Abstract 13243: Oxidative Stress From Upregulation of Cardiac β3-AR-Activated iNOS Uncoupling Promotes Diabetic Cardiomyopathy in Cynomolgous Monkeys With Naturally-Occurring Type 2 Diabetes

Background: Recent evidence highlights oxidative stress as an important mechanism to cardiac dysfunction in diabetic cardiomyopathy (DM) and altered β 3 -adrenergic receptor (AR)-activated nitric oxide synthase (NOS) pathway contributing to this process. However, the NOS isoforms involved are controversial. The mechanism of how β 3 -AR stimulation impacts ROS, SERCA2a, and cardiac function in diabetes is unclear. We tested the hypothesis that oxidant stress from upregulation of LV β 3 -AR-activated iNOS uncoupling promotes DM. Methods: We compared myocyte β 1 - and β 3 -AR, 3 NOS, peroxynitrite (NT), NADPH and SERCA2a expressions and myocyte functional responses to β- and β 3 -AR stimulation with isoproterenol (ISO,10 -8 M) and BRL-37344 (BRL,10 -8 M), respectively, in the absence and presence of iNOS inhibitor, 1400W (10 -5 M), of 10 male monkeys: 5 normal and 5 with naturally-occurring type 2 diabetes. Myocytes were freshly isolated from LV tissue collected during terminal study. Results: Compared to normal myocytes, DM myocytes had significantly increased protein levels of β 3 -AR (0.27 vs 0.15) and iNOS (0.46 vs 0.23) accompanied with increased oxidative stress indicated by significantly-elevated NT formation, NADPH (P67 -phox , 33% and p22 -phox , 25%) and decreased GTPCH expression (0.48 vs 0.82) and activity. DM myocytes had significantly-decreased protein levels of β 1 -AR (0.38 vs 0.59) and SERCA2a (0.95 vs 1.15). These changes were associated with reduced basal cell contraction of dL/dt max (68.4 vs 121.1 μm/s), relaxation (-63.4 vs -110.2 μm/s), and [Ca 2+ ] iT (0.15 vs 0.27) accompanied by diminished β-AR-stimulated positive inotropic response, but enhanced β 3 -AR-induced negative inotropic response. Only in DM myocytes, pretreatment with 1400W improved basal cell function and augmented ISO-increased dL/dt max (63.2%) and [Ca 2+ ] iT (27.8%), but significantly limited BRL-induced decrease in dL/dt max (-13.8%) and [Ca 2+ ] iT (-10.9%). Conclusions: DM is associated with contrasting changes on myocyte β 1 - and β 3 -AR expression with decreased SERCA2a and increased iNOS. Upregulation of β 3 -AR triggers iNOS uncoupling, leading to oxidative stress, thus promoting intrinsic myocyte dysfunction with impaired [Ca 2+ ] i regulation and reduced β-AR reserve.