Overexpression of miR-142-3p improves mitochondrial function in cardiac hypertrophy

Our previous studies have shown that Src homology 2 (SH2) B adaptor protein 1 (SH2B1) plays an important role in cardiac hypertrophy, but the specific mechanism remains to be studied. Through bioinformatics and related research, it is found that miR-14 2-3 p is closely related to SH2B1. Exploring the relationship between miR-14 2-3 p and gene SH2B1 expression is beneficial for the treatment of cardiac hypertrophy. SH2B1 is a key factor regulating energy metabolism, mitochondria are the main organelles of energy metabolism and cardiac hypertrophy are closely related to mitochondrial dysfunction. So it is particularly important to explore the relationship between miR-14 2-3 p and SH2B1 and myocardial mitochondrial function. In this study, we investigated whether overexpression of miR-14 2-3 p can inhibit the expression of gene SH2B1, ameliorate cardiac mitochondrial dysfunction and cardiac hypertrophy.We first constructed a pressure overload myocardial hypertrophy model by ligation of the abdominal aorta(AB) of rats. After 4 weeks of modeling, echocardiographic examination showed that the heart volume of the model group became larger, and Hematoxylin and Eosin Staining Kit (HE) staining showed that the cross-sectional area of the heart tissue became larger. The expression of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), β-Myosin Heavy Chain (β-MHC) messenger RNA (mRNA) increased by real‑time polymerase chain reaction (PCR), which proved that the model of cardiac hypertrophy was successfully constructed. Then, miR-14 2-3 p agomir was injected into the tail vein of rats 2 weeks and 4 weeks respectively. The expression of miR-4 2-3 p mRNA was increased by PCR, suggesting that the miR-14 2-3 p plasmid was successfully transfected. At 4 weeks of pressure overload myocardial hypertrophy model, echocardiography was used to detect cardiac function. HE staining of heart tissue and the expression of ANP, BNP, β-MHC mRNA were used to detect cardiac hypertrophy. Flow cytometry was used to detect changes in mitochondrial membrane potential. Secondly, we observed the effect of miR-14 2-3 p on cardiomyocyte hypertrophy and mitochondrial function in vitro by culture neonatal rat cardiomyocytes. Afterwards, using angiotensin (Ang)II-, miRNA mimic- and miRNA mimic nc- treated cardiomyocytes for a given time. α-actin staining found that the myocardial cells became larger, The expression of ANP, BNP, β-MHC mRNA increased by PCR, which proved that AngII-induced cardiac hypertrophy was successfully constructed. Then, the mitochondrial density was measured using mitochondrial Mito-Red staining by Confocal microscope, the mitochondrial membrane potential was evaluated using flow cytometry, Mitochondrial respiration oxygen consumption rate (OCR) was measured by a Seahorse Extracellular Flux Analyzer XF96, and the expression levels of miR-14 2-3 p, ANP, BNP, β-MHC mRNA, SH2B1 in the cardiomyocytes of different groups were measured by RT-PCR and Western blotting. Finally, we used luciferase assay and transfected miR-14 2-3 p agomir in rats, transfected miR-14 2-3 p mimic in Cardiomyocytes, it is found that myocardial SH2B1 mRNA and protein expression both were reduced.When the pressure overload myocardial hypertrophy model was constructed for four weeks, echocardiography revealed that the heart volume, Left ventricular end diastolic diameter(LVIDd), Left ventricular end systolic diameter (LVIDs), Left ventricular posterior wall thickness (LVPWd), Systolic left ventricular posterior wall (LVPWs), Left ventricle (LV) Mass increased, Ejection fraction (EF) % decreased of AB group increased, but transfected with miR-14 2-3 p agomir of AB, these increase was not significant, EF% reduction was not obvious. HE staining showed that the myocardial cross-sectional area of AB group increased significantly, but the miR-14 2-3 p agomir treatment of AB group did not increase significantly. PCR analysis showed that the expression of ANP, BNP,β-MHC mRNA was significantly increased in AB group, but the miR-14 2-3 p agomir treatment of AB group was not significantly increased. Flow cytometry showed that the mitochondrial membrane potential of AB group was significantly reduced, and the miR-14 2-3 p agomir treatment of AB group was not significantly decreased. During AngII-induced cardiomyocyte hypertrophy, ANP, BNP,β-MHC mRNA expression was increased, while these factors was not significantly increased in miR-14 2-3 p mimic treatment group; mitochondrial membrane potential, mitochondrial density and OCR was significantly decreased in AngII treated group, and these were not significantly reduced in miR-14 2-3 p mimic treatment group; CONCLUSIONS: miR-14 2-3 p not only mitigate cardiac hypertrophy by directly inhibit the expression of gene SH2B1, but also can protect mitochondrial function in cardiac hypertrophy of vitro and vivo.