Cardiac miR-17-92 protects cardiomyocytes against simulated ischemia/reoxygenation injury under hyperglycemic condition via suppressing sphingolipid biosynthesis

Background: Dysregulation of microRNAs (miRNAs), including the miR-17-92 cluster, has been implicated in the pathogenesis of heart disease and related conditions, such as diabetes, hypertension, and ischemia/reperfusion (I/R) injury. Aberrancies in bioactive sphingolipid metabolism, specifically ceramide (CER), contribute to diabetic cardiomyopathy, where induction of ceramide synthases (CerS) predicts elevated risk of cardiovascular complications. This study investigates whether miR-17-92 can mitigate cardiomyocyte death following simulated ischemia/reoxygenation (SI/RO) under diabetic condition by targeting CerS. Methods & Results: Isolated cardiomyocytes from adult tamoxifen-inducible cardiac-specific miR-17-92 knock-in (KI) and knock-out (KO) male mice were subjected to SI for 40 minutes and RO for 1 hour with high glucose (25 mM). Cardiomyocytes from miR-17-92 KO mice exhibited significantly higher cell death (assessed by trypan blue staining) and apoptosis (assessed by TUNEL staining) post-SI/RO compared to wild type, while cell death was markedly reduced in miR-17-92 KI cardiomyocytes under hyperglycemic conditions. Human cardiomyocyte cells, AC16, were transfected with miR-20a mimic (a member of the miR-17-92 cluster) and incubated with high glucose (25 mM). After 24 hours, cells were exposed to SI (4 hours) followed by 24 hours of RO in high glucose conditions. Overexpression of miR-20a (confirmed by Real-time PCR) protected AC16 cardiomyocytes from SI/RO injury in both normal and hyperglycemic conditions. Western blot analysis and Real-time PCR confirmed that overexpression of miR-20a in AC-16 cells reduced expression of sphingolipid biosynthesis enzymes, ceramide synthase 6 (CerS6) and sphingomyelin synthase 2 (SMS2) after SI/RO under hyperglycemic condition. Conclusion: This study demonstrated that overexpression of miR-20, a member of miR-17-92 cluster, protects cardiomyocytes from ischemia/reperfusion injury by downregulating CerS6 and SMS2, thereby reducing lipotoxicity and pro-apoptotic signaling. These findings suggest that miR-17-92 as a potential therapeutic target for mitigating myocardial infarction in diabetic patients. This work was supported in part by grants from the National Institutes of Health RO1HL134366 (A.D.) and RO1HL158951 (A.D.) This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.