Molecular mechanism of miR-142-3p in the injury induced by hydrogen peroxide by regulating ELAVL1

doi:10.3877/cma.j.issn.2095-1221.2019.03.002

Abstract

Abstract:

Objective

To investigate the effect of microRNA-142-3p (miR-142-3p) on hydrogen peroxide-induced cardiomyocyte injury and its mechanism.

Methods

The oxidative stress injury model of H9C2 cardiomyocytes was established. H9C2 cardiomyocytes were randomly divided into normal control group, H₂O₂ group, H₂O₂+miR-142-3p group, H₂O₂+miR negative control group, H₂O₂+ si-ELAVL1 group, H₂O₂+ group. siRNA control group, H₂O₂+miR-142-3p+pcDNA-ELAVL1 group, H₂O₂+miR-142-3p+pcDNA group. qRT-PCR and Western Blot were used to detect the expression of miR-142-3p and ELAVL1 in H9C2 cardiomyocytes, respectively. Fluorescence probe method was used to detect the level of reactive oxygen species (ROS) production in H9C2 cardiomyocytes of each group. MTT assay and flow cytometry were used to detect H9C2 cardiomyocyte survival rate and apoptosis rate, respectively. The dual luciferase reporter assay validated the targeting of miR-142-3p and ELAVL1. Western Blot was used to detect the expression of Cleaved Caspase-3, STAT3, Caspase-3 and p-STAT3.

Results

The expression of miR-142-?3p (0.26±0.06) , p-STAT3 expression (0.36±0.04) and cell viability (61.73±6.48) in the H₂O₂ group were significantly lower than those in the normal control group (P?< 0.01) . The ROS level (1566.38±121.57) , apoptosis rate (27.46±1.73) , Cleaved Caspase-3 (0.68±0.08) and ELAVL1 expression level (4.23±0.31) were significantly increased (P?< 0.01) . Dual luciferase reporter assay confirmed that ELAVL1 was a target gene of miR-142-3p. The miR-142-3p overexpression or silencing ELAVL1 could significantly promote the myocardial cell survival, up-regulate the p-STAT3 expression, while inhibit the cardiomyocyte apoptosis and expression of Cleaved Caspase-3.Overexpression of ELAVL1 reversed the protective effect of miR-142-3p on H9C2 cells treated with hydrogen peroxide.

Conclusion

miR-142-3p may attenuate the hydrogen peroxide-induced cardiomyocyte injury by inhibiting the ELAVL1 expression, which may protect cardiomyocytes by affecting the STAT3 signaling pathway.

Key words: MiR-142-3p, ELAVL1, Hydrogen peroxide, Cardiomyocytes

Sen Bing, Bo Yuan, Changyu Wang, Yi Wan, Li Wang, Jinguo Zou. Molecular mechanism of miR-142-3p in the injury induced by hydrogen peroxide by regulating ELAVL1[J]. Chinese Journal of Cell and Stem Cell(Electronic Edition), 2019, 09(03): 135-143.

/ / Recommend

Add to citation manager EndNote|Ris|BibTeX

URL: https://zhxbygxbzz.cma-cmc.com.cn/EN/10.3877/cma.j.issn.2095-1221.2019.03.002

https://zhxbygxbzz.cma-cmc.com.cn/EN/Y2019/V09/I03/135

Figures/Tables 15

References 21

[1]	Frangogiannis NG. Regulation of the inflammatory response in cardiac repair[J]. Circ Res, 2012, 110(1):159-173.
[2]	Gjesing A,Gislason GH,Køber L, et al. Nationwide trends in development of heart failure and mortality after first-time myocardial infarction 1997-2010: A Danish cohort study[J]. Eur J Intern Med, 2014, 25(8):731-738.
[3]	Carvajal K,Hafidi ME,Baños G. Myocardial damage due to ischemia and reperfusion in hypertriglyceridemic and hypertensive rats[J]. J Hypertens, 1999, 17(11):1607-1616.
[4]	Jin JK,Blackwood EA,Azizi K, et al. ATF6 decreases myocardial ischemia/reperfusion damage and links ER stress and oxidative stress signaling pathways in the heart[J]. Circ Res, 2017, 120(5):862-875.
[5]	Liu XR,Li T,Cao L, et al. Dexmedetomidine attenuates H₂O₂-induced neonatal rat cardiomyocytes apoptosis through mitochondria- and ER-medicated oxidative stress pathways[J]. Mol Med Rep, 2018, 17(5):7258-7264.
[6]	Wang Y,Ouyang M,Wang Q, et al. MicroRNA-142-3p inhibits hypoxia/reoxygenationinduced apoptosis and fibrosis of cardiomyocytes by targeting high mobility group box 1[J]. Int J Mol Med, 2016, 38(5):1377-1386.
[7]	Jeyabal P,Thandavarayan RA,Joladarashi D, et al. MicroRNA-9 inhibits hyperglycemia-induced pyroptosis in human ventricular cardiomyocytes by targeting ELAVL1[J]. Biochem Biophys Res Commun, 2016, 471(4):423-429.
[8]	曹瑞平,王文连,方婷婷, 等. 激活乙醛脱氢酶2的表达可减轻过氧化氢诱导的心肌细胞损伤[J]. 南方医科大学学报, 2018, 38(8):938-942.
[9]	符丽珍,黄茂芹,劳之勇, 等. miR-221在H₂O₂诱导的大鼠心肌细胞损伤中的作用及机制研究[J]. 中国比较医学杂志, 2017, 27(5):83-88.
[10]	王全伟,凡文博,王智昊, 等. 氧化应激与心血管疾病关系的研究进展[J]. 中国老年学杂志, 2014, 34(1):270-273.
[11]	Coskunpinar E,Cakmak HA,Kalkan AK, et al. Circulating miR- 221- 3p as a novel marker for early prediction of acute myocardial infarction[J]. Gene, 2016, 591(1):90-96.
[12]	Su Q,Li L,Zhao J, et al. MiRNA expression profile of the myocardial tissue of Pigs with coronary microembolization[J]. Cell Physiol Biochem, 2017, 43(3):1012-1024.
[13]	Su Q,Lv X,Ye Z, et al. The mechanism of miR-142-3p in coronary microembolization-induced myocardiac injury via regulating target gene IRAK-1[J]. Cell Death Dis, 2019, 10(2):61.
[14]	Liu BL,Cheng M,Hu S, et al. Overexpression of miR-142-3p improves mitochondrial function in cardiac hypertrophy[J]. Biomedicine c Pharmacotherapy, 2018, 108(1):1347-1356.
[15]	Qin B,Shu Y,Long L, et al. MicroRNA-142-3p induces Atherosclerosis-Associated endothelial cell apoptosis by directly targeting rictor[J]. Cell Physiol Biochem, 2018, 47(4):1589-1603.
[16]	Gu C,Zhang M,Sun W, et al. Upregulation of miR-324-5p inhibits proliferation and invasion of colorectal cancer cells by targeting ELAVL1[J]. Oncol Res, 2019, 27(5):515-524.
[17]	Li X,Zeng L,Cao C, et al. Long noncoding RNA MALAT1 regulates renal tubular epithelial pyroptosis by modulated miR-23c targeting of ELAVL1 in diabetic nephropathy[J]. Exp Cell Res, 2017, 350(2):327-335.
[18]	Nguyen MA,Karunakaran D,Geoffrion M, et al. Extracellular vesicles secreted by atherogenic macrophages transfer MicroRNA to inhibit cell migration[J]. Arterioscler Thromb Vasc Biol, 2018, 38(1):49-63.
[19]	卢叶,陈锐,万静, 等. 微小RNA-30a诱导人心肌细胞凋亡及其机制研究[J]. 临床和实验医学杂志, 2016, 15(22):2180-2185.
[20]	薛玉刚,程锦,曾广伟, 等. miR-214对过氧化氢诱导的心肌细胞凋亡的影响[J]. 临床和实验医学杂志, 2017, 16(17):1692-1695.
[21]	闫世林,罗永百,朱浩锋, 等. CaSR对过氧化氢作用后心肌细胞活性和凋亡影响[J]. 中国循证心血管医学杂志, 2018, 10(4):488-492.

分组	miR-142-3p	ELAVL1 mRNA	ELAVL1蛋白
正常对照组	1.00±0.12	1.01±0.08	0.39±0.04
H₂O₂组	0.26±0.06^a	4.23±0.31^a	0.59±0.06^a
t值	13.511	24.636	6.794
P值	< 0.01	< 0.01	< 0.01

分组	miR-142-3p	ELAVL1 mRNA	ELAVL1蛋白
正常对照组	1.00±0.12	1.01±0.08	0.39±0.04
H₂O₂组	0.26±0.06^a	4.23±0.31^a	0.59±0.06^a
t值	13.511	24.636	6.794
P值	< 0.01	< 0.01	< 0.01

分组	miR-142-3p	细胞存活率（﹪）	细胞凋亡率（﹪）	ROS（MFI）
正常对照组	1.00±0.09	100.00±4.89	6.83±0.66	468.43± 68.56
H₂O₂组	0.32±0.04^a	61.73±6.48^a	27.46±1.73^a	1367.17±103.25^a
H₂O₂+ miR阴性对照组	1.27±0.15	56.92±5.79	29.46±1.87	1456.76±121.49
H₂O₂+miR-142-3p组	3.86±0.42^b	79.54±6.28^b	16.43±0.96^b	796.28± 94.45
F值	276.674	66.123	337.789	136.377
P值	< 0.01	< 0.01	< 0.01	< 0.01

分组	miR-142-3p	细胞存活率（﹪）	细胞凋亡率（﹪）	ROS（MFI）
正常对照组	1.00±0.09	100.00±4.89	6.83±0.66	468.43± 68.56
H₂O₂组	0.32±0.04^a	61.73±6.48^a	27.46±1.73^a	1367.17±103.25^a
H₂O₂+ miR阴性对照组	1.27±0.15	56.92±5.79	29.46±1.87	1456.76±121.49
H₂O₂+miR-142-3p组	3.86±0.42^b	79.54±6.28^b	16.43±0.96^b	796.28± 94.45
F值	276.674	66.123	337.789	136.377
P值	< 0.01	< 0.01	< 0.01	< 0.01

分组	WT- ELAVL1	MUT- ELAVL1
miR阴性对照组	1.03±0.09	1.08±0.09
miR-142-3p组	0.58±0.07^a	0.99±0.08
t值	9.668	1.831
P值	0.000	0.097

Please choose a citation manager

Content to export