Research ArticlemiR-150 regulates high glucose-induced cardiomyocyte hypertrophy by targeting the transcriptional co-activator p300
Introduction
Cardiomyocyte hypertrophy is one of the key structural hallmarks of diabetic cardiomyopathy, which ultimately leads to cardiac failure [1]. The pathogenesis of cardiomyocyte hypertrophy at the transcriptional level involves the epigenetic modification of histones and the activation of multiple transcription factors, eventually resulting in a characteristic reversion to fetal genes expression [2], [3]. Growing evidence suggests that excessive acetylation in cardiomyocyte nuclei, which is regulated by the activity of histone acetyl transferases (HATs) and histone deacetylases (HDACs), plays an important role in coordinating this transcriptional response in cardiac hypertrophy [4], [5]. Transcriptional co-activator p300 is the most widely studied of the HATs due to its role in cardiac hypertrophy. Prohypertrophic stimuli, including hyperglycemia, initiate the expression and activation of the p300 gene [3], [5], [6]. p300, acting as an epigenetic regulator, acetylates histones to loosen the nucleosome, facilitate protein-DNA interactions, and ultimately promote gene transcription [7], [8]. In addition, p300 can directly acetylate or interact with hypertrophy-responsive transcription factors, such as GATA4 and MEF2, to increase their DNA binding and transcriptional activities [3], [5], [9], [10]. The binding of GATA4 to p300 is essential for the transcription of genes encoding ANP, BNP, ET-1 and β-MHC in both phenylephrine (PE)- and aortic banding (AB)-induced cardiac hypertrophy [9], [10]. The MEF2-mediated transcription of fetal cardiac genes in diabetes-induced cardiomyocyte hypertrophy is also p300-dependent [3], [5]. Furthermore, the redox-sensitive transcription factors NF-κB and AP-1, which are involved in diabetes-induced cardiac hypertrophy, are also downstream effectors of p300 [11], [12]. Thus, by regulating both histones and transcription factors, p300 is considered as a master switch to trigger the expression of pivotal regulators of cardiomyocyte hypertrophy. However, the mechanism of upstream regulation of p300 expression in cardiomyocyte hypertrophy is not fully understood, especially with regard to microRNAs (miRNAs), which regulate gene expression post-transcriptionally.
miRNAs represent a family of short, noncoding RNAs that are endogenous regulators of gene expression [13]. Mature miRNA sequences are single-stranded, 18–24 nucleotides in length, and are often highly conserved among species. They mostly interact with the 3′-untranslated region (UTR) of their target mRNA transcripts, leading to mRNA degradation and/or translational repression, and ultimately to the negative regulation of gene expression [14]. A variety of studies indicate that many miRNAs exhibit altered expression profiles in cardiac hypertrophy induced by diabetes or other conditions [15], [16], [17]. However, whether miRNAs regulate the expression of p300 in cardiomyocyte hypertrophy has not been determined.
Many miRNA profiling studies using microarray technology have observed that miR-150 is decreased in cardiac hypertrophy, although its functional role has not been validated [13], [17], [18], [19]. To identify candidate miRNAs that regulate p300, we used open source software (TargetScan, miRanda, PITA) and identified miR-150 as a potential p300 targeting miRNA. Thus, the purpose of this study was to investigate the potential role of miR-150 in the regulation of p300 gene expression and high glucose-induced cardiomyocyte hypertrophy. We first investigated whether alterations in the expression of miR-150 or p300 are involved in cardiac hypertrophy in streptozotocin (STZ)-induced diabetic rats. Neonatal rat cardiomyocytes were then used to explore the relationship between miRNA-150 levels and p300 expression.
Section snippets
Animal studies
Age- and weight-matched male Sprague-Dawley rats (200–250 g body weight; Chongqing Medical University, Chongqing, China) were housed in a climate-controlled room with a 12-hour light/dark cycle and free access to food and water. All procedures were performed according to the National Institute of Health (NIH) guidelines for the care and use of experimental animals (no. 85-23, revised 1996). Diabetes was induced by a single i.v. injection of STZ in citrate buffer (65 mg/kg, pH 5.6). Nondiabetic
High glucose-induced cardiomyocyte hypertrophy is associated with alterations in miR-150 and p300 expression in vivo and in vitro
We first assessed the morphological and biochemical changes that occurred with diabetes-induced cardiomyocyte hypertrophy, as were previously reported [3], [16]. After three months of follow-up, diabetic animals displayed reduced body weight and an increased HW/BW ratio compared with the age- and gender-matched normal control group (Table 1). Histological analysis further revealed that the cardiomyocyte cross-sectional area was increased in the diabetic hearts (Fig. 1A). Moreover, mRNA levels
Discussion
This study demonstrated a novel miRNA-dependent pathway regulating p300 expression and subsequent alterations in glucose-induced cardiomyocyte hypertrophy. We have used bioinformatics to predict and show that miR-150 is a potential p300 targeting miRNA. High glucose levels caused the down-regulation of miR-150 levels and an increase in p300 expression concomitantly with cardiomyocyte hypertrophy both in vivo and in vitro. Luciferase assays confirmed that miR-150 directly interacts with the
Acknowledgments
This work was supported by the National Natural Science Foundation of China (No. 30570877). The authors would like to acknowledge support from the Ophthalmology Laboratory of Chongqing Medical University.
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