Long non-coding ribonucleic acid zinc finger E-box binding homeobox 1 antisense RNA 1 regulates myocardial fibrosis in diabetes through the Hippo–Yes-associated protein signaling pathway

IF 3.2 3区医学

Journal of Diabetes Investigation Pub Date : 2023-06-12 DOI:10.1111/jdi.13989

Jing Wu, Rui Lyu, Shumin Chen, Xiaoguang Wang

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引用次数: 1

Abstract

Aims/Introduction

Fibrosis is the principle reason for heart failure in diabetes. Regarding the involvement of long non-coding ribonucleic acid zinc finger E-box binding homeobox 1 antisense 1 (ZEB1-AS1) in diabetic myocardial fibrosis, we explored its specific mechanism.

Materials and Methods

Human cardiac fibroblasts (HCF) were treated with high glucose (HG) and manipulated with plasmid cloning deoxyribonucleic acid 3.1-ZEB1-AS1/microribonucleic acid (miR)-181c-5p mimic/short hairpin RNA specific to sirtuin 1 (sh-SIRT1). ZEB1-AS1, miR-181c-5p expression patterns, cell viability, collagen I and III, α-smooth muscle actin (α-SMA), fibronectin levels and cell migration were assessed by reverse transcription quantitative polymerase chain reaction, cell counting kit-8, western blot and scratch tests. Nuclear/cytosol fractionation assay verified ZEB1-AS1 subcellular localization. The binding sites between ZEB1-AS1 and miR-181c-5p, and between miR-181c-5p and SIRT1 were predicted and verified by Starbase and dual-luciferase assays. The binding of SIRT1 to Yes-associated protein (YAP) and YAP acetylation levels were detected by co-immunoprecipitation. Diabetic mouse models were established. SIRT1, collagen I, collagen III, α-SMA and fibronectin levels, mouse myocardium morphology and collagen deposition were determined by western blot, and hematoxylin–eosin and Masson trichrome staining.

Results

Zinc finger E-box binding homeobox 1 antisense 1 was repressed in HG-induced HCFs. ZEB1-AS1 overexpression inhibited HG-induced HCF excessive proliferation, migration and fibrosis, and diminished collagen I, collagen III, α-SMA and fibronectin protein levels in cells. miR-181c-5p had targeted binding sites with ZEB1-AS1 and SIRT1. SIRT1 silencing/miR-181c-5p overexpression abrogated ZEB1-AS1-inhibited HG-induced HCF proliferation, migration and fibrosis. ZEB1-AS1 suppressed HG-induced HCF fibrosis through SIRT1-mediated YAP deacetylation. ZEB1-AS1 and SIRT1 were repressed in diabetic mice, and miR-181c-5p was promoted. ZEB1-AS1 overexpression improved myocardial fibrosis in diabetic mice, and reduced collagen I, collagen III, α-SMA and fibronectin protein levels in myocardial tissues.

Conclusion

Long non-coding ribonucleic acid ZEB1-AS1 alleviated myocardial fibrosis through the miR-181c-5p–SIRT1–YAP axis in diabetic mice.

Abstract Image

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长链非编码核糖核酸锌指E-box结合同源盒1反义RNA 1通过希波- yes相关蛋白信号通路调控糖尿病心肌纤维化

目的/介绍纤维化是糖尿病患者心力衰竭的主要原因。关于长链非编码核糖核酸锌指E-box结合同源盒1反义1 (ZEB1-AS1)参与糖尿病心肌纤维化，我们探讨其具体机制。材料和方法用高糖(HG)处理人心脏成纤维细胞(HCF)，用质粒克隆脱氧核糖核酸3.1-ZEB1-AS1/微核糖核酸(miR)-181c-5p模拟物/ sirtuin 1特异性短发夹RNA (sh-SIRT1)进行修饰。通过逆转录定量聚合酶链反应、细胞计数试剂盒-8、western blot和划痕试验评估ZEB1-AS1、miR-181c-5p表达模式、细胞活力、胶原I和III、α-平滑肌肌动蛋白(α-SMA)、纤维连接蛋白水平和细胞迁移。核/细胞质分离实验证实ZEB1-AS1亚细胞定位。通过Starbase和双荧光素酶检测预测并验证ZEB1-AS1与miR-181c-5p之间、miR-181c-5p与SIRT1之间的结合位点。通过共免疫沉淀检测SIRT1与yes相关蛋白(YAP)的结合和YAP乙酰化水平。建立糖尿病小鼠模型。采用western blot、苏木精-伊红、马松三色染色检测小鼠SIRT1、ⅰ型胶原、ⅲ型胶原、α-SMA、纤维连接蛋白水平、心肌形态及胶原沉积。结果锌指E-box结合同源盒1反义1在hg诱导的hcf中受到抑制。ZEB1-AS1过表达抑制hg诱导的HCF过度增殖、迁移和纤维化，降低细胞中I型胶原、III型胶原、α-SMA和纤维连接蛋白水平。miR-181c-5p靶向ZEB1-AS1和SIRT1的结合位点。SIRT1沉默/miR-181c-5p过表达消除了zeb1 - as1抑制hg诱导的HCF增殖、迁移和纤维化。ZEB1-AS1通过sirt1介导的YAP去乙酰化抑制hg诱导的HCF纤维化。在糖尿病小鼠中，ZEB1-AS1和SIRT1被抑制，miR-181c-5p被提升。ZEB1-AS1过表达可改善糖尿病小鼠心肌纤维化，降低心肌组织中I型胶原、III型胶原、α-SMA和纤维连接蛋白水平。结论长链非编码核糖核酸ZEB1-AS1通过miR-181c-5p-SIRT1-YAP轴减轻糖尿病小鼠心肌纤维化。

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来源期刊

Journal of Diabetes Investigation Medicine-Internal Medicine

自引率

9.40%

发文量

218

期刊介绍： Journal of Diabetes Investigation is your core diabetes journal from Asia; the official journal of the Asian Association for the Study of Diabetes (AASD). The journal publishes original research, country reports, commentaries, reviews, mini-reviews, case reports, letters, as well as editorials and news. Embracing clinical and experimental research in diabetes and related areas, the Journal of Diabetes Investigation includes aspects of prevention, treatment, as well as molecular aspects and pathophysiology. Translational research focused on the exchange of ideas between clinicians and researchers is also welcome. Journal of Diabetes Investigation is indexed by Science Citation Index Expanded (SCIE).