Myosin Heavy Chain 7 Gene-derived miRNA-208b-3p Enhances the Fibrosis-related Gene Expression in Cardiac Fibroblasts

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Myosin Heavy Chain 7 Gene-derived miRNA-208b-3p Enhances the Fibrosis-related Gene Expression in Cardiac Fibroblasts

Basic Research | 更新时间：2024-02-19

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- Myosin Heavy Chain 7 Gene-derived miRNA-208b-3p Enhances the Fibrosis-related Gene Expression in Cardiac Fibroblasts
- Journal of Sun Yat-sen University(Medical Sciences) Vol. 44, Issue 4, Pages: 642-650(2023)
- 作者机构：
  
  1.南方医科大学附属广东省人民医院//广东省医学科学院医学研究部，广东广州 510080
  2.南方医科大学附属广东省人民医院//广东省医学科学院检验科，广东广州 510080
  3.南方医科大学附属广东省人民医院//广东省医学科学院麻醉科，广东广州 510080
- 作者简介：
  
  SHAN Zhi-xin； E-mail： shanzhixin@gdph.org.cn
- 基金信息：
- DOI：10.13471/j.cnki.j.sun.yat-sen.univ(med.sci).20230421.002
  CLC： R363.2
- Published：20 July 2023，
  
  Received：04 December 2022，
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张梦珍,翟琳,郭林林等.肌球蛋白重链7基因来源的miR-208b-3p促进心肌成纤维细胞中纤维化相关基因表达[J].中山大学学报（医学科学版）,2023,44(04):642-650.

ZHANG Meng-zhen,ZHAI Lin,GOU Lin-lin,et al.Myosin Heavy Chain 7 Gene-derived miRNA-208b-3p Enhances the Fibrosis-related Gene Expression in Cardiac Fibroblasts[J].Journal of Sun Yat-sen University(Medical Sciences),2023,44(04):642-650.
张梦珍,翟琳,郭林林等.肌球蛋白重链7基因来源的miR-208b-3p促进心肌成纤维细胞中纤维化相关基因表达[J].中山大学学报（医学科学版）,2023,44(04):642-650. DOI： 10.13471/j.cnki.j.sun.yat-sen.univ(med.sci).20230421.002.

ZHANG Meng-zhen,ZHAI Lin,GOU Lin-lin,et al.Myosin Heavy Chain 7 Gene-derived miRNA-208b-3p Enhances the Fibrosis-related Gene Expression in Cardiac Fibroblasts[J].Journal of Sun Yat-sen University(Medical Sciences),2023,44(04):642-650. DOI： 10.13471/j.cnki.j.sun.yat-sen.univ(med.sci).20230421.002.

摘要

目的

2

探究肌球蛋白重链7基因来源的微小RNA-208b-3p（miR-208b-3p）对心肌成纤维细胞纤维化表型的调控作用。

方法

2

通过miRNA 表达谱芯片分析18周龄的糖尿病db/db小鼠和db/m对照小鼠心肌中差异的miRNAs。通过实时荧光定量PCR（RT-qPCR）检测miR-208b-3p 在血管紧张素Ⅱ（Ang Ⅱ）和高糖/葡萄糖氧化酶（G/Go）处理的C57BL/6乳小鼠心肌细胞（NMVCs）和心肌成纤维细胞（mCFs）中的表达；利用CCK8细胞增殖实验、流式细胞术和纤维化相关蛋白（包括COL1A1、COL3A1和α-SMA）表达检测来了解miR-208b-3p对mCFs纤维化表型的影响；双萤光素酶报告基因实验检测miR-208b-3p与潜在靶基因Mtf2和Pgrmc1 3'端非翻译区（3'-UTR）的结合作用；利用RT-qPCR和Western blot 法分别检测miR-208b-3p转染的 mCFs中Mtf2 和Pgrmc1 表达；通过小干扰RNA（siRNA）抑制Mtf2和Pgrmc1表达，并检测对mCFs中纤维化相关蛋白表达的作用。

结果

2

miRNA 表达谱和RT-qPCR结果证实miR-208b-3p在糖尿病db/db小鼠心肌中表达显著升高。miR-208b-3p前体与宿主基因Myh7在db/db小鼠心肌中表达显著升高，miR-208b-3p与Myh7 mRNA在乳小鼠来源的mCFs和NMVCs中均有表达，但在NMVCs中水平更高。miR-208b-3p在Ang Ⅱ和G/Go处理后的mCFs和NMVCs中表达均明显升高。miR-208b-3p可显著增加mCFs中纤维化相关蛋白COL1A1、COL3A1和α-SMA表达，但不影响mCFs的增殖能力和细胞周期分布特征。双萤光素酶报告基因实验显示miR-208b-3p与Mtf2和Pgrmc1 3'-UTR有结合作用。miR-208b-3p可在转录后水平抑制mCFs中Mtf2和Pgrmc1表达。miR-208b-3p、Mtf2 siRNA和Pgrmc1 siRNA均能一致性地促进mCFs中纤维化相关蛋白表达。

结论

2

miR-208b-3p通过靶向Mtf2和Pgrmc1基因发挥促进mCFs中纤维化相关基因表达的作用。

Abstract

Objective

2

To investigate the effect of myosin heavy chain 7 gene-derived miRNA-208b-3p on the fibrotic phenotype of cardiac fibroblasts.

Methods

2

miRNA chip array was performed to detect the dysregulated miRNAs in the myocardium of diabetic db/db mice and db/m control mice. Neonatal mouse ventricular cardiomyocytes （NMVCs） and cardiac fibroblasts （CFs） were isolated from C57BL/6 mice and cultured. Real-time quantitative PCR （RT-qPCR） was conducted to determine the expression of miR-208b-3p in mouse CFs and NMVCs subjected to angiotensinⅡ（AngⅡ） and high glucose plus glucose oxidase （G/Go） treatment， respectively. Cell counting kit 8（CCk8） assay， flow cytometry and determination of fibrosis-related protein， including COL1A1， COL3A1and α-SMA， were performed in mCFs transfected with miR-208b-3p. Dual luciferase reporter assay was performed to confirm the interaction between miR-208b-3p and the 3'-UTR of metal response element binding transcription factor 2 （Mtf2） and progesterone receptor membrane component 1（Pgrmc1）， respectively. The expressions of Mtf2 and Pgrmc1 at the mRNA and protein levels in mCFs after miR-208b-3p mimic transfection were determined using RT-qPCR and Western blot assay， respectively. The small interfering RNA （siRNA） was used to inhibit Mtf2 and Pgrmc1 expression in mCFs， and the effects of Mtf2 siRNA， Pgrmc1 siRNA and miR-208b-3p on fibrosis-related protein expression in mCFs were investigated.

Results

2

Results of miRNA chip array and RT-qPCR assay showed that miR-208b-3p was up-regulated in the myocardium of the diabetic db/db mice. miR-208b precursor and the host gene of Myh7 were consistently increased in db/db mice. miR-208b-3p and Myh7 mRNA were expressed in mCFs and NMVCs， but the levels of miR-208b-3p and Myh7 mRNA in NMVCs were much higher than those in mCFs. miR-208b-3p was up-regulated in mCFs and NMVCs subjected to Ang Ⅱ and G/Go treatment， respectively. miR-208b-3p could significantly enhance fibrosis-related protein， including COL1A1， COL3A1 and α-SMA， in mCFs， without affecting the proliferation activity and cell cycle distribution of mCFs. Dual luciferase reporter assay revealed the interactions of miR-208b-3p with the 3'-UTR of Mtf2 and Pgrmc1. The results of RT-qPCR and Western blotting confirmed that miR-208b-3p inhibited Mtf2 and Pgrmc1 expression at the post- transcriptional level. Transfection with miR-208b-3p mimic， Mtf2 siRNA and Pgrmc1 siRNA could consistently enhance the fibrosis-related protein expression in the cardiac fibroblasts.

Conclusions

2

miR-208b-3p enhances fibrosis-related gene expression by targeting Mtf2 and Pgrmc1in mCFs.

关键词

心肌纤维化微小RNAmiR-208b-3p心肌成纤维细胞

Keywords

cardiac fibrosismicroRNAmiR-208b-3pcardiac fibroblast

references

Rubler S， Dlugash J， Yuceoglu YZ， et al. New type of cardiomyopathy associated with diabetic glomerulosclerosis［J］. Am J Cardiol， 1972， 30：595-602.

Regan TJ， Lyons MM， Ahmed SS， et al. Evidence for cardiomyopathy in familial diabetes mellitus［J］. J Clin Invest， 1977， 60（4）：884-899.

Park S， Nguyen NB， Pezhouman A， et al. Cardiac fibrosis： potential therapeutic targets［J］. Transl Res， 2019， 209：121-137.

Saliminejad K， Khorram Khorshid HR， Soleymani Fard S， et al. An overview of microRNAs： biology， functions， therapeutics， and analysis methods［J］. J Cell Physiol， 2019， 234（5）：5451-5465.

陈丽文，郭晶，陈泽润，等. MicroRNA-99b-5p通过抑制成纤维细胞生长因子21表达促进心肌细胞肥大［J］. 中山大学学报（医学科学版）， 2022， 43（2）：192-202.

Chen LW， Guo J， Chen ZR， et al. MicroRNA-99b-5p aggravates cardiomyocyte hypertrophy by down-regulating fgf21［J］. J Sun Yat-sen Univ （Med Sci）， 2022， 43（2）：192-202.

易芷瑶，赵安职，张铭，等. 核内miR-199b-5p通过上调CDK9表达促进心肌细胞肥大［J］. 中国病理生理杂志， 2021， 37（2）：193-201.

Yi ZY， Zhao AZ， Zhang M， et al. Nuclear miR-199b-5p promotes cardiomyocyte hypertrophy by up-regulating CDK9 expression ［J］. Chin J Pathophysiol， 2021， 37（2）：193-201.

赵安职，郭晶，陈丽文，等. miR-25-3p通过BTG2/SOD2轴促进心肌成纤维细胞中纤维化相关基因表达［J］. 中国动脉硬化杂志， 2022， 30（4）： 328-334； +351.

Zhao AZ， Guo J， Chen LW， et al. MiR-25-3p promotes the expression of fibrosis-related genes in cardiac fibroblasts through BTG2/ SOD2 axis［J］. Chin J Arterioscler， 2022， 30（4）： 328-334； +351.

Wojciechowska A， Braniewska A， Kozar-Kamińska K. MicroRNA in cardiovascular biology and disease［J］. Adv Clin Exp Med， 2017， 26（5）：865-874.

Ahmed U， Khaliq S， Ahmad HU， et al. Pathogenesis of diabetic cardiomyopathy and role of miRNA［J］. Crit Rev Eukaryot Gene Expr， 2021，31（1）：79-92.

van Rooij E， Quiat D， Johnson BA， et al. A family of microRNAs encoded by myosin genes governs myosin expression and muscle performance［J］. Dev Cell， 2009，17（5）：662-673.

Mekala N， Kurdys J， Vicenzi AP， et al. MiR 208a regulates mitochondrial biogenesis in metabolically challenged cardiomyocytes［J］. Cells， 2021，10（11）：3152.

Blumensatt M， Fahlbusch P， Hilgers R， et al. Secretory products from epicardial adipose tissue from patients with type 2 diabetes impair mitochondrial β-oxidation in cardiomyocytes via activation of the cardiac renin-angiotensin system and induction of miR-208a［J］. Basic Res Cardiol， 2017，112（1）：2.

Rawal S， Nagesh PT， Coffey S， et al. Early dysregulation of cardiac-specific microRNA-208a is linked to maladaptive cardiac remodelling in diabetic myocardium［J］. Cardiovasc Diabetol， 2019，18（1）：13.

Zhao L， Li W， Zhao H. Inhibition of long non-coding RNA TUG1 protects against diabetic cardiomyopathy induced diastolic dysfunction by regulating miR-499-5p［J］. Am J Transl Res， 2020， 12（3）：718-730.

Mathiyalagan P， Okabe J， Chang L， et al. The primary microRNA-208b interacts with Polycomb-group protein， Ezh2， to regulate gene expression in the heart［J］. Nucleic Acids Res， 2014， 42（2）：790-803.

丰嘉欣，郭继深，梁俣，等. Circ_0018478通过编码HERC4-193发挥抑制心肌成纤维细胞纤维化表型的作用［J］.中山大学学报（医学科学版），2022，43（6）：995-1004.

Feng JX， Guo JS， Liang Y， et al. Circ_0018478 inhibits the fibrotic phenotype of cardiac fibroblasts via encoding protein HERC4-193［J］. J Sun Yat⁃sen Univ （Med Sci）， 2022， 43（6）：995-1004.

Rawal S， Nagesh PT， Coffey S， et al. Early dysregulation of cardiac-specific microRNA-208a is linked to maladaptive cardiac remodelling in diabetic myocardium［J］. Cardiovasc Diabetol， 2019 ，18（1）：13.

Stanton AM， Vaduganathan M， Chang LS， et al. Asymptomatic diabetic cardiomyopathy： an underrecognized entity in type 2 diabetes［J］. Curr Diab Rep， 2021， 21（10）：41.

Tian J， Zhao Y， Liu Y， et al. Roles and mechanisms of herbal medicine for diabetic cardiomyopathy： current status and perspective［J］. Oxid Med Cell Longev， 2017， 2017：8214541.

Turner NA. Therapeutic regulation of cardiac fibroblast function： targeting stress-activated protein kinase pathways［J］. Future Cardiol， 2011，7（5）：673-691.

Liu B， Li J， Cairns MJ. Identifying miRNAs， targets and functions［J］. Brief Bioinform， 2014，5（1）：1-19.

Wang BW， Wu GJ， Cheng WP， et al. MicroRNA-208a increases myocardial fibrosis via endoglin in volume overloading heart［J］. PLoS One， 2014， 9（1）：e84188.

Coulis G， Londhe AD， Sagabala RS， et al. Protein tyrosine phosphatase 1B regulates miR-208b-argonaute 2 association and thyroid hormone responsiveness in cardiac hypertrophy［J］. Sci Signal， 2022，15（730）：eabn6875.

Yang J， Yu X， Xue F， et al. Exosomes derived from cardiomyocytes promote cardiac fibrosis via myocyte-fibroblast cross-talk［J］. Am J Transl Res， 2018，10（12）：4350-4366.

Dong C， Nakagawa R， Oyama K， et al. Structural basis for histone variant H3tK27me3 recognition by PHF1 and PHF19［J］. Elife， 2020， 9：e58675.

Zhang Z， Jones A， Sun CW， et al. PRC2 complexes with JARID2， MTF2， and esPRC2p48 in ES cells to modulate ES cell pluripotency and somatic cell reprogramming［J］. Stem Cells， 2011， 29（2）：229-240.

Lee SR， Heo JH， Jo SL， et al. Progesterone receptor membrane component 1 reduces cardiac steatosis and lipotoxicity via activation of fatty acid oxidation and mitochondrial respiration［J］. Sci Rep， 2021，11（1）：8781.

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LI Hui

FANG Xianhong

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Related Institution

Department of Clinical Laboratory

Guangdong Provincial Key Laboratory of Clinical Pharmacology// Guangdong Provincial People’s Hospital// Guangdong Academy of Medical Sciences， Southern Medical University

School of Medicine， South China University of Technology

Department of Pharmacy， The Fifth Affiliated Hospital of Guangzhou Medical University

Guangdong Cardiovascular Institute//Guangdong Provincial People’s Hospital//Guangdong Academy of Medical Sciences， Southern Medical University

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