Allopurinol reduces cardiac keap1 and activates NrF2/p62 to attenuate diabetic cardiomyopathy

Abstract

Introduction: Allopurinol (ALP), a xanthine oxidase inhibitor, attenuates diabetes-induced oxidative stress and diabetic cardiomyopathy(DCM), but the mechanism is unclear. Activation of Nuclear factor erythroid 2-related factor-2 (Nrf2) following the disassociation with its repressor Keap1 under oxidative stress plays key roles in maintaining inner redox homeostasis and in attenuating DCM, which is associated with attenuation of autophagy and upregulation of SQSTM1/P62. However, the interactions among Keap1, Nrf2 and p62 in DCM and in ALP-mediated attenuation of DCM is unknown. We postulated that suppressing Keap1 and the subsequent activation of Nrf2 and mitigating autophagy over activation may represent major mechanisms whereby ALP attenuates DCM.

Methods and Results: Streptozotocin-induced diabetic (D) rats were untreated or treated with allopurinol (100 mg/kg/day, n=6 per group) for 1 week starting at 4 weeks of diabetes induction, and terminated after measuring left ventricular (LV) function by echocardiography. D rats demonstrated asymptomatic cardiac dysfunction manifested as significant reductions in heart rate(HR), left ventricular eject fraction(LVEF), stroke work(SW) and cardiac output(CO), Left ventricular end-systolic volume (LVVs)) as compared to non-diabetic control (all p<0.05) and ALP improved or normalized HR, LVEF, SW, CO, LVVs in D rats(all p<0.05, vs. diabetic untreated). Biochemically, hearts of D rats displayed excessive oxidative stress manifested as increased levels of 15-F2t-Isoprostane, increased apoptotic cell death and cardiomyocyte autophagy (enhanced LC3 II/I ratio) that were concomitant with increased Nrf2 and Nrf2- antioxidant-response element (ARE) (HO-1 expression, superoxide dysmutase activity and catalase expression) but decreased P62 expression. ALP reverted all the above mentioned diabetes-induced biochemical changes except that it further activated Nrf2 with concomitant inhibition of the Nrf2 repressor Keap1.

Conclusion: Suppressing Keap1 and the subsequent activation of Nrf2 and p62 to mitigate apoptosis and autophagy over activation may represent major mechanisms whereby ALP attenuates DCM and cardiac dysfunction.