Effect of miR-134 against myocardial hypoxia/reoxygenation injury by directly targeting NOS3 and regulating PI3K/Akt pathway

Abstract

To reveal the function of miR-134 in myocardial ischemia. Methods: Real-time PCR and western blotting were performed to measure the expression of miR-134, nitric oxide synthase 3 (NOS3) and apoptotic-associated proteins. Lactic dehydrogenase (LDH) assay, cell counting kit-8 (CCK-8), Hoechst 33342/PI double staining and flow cytometry assay were implemented in H9c2 cells, respectively. MiR-134 mimic/inhibitor was used to regulate miR-134 expression. Bioinformatic analysis and luciferase reporter assay were utilized to identify the interrelation between miR-134 and NOS3. Rescue experiments exhibited the role of NOS3. The involvement of PI3K/AKT was assessed by western blot analysis. Results: MiR-134 was high regulated in the myocardial ischemia model, and miR-134 mimic/ inhibitor transfection accelerated/impaired the speed of cell apoptosis and attenuated/exerted the cell proliferative prosperity induced by H/R regulating active status of PI3K/AKT signaling. LDH activity was also changed due to the different treatments. Moreover, miR-134 could target NOS3 directly and simultaneously attenuated the expression of NOS3. Co-transfection miR-134 inhibitor and pcDNA3.1-NOS3 highlighted the inhibitory effects of miR-134 on myocardial H/R injury. Conclusion: This present work puts insights into the crucial effects of the miR-134/NOS3 axis in myocardial H/R injury, delivering a potential therapeutic technology in future.

Pathophysiological implications of hypoxia in human diseases

“…Numerous miRNAs had been reported to be up-or down-regulated in patients with myocardial ischemia/reperfusion injury (Table 1). Among them, MiR-22, miR-134, miR-135a, miR-203, miR-144, miR-98, miR-18a, miR-210, miR-340-5p, miR-374a-5p, and miR-1192 exert protective effects in cardiovascular ischemic injury through downregulating their target genes [37,41,56,77,79,112,160,163,175,183,184]. On the other hand, a specific set of miRNAs has been linked to cardiac dysfunctions in varied cardiac injury models.…”

Regulation of NOS expression in vascular diseases

“…Hypoxia upregulates the expression of miR-134 in rat cardiomyoblast H9c2 cells. As miR-134 directly targets NOS3 mRNA and reduce NOS3 protein expression this post-transcriptional mechanisms seem to be part of the hypoxia related downregulation of NOS3 expression [190].…”

The microRNA miR‐134‐5p induces calcium deposition by inhibiting histone deacetylase 5 in vascular smooth muscle cells

“…56 The function of miR-134 in myocardial infarction is to accelerate myocardial hypoxia/reoxygenation injury by targeting nitric oxide synthase 3, encoding endothelial nitric oxide synthase, which generates nitric oxide and plays a protective role in the cardiovascular system. 57 Xiao et al revealed that inhibition of miR-134 could protect myocardial ischaemia/reperfusion injury through up-regulation of NOS3 and activation of the PI3K/AKT pathway. 57 miR-134 seems to actively participate in the progression of atherosclerosis not only in cardiomyocytes but also by targeting angiopoietin-like 4 in macrophages.…”

“…57 Xiao et al revealed that inhibition of miR-134 could protect myocardial ischaemia/reperfusion injury through up-regulation of NOS3 and activation of the PI3K/AKT pathway. 57 miR-134 seems to actively participate in the progression of atherosclerosis not only in cardiomyocytes but also by targeting angiopoietin-like 4 in macrophages. 58 It is noteworthy that miR-134-5p is up-regulated in the serum of patients with coronary artery calcification and could be used as a biomarker.…”