Catalpol attenuates cardiomyocyte apoptosis in diabetic cardiomyopathy via Neat1/miR-140–5p/HDAC4 axis

doi:10.1016/j.biochi.2019.05.005

Biochimie

Volume 165, October 2019, Pages 90-99

https://doi.org/10.1016/j.biochi.2019.05.005 Get rights and content

Highlights

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Catalpol attenuated HG-induced NEAT1 expression in mouse cardiomyocytes.
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NEAT1 could act as an endogenous sponge of miR-140–5p to positively regulate HDAC4 expression.
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Catalpol alleviated myocardial damage via NEAT1/miR-140–5p/HDAC4 axis.

Abstract

The cardioprotection of catalpol and its mechanism in diabetic cardiomyopathy (DCM) remains unclear. Here, mouse cardiomyocytes were treated with high glucose (HG) to establish a model of cellular injury induced by HG. In vitro experiments were carried out and confirmed that Catalpol attenuated HG-induced long noncoding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (Neat1) expression in mouse cardiomyocytes. Mechanistically, luciferase reporter analysis indicated that Neat1 could decrease the transcription of miR-140–5p to positively regulate histone deacetylase 4 (HDAC4) expression. Notably, overexpression of miR-140–5p or silencing of HDAC4 rescued Neat1-induced cardiomyocyte apoptosis. DCM was induced in male C57BL/6 mice by intraperitoneal injection with streptozotocin (STZ) combined with a high-fat/high-sugar diet. Further in vivo experiments identified that Catalpol alleviated myocardial damage by regulating Neat1/miR-140–5p/HDAC4 axis in DCM mice. Thus, our results demonstrated that Catalpol could exert cardioprotective effect against DCM via Neat1/miR-140–5p/HDAC4 pathway.

Introduction

As the one of the most frequent metabolic disorder caused by multiple factors, diabetes mellitus (DM) is a chronic metabolic syndrome with high incidences worldwide and is one of the most serious public health problems of the 21st century [1]. With the hallmark of chronic hyperglycemia attributing to the defects in insulin secretion, action or both, DM will implicate the function of multi-organs, such as kidney, blood vessels, nervous system and heart [2], which causes great pain for the DM patients and their family due to the complications of DM. Diabetic cardiomyopathy (DCM) is one of the most vital DM complications characterized by myocardial hypertrophy, focal necrosis, extracellular matrix deposition, cardiomyocyte apoptosis and interstitial fibrosis, which is also the leading cause of heart failure [3]. The development of DCM presents with highly complex pathogenesis attributing to implicating multiple interconnected mechanisms resulting in cellular damage, even and apoptosis in the cardiac muscles [4]. Hence, it is essential to explore the potential mechanism underlying cardiomyocyte apoptosis to bring the novel insight for the clinical therapy of DCM.

Catalpol is an iridoid glycoside extracted from the roots of Rehmannia glutinosa, which has been previously reported to exert antihyperglycemic effect by enhancing the synthesis of hepatic glycogen and inhibiting glycosylation [5]. Emerging evidence highlighted the anti-diabetic effect of Catalpol in streptozotocin (STZ)-induced rat diabetes [6]. However, the role and molecular mechanism of catalpol on myocardial cell apoptosis in DCM are still unclear. As a class of non-coding RNA without the capacity of coding protein, long non-coding RNA (lncRNA) is longer than 200 nts. Several lines of evidence suggest that several lncRNAs were involved in the development of DCM [7]. LncRNA nuclear paraspeckle assembly transcript 1 (Neat1) located at 11q13.1 locus is implicated in the mechanisms of diverse diseases [8]. Recently, Ma et al. reported that highly expressed Neat1 in ischemia reperfusion (I/R) -treated diabetic rat myocardial tissues promoted myocardial apoptosis and autophagy to aggravate myocardial I/R injury [9]. However, the underlying mechanism of Neat1 involved in DCM remains unclear. Gernapudi et al. identified a miR-140 binding site in Neat1 and found that miR-140 could physically bind to Neat1 in the nucleus, leading to increased Neat1 expression in adipocyte-derived stem cells (ADSCs) [10]. Furthermore, Yang et al. demonstrated that miR-140 was significantly downregulated in adult mouse myocardium with I/R injury. Overexpression of miR-140 could improved myocardial infarction and apoptosis in mice [11]. Hence, we hypothesized that miR-140 participated in the underlying mechanism of Neat1 in DCM.

Histone deacetylase 4 (HDAC4), a member of the Class IIa HDACs family, is highly expressed in heart, brain, skeletal muscle and thymus [12]. A large quantity of evidence has revealed that HDAC4 promoted cardiac regeneration and improved the restoration of cardiac function [13,14]. Moreover, miR-140–5p was predicted to have the binding site with HDAC4-3′UTR. Therefore, we speculated that the cardioprotective mechanism of Catalpol in DCM might involve the regulatory association among Neat1, miR-140–5p and HDAC4.

Section snippets

Isolation and culture of mouse cardiomyocytes

Myocardial tissues were separated from newborn C57BL/6J mice obtained from Center for Animal Experiment of First Affiliated Hospital, Heilongjiang University of Chinese Medicine, minced and transferred into a conical tube containing HEPES-buffered saline solution at 4 °C overnight. The suspension was digested in 0.15% collagenase/dispase mixture (Roche Applied Science, Mannheim, Germany) for 20 min at 37 °C followed by centrifugation at 1000 rpm for 5min. Dulbecco's modified Eagle medium (DMEM;

Catalpol suppressed HG-induced apoptosis in cardiomyocytes

To the best of our knowledge, HG-induced myocardial apoptosis is the major cause of DCM [4]. To verify effects of Catalpol on HG-induced apoptosis of cardiomyocytes, mouse cardiomyocytes were treated with NG, HM as an osmotic control, HG or HG plus different concentrations of Catalpol. Apoptosis was determined by TUNEL staining. As shown in Fig. 1A, the apoptotic index was found to be higher in the HG group, while Catalpol significantly reduced the HG-induced apoptosis in a dose-dependent

Discussion

Our study was the first to investigate the involvement of Neat1/miR-140–5p/HDAC4 axis in DCM progression. The present study generated the major findings as following: 1) Catalpol ameliorated cardiomyocyte apoptosis induced by HG, which was abrogated by Neat1 overexpression, indicating the important role of Neat1 in cardioprotective effect of Catalpol; 2) Neat1 could function as a ceRNA to upregulate HDAC4 expression by downregulating the transcription of miR-140–5p, which consequently resulted

Conclusion

In conclusion, we initiated an exploratory analysis on the expression of lncRNA Neat1 in process of cardioprotective effect of Catalpol in DCM and determined its downstream mechanism of miR-140–5p/HDAC4.

Conflict of interest

The authors declare no conflict of interest.

Acknowledgements

This study was funded by Supporting Plan for Excellent Innovative Talents of Heilongjiang University of Traditional Chinese Medicine; Heilongjiang Post-doctoral Research Initiation Fund; Two-class and Superiority Discipline Construction Project (051022).

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Research paperCatalpol attenuates cardiomyocyte apoptosis in diabetic cardiomyopathy via Neat1/miR-140–5p/HDAC4 axis

Highlights

Abstract

Introduction

Section snippets

Isolation and culture of mouse cardiomyocytes

Catalpol suppressed HG-induced apoptosis in cardiomyocytes

Discussion

Conclusion

Conflict of interest

Acknowledgements

Diabetes & endocrinology

Chin. Med.

Brain Res.

Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie

Mol. Cell

Mol. Cell

Diabetes mellitus statistics on prevalence and mortality: facts and fallacies

Nat. Rev. Endocrinol.

Diabetic cardiomyopathy: a hyperglycaemia- and insulin-resistance-induced heart disease

Diabetologia

Diabetic cardiomyopathy: understanding the molecular and cellular basis to progress in diagnosis and treatment

Heart Fail. Rev.

Plasma glucose lowering mechanisms of catalpol, an active principle from roots of Rehmannia glutinosa, in streptozotocin-induced diabetic rats

J. Agric. Food Chem.

lncRNA H19/miR-675 axis regulates cardiomyocyte apoptosis by targeting VDAC1 in diabetic cardiomyopathy

Sci. Rep.

Research paper
Catalpol attenuates cardiomyocyte apoptosis in diabetic cardiomyopathy via Neat1/miR-140–5p/HDAC4 axis