Dysfunction of endothelium-dependent relaxation to insulin via PKC-mediated GRK2/Akt activation in aortas of ob/ob mice

Abstract

In diabetic states, hyperinsulinemia may negatively regulate Akt/endothelial nitric oxide synthase (eNOS) activation. Our main aim was to investigate whether and how insulin might negatively regulate Akt/eNOS activities via G protein-coupled receptor kinase 2 (GRK2) in aortas from ob/ob mice. Endothelium-dependent relaxation was measured in aortic rings from ob/ob mice (a type 2 diabetes model). GRK2, β-arrestin2, and Akt/eNOS signaling-pathway protein levels and activities were mainly assayed by Western blotting. Plasma insulin was significantly elevated in ob/ob mice. Insulin-induced relaxation was significantly decreased in the ob/ob aortas [vs. age-matched control (lean) ones]. The response in ob/ob aortas was enhanced by PKC inhibitor or GRK2 inhibitor. Akt (at Thr(308)) phosphorylation and eNOS (at Ser(1177)) phosphorylation, and also the β-arrestin2 protein level, were markedly decreased in the membrane fraction of insulin-stimulated ob/ob aortas (vs. insulin-stimulated lean ones). These membrane-fraction expressions were enhanced by GRK2 inhibitor and by PKC inhibitor in the ob/ob group but not in the lean group. PKC activity was much greater in ob/ob than in lean aortas. GRK2 protein and activity levels were increased in ob/ob and were greatly reduced by GRK2 inhibitor or PKC inhibitor pretreatment. These results suggest that in the aorta in diabetic mice with hyperinsulinemia an upregulation of GRK2 and a decrease in β-arrestin2 inhibit insulin-induced stimulation of the Akt/eNOS pathway and that GRK2 overactivation may result from an increase in PKC activity.

Tonic Inhibition by G Protein–Coupled Receptor Kinase 2 of Akt/Endothelial Nitric-Oxide Synthase Signaling in Human Vascular Endothelial Cells under Conditions of Hyperglycemia with High Insulin Levels

“…We interpret this finding to indicate that GRK2, which was upregulated in a high glucose/high insulin environment, led to a tonic inhibition of the Akt/eNOS pathway in endothelial cells. This could fully explain the theoretical basis of the beneficial effect of GRK2 inhibitor on impaired endotheliumdependent relaxation to insulin in aorta of insulin-resistant diabetic ob/ob mouse, an effect that has been demonstrated in an earlier report (Taguchi et al, 2011).…”

“…Interestingly, recent reports have demonstrated that vascular levels of ERK1/2 activation are elevated in type 2 diabetic mice (Tian et al, 2011;Choi et al, 2012), leading to vascular endothelial dysfunction. The results of those studies might be attributed to increased expression and activity of vascular GRK2 in the disease state (Taguchi et al, 2011(Taguchi et al, , 2012b.…”

“…GRK2 has been described to be involved in impaired endothelium-dependent relaxations in aorta of the ob/ob mouse (Taguchi et al, 2011(Taguchi et al, , 2012a, a model of severe obesity, insulin resistance, and diabetes caused by leptin deficiency (Chen and Wang, 2005). Despite the degree of obesity and hyperinsulinemia, this type 2 diabetic model displays milder hyperglycemia.…”

Improvement of vascular insulin sensitivity by downregulation of GRK2 mediates exercise-induced alleviation of hypertension in spontaneously hypertensive rats

“…In vitro studies using liver cells and adipocytes show that GRK2 inhibits basal and insulin-stimulated glycogen synthesis and insulin-mediated glucose transport (27,31). Recent data highlight the important role of GRK2 in endothelial dysfunction in obese and diabetic rodent models (28,29). However, it is largely unclear whether GRK2 participates in exercise-associated improvement of vascular insulin sensitivity in hypertension.…”

Examining the Role of Insulin in the Regulation of Cardiovascular Health

“…Insulin-stimulated relaxation is impaired in rodent models of type 2 diabetes or obesity [38,72] and can be enhanced by PKC inhibitors [73].…”