wang W: Overexpression of miR-22 attenuates oxidative stress injury in diabetic cardiomyopathy via Sirt 1. Cardiovasc Ther. Dec 29–2017.(Epub ahead of print). doi: 10.1111/1755-5922.12318.
45	Luo G, Jian Z, Zhu Y, Zhu Y, Chen B, Ma R, Tang F and Xiao Y: Sirt1 promotes autophagy and inhibits apoptosis to protect cardiomyocytes from hypoxic stress. Int J Mol Med. 43:2033–2043. 2019.PubMed/NCBI
46	Potenza MA, Sgarra L, Nacci C, Leo V, De Salvia MA and Montagnani M: Activation of AMPK/SIRT1 axis is required for adiponectin-mediated preconditioning on myocardial ischemia-reperfusion (I/R) injury in rats. PLoS One. 14:e02106542019. View Article : Google Scholar : PubMed/NCBI
47	Huang G, Hao F and Hu X: Downregulation of microRNA-155 stimulates sevoflurane-mediated cardioprotection against myocardial ischemia/reperfusion injury by binding to SIRT1 in mice. J Cell Biochem. 120:15494–15505. 2019. View Article : Google Scholar : PubMed/NCBI
48	Ding S, Liu D, Wang L, Wang G and Zhu Y: Inhibiting MicroRNA-29a protects myocardial ischemia-reperfusion injury by targeting SIRT1 and suppressing oxidative stress and NLRP3-mediated pyroptosis pathway. J Pharmacol Exp Ther. 372:128–135. 2020. View Article : Google Scholar : PubMed/NCBI
49	Hsu CP, Zhai P, Yamamoto T, Maejima Y, Matsushima S, Hariharan N, Shao D, Takagi H, Oka S and Sadoshima J: Silent information regulator 1 protects the heart from ischemia/reperfusion. Circulation. 122:2170–2182. 2010. View Article : Google Scholar : PubMed/NCBI
50	McKenna M, McGarrigle S and Pidgeon GP: The next generation of PI3K-Akt-mTOR pathway inhibitors in breast cancer cohorts. Biochim Biophys Acta Rev Cancer. 1870:185–197. 2018. View Article : Google Scholar : PubMed/NCBI
51	Chi Y, Ma Q, Ding XQ, Qin X, Wang C and Zhang J: Research on protective mechanism of ibuprofen in myocardial ischemia-reperfusion injury in rats through the PI3K/Akt/mTOR signaling pathway. Eur Rev Med Pharmacol Sci. 23:4465–4473. 2019.PubMed/NCBI
52	Li X, Hu X, Wang J, Xu W, Yi C, Ma R and Jiang H: Inhibition of autophagy via activation of PI3K/Akt/mTOR pathway contributes to the protection of hesperidin against myocardial ischemia/reperfusion injury. Int J Mol Med. 42:1917–1924. 2018.PubMed/NCBI
53	Wang H, Liu H, Chen K, Xiao J, He K, Zhang J and Xiang G: SIRT1 promotes tumorigenesis of hepatocellular carcinoma through PI3K/PTEN/AKT signaling. Oncol Rep. 28:311–318. 2012.PubMed/NCBI
54	Zhang W, Zhang Y, Wang Z, Xu T, Huang C, Yin W, Wang J, Xiong W, Lu W, Zheng H, et al: Tris(2-chloroethyl)phosphate-induced cell growth arrest via attenuation of SIRT1-independent PI3K/Akt/mTOR pathway. J Appl Toxicol. 36:914–924. 2016. View Article : Google Scholar : PubMed/NCBI