MicroRNA-499-5p Inhibits Transforming Growth Factor-β1-induced Smad2 Signaling Pathway and Suppresses Fibroblast Proliferation and Collagen Synthesis in Rat by Targeting TGFβ-R1

Overview

Journal Mol Biol Rep

Specialty Molecular Biology

Date 2023 Sep 7

PMID 37676431

Authors

Qing Zhao

Wentao Yang

Xiangdong Li

Hongtao Yuan

Jianping Guo

Yutang Wang

Zhaoliang Shan

Affiliations

Soon will be listed here.

Abstract

Background: Artial fibrosis has been recognized as a typical pathological change in atrial fibrillation. Although present evidence suggests that microRNA-499-5p (miR-499-5p) plays an important role in the development of atrial fibrosis, the specific mechanism is not fully understood. Therefore, this study attempted to assess the influence of miR-499-5p on atrial fibroblasts and explore the potential molecular mechanism.

Methods: Atrial fibroblasts from sprague dawley rat were respectively transfected with miR-499-5p mimic, miR-499-5p negative control and miR-499-5p inhibitor, atrial fibroblasts without any treatment were also established. Cell counting kit-8 assay and transwell assay were used to detect the proliferation and migration of atrial fibroblasts in each group. Expressions of miR-499-5p, TGF-β1, smad2, α-SMA, collagen-I and TGFβ-R1 in mRNA and protein level were subsequently detected via quantitative real-time polymerase chain reaction and western blot. Furthermore, the prediction of the binding sites of miR-499-5p and TGFβ-R1 was performed via the bioinformatics online software TargetScan and verified by dual luciferase reporter.

Results: By utilizing miR-499-5p-transfected atrial fibroblasts model, expression of miR-499-5p in the miR-499-5p mimic group was upregulated, while it was downregulated in the miR-499-5p inhibitors group. Upregulated miR-499-5p expression led to to a significant decrease in the proliferative and migratory ability of cultured atrial fibroblasts, while downregulated miR-499-5p expression led to a significant increase in the proliferative and migratory ability of cultured atrial fibroblasts. Additionally, upregulated miR-499-5p expression made a significant rise in TGF-β1-induced mRNA and protein expression of TGF-β1, TGFβ-R1, smad2, α-SMA and collagen-I in atrial fibroblasts. Furthermore, results from the dual luciferase reporter conformed that miR-499-5p may repress TGFβ-R1 by binding the 3'UTR of TGFβ-R1 directly.

Conclusions: miR-499-5p is able to inhibit the activation of transforming growth factor β-induced Smad2 signaling and eventually suppressed the proliferation, migration and invasion of atrial fibroblasts and collagen synthesis by targeting TGFβ-R1.

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References

Xin Y, Yang C, Han Z . Circulating miR-499 as a potential biomarker for acute myocardial infarction. Ann Transl Med. 2016; 4(7):135. PMC: 4842393. DOI: 10.21037/atm.2016.03.40. View

Matkovich S, Hu Y, Eschenbacher W, Dorn L, Dorn 2nd G . Direct and indirect involvement of microRNA-499 in clinical and experimental cardiomyopathy. Circ Res. 2012; 111(5):521-31. PMC: 3429338. DOI: 10.1161/CIRCRESAHA.112.265736. View

Wojciechowska A, Braniewska A, Kozar-Kaminska K . MicroRNA in cardiovascular biology and disease. Adv Clin Exp Med. 2017; 26(5):865-874. DOI: 10.17219/acem/62915. View

Zhao C, Cheng G, He R, Hong Y, Wan Q, Wang Z . Analysis and clinical significance of microRNA-499 expression levels in serum of patients with acute myocardial infarction. Genet Mol Res. 2015; 14(2):4027-34. DOI: 10.4238/2015.April.27.17. View

Dong S, Cheng Y, Yang J, Li J, Liu X, Wang X . MicroRNA expression signature and the role of microRNA-21 in the early phase of acute myocardial infarction. J Biol Chem. 2009; 284(43):29514-25. PMC: 2785585. DOI: 10.1074/jbc.M109.027896. View

Li Y, Lu J, Bao X, Wang X, Wu J, Li X . MiR-499-5p protects cardiomyocytes against ischaemic injury via anti-apoptosis by targeting PDCD4. Oncotarget. 2016; 7(24):35607-35617. PMC: 5094948. DOI: 10.18632/oncotarget.9597. View

Shen N, Zhang C, Li Z, Kong L, Wang X, Gu Z . MicroRNA expression signatures of atrial fibrillation: The critical systematic review and bioinformatics analysis. Exp Biol Med (Maywood). 2019; 245(1):42-53. PMC: 6987745. DOI: 10.1177/1535370219890303. View

Burstein B, Libby E, Calderone A, Nattel S . Differential behaviors of atrial versus ventricular fibroblasts: a potential role for platelet-derived growth factor in atrial-ventricular remodeling differences. Circulation. 2008; 117(13):1630-41. DOI: 10.1161/CIRCULATIONAHA.107.748053. View

Nattel S . Molecular and Cellular Mechanisms of Atrial Fibrosis in Atrial Fibrillation. JACC Clin Electrophysiol. 2018; 3(5):425-435. DOI: 10.1016/j.jacep.2017.03.002. View

10.

Xu J, Wu H, Chen S, Qi B, Zhou G, Cai L . MicroRNA-30c suppresses the pro-fibrogenic effects of cardiac fibroblasts induced by TGF-β1 and prevents atrial fibrosis by targeting TGFβRII. J Cell Mol Med. 2018; 22(6):3045-3057. PMC: 5980214. DOI: 10.1111/jcmm.13548. View

11.

Lee T, Yeh C, Lee Y, Shih Y, Chen Y, Hung C . Fibroblast-enriched endoplasmic reticulum protein TXNDC5 promotes pulmonary fibrosis by augmenting TGFβ signaling through TGFBR1 stabilization. Nat Commun. 2020; 11(1):4254. PMC: 7449970. DOI: 10.1038/s41467-020-18047-x. View

12.

Feinberg M, Moore K . MicroRNA Regulation of Atherosclerosis. Circ Res. 2016; 118(4):703-20. PMC: 4762069. DOI: 10.1161/CIRCRESAHA.115.306300. View

13.

Wang Y, Cai H, Li H, Gao Z, Song K . Atrial overexpression of microRNA-27b attenuates angiotensin II-induced atrial fibrosis and fibrillation by targeting ALK5. Hum Cell. 2018; 31(3):251-260. DOI: 10.1007/s13577-018-0208-z. View

14.

Wang J, Jia Z, Zhang C, Sun M, Wang W, Chen P . miR-499 protects cardiomyocytes from H 2O 2-induced apoptosis via its effects on Pdcd4 and Pacs2. RNA Biol. 2014; 11(4):339-50. PMC: 4075519. DOI: 10.4161/rna.28300. View

15.

Vander Ark A, Cao J, Li X . TGF-β receptors: In and beyond TGF-β signaling. Cell Signal. 2018; 52:112-120. DOI: 10.1016/j.cellsig.2018.09.002. View

16.

Shi Y, Han Y, Niu L, Li J, Chen Y . MiR-499 inhibited hypoxia/reoxygenation induced cardiomyocytes injury by targeting SOX6. Biotechnol Lett. 2019; 41(6-7):837-847. PMC: 6551346. DOI: 10.1007/s10529-019-02685-3. View

17.

Liu L, Gong W, Zhang S, Shen J, Wang Y, Chen Y . Hydrogen Sulfide Attenuates Angiotensin II-Induced Cardiac Fibroblast Proliferation and Transverse Aortic Constriction-Induced Myocardial Fibrosis through Oxidative Stress Inhibition via Sirtuin 3. Oxid Med Cell Longev. 2021; 2021:9925771. PMC: 8486544. DOI: 10.1155/2021/9925771. View

18.

Feng Y, Bao Y, Ding J, Li H, Liu W, Wang X . MicroRNA-130a attenuates cardiac fibrosis after myocardial infarction through TGF-β/Smad signaling by directly targeting TGF-β receptor 1. Bioengineered. 2022; 13(3):5779-5791. PMC: 8973730. DOI: 10.1080/21655979.2022.2033380. View

19.

Gal P, Marrouche N . Magnetic resonance imaging of atrial fibrosis: redefining atrial fibrillation to a syndrome. Eur Heart J. 2015; 38(1):14-19. DOI: 10.1093/eurheartj/ehv514. View

20.

Centurion O, Alderete J, Torales J, Garcia L, Scavenius K, Mino L . Myocardial Fibrosis as a Pathway of Prediction of Ventricular Arrhythmias and Sudden Cardiac Death in Patients With Nonischemic Dilated Cardiomyopathy. Crit Pathw Cardiol. 2019; 18(2):89-97. DOI: 10.1097/HPC.0000000000000171. View