Abstract

Optimum level of autophagy is indispensable for survival and cellular function, and miRNA tightly regulates biological functions. In diabetic hearts, miR‐133a (anti‐hypertrophy and anti‐fibrosis) is attenuated and autophagy is induced. Based on these observations, we hypothesize that inhibition of miR‐133a induces autophagy in diabetic hearts. To test the hypothesis, HL1 cardiomyocytes were treated with low (LG‐5mM) and high (HG‐25mM) dose of D‐glucose, and miR‐133a and scrambled control. To determine whether miR‐133a mitigates autophagy in diabetic hearts, we measured the levels of monodansylcadaverin intensity (marker of autophagolysosome) and then the upstream targets ATG3, LC3B (autophagophore formation) and Beclin‐1 (initiator of autophagy signaling). We also determined the activity and expression of mTOR (upstream inhibitor of Beclin‐1) and AMPK (upstream inhibitor of mTOR) by RT‐qPCR, immunoblotting and immunofluorescence. The specific role of miR‐133a in mitigation of high glucose mediated autophagophore formation is determined by transfecting HL1 cardiomyocytes with LC3‐RFP plasmid. In hyperglycemic cardiomyocytes, miR‐133a decreases the protein levels of AMPK (4.1±0.38 fold), that in turn increases mTOR (3.2±0.4 fold), which suppresses Beclin1 (2.5± 0.33 fold) and the downstream targets LC3B (3.5±0.4 fold) and ATG3 (2.5±0.18 fold), and fluorescent intensity of monodansylcadaverin (2.5±0.47 fold). The in vivo studies were performed on C57BL/6J and Ins2+/‐ (spontaneous, chronic T1D) mice. Both groups of mice were treated with miR‐133a mimic (10^5lentivirus, iv,/day for 5 days) and hearts were extracted after 10 days. The in vivo results corroborate that miR‐133a mitigates cardiac autophagy in diabetes. Further investigation revealed that AMPK 3’UTR is a direct target of miR‐133a. These findings demonstrate a novel mechanism of miR‐133a mediated mitigation of autophagy in diabetic hearts.