Diabetes-Induced Cardiomyocyte Passive Stiffening Is Caused by Impaired Insulin-Dependent Titin Modification and Can Be Modulated by Neuregulin-1.

RATIONALE: Increased titin-dependent cardiomyocyte tension is a hallmark of heart failure with preserved ejection fraction associated with type-2 diabetes mellitus. However, the insulin-related signaling pathways that modify titin-based cardiomyocyte tension, thereby contributing to modulation of diastolic function, are largely unknown. OBJECTIVE: We aimed to determine how impaired insulin signaling affects titin expression and phosphorylation and thus increases passive cardiomyocyte tension, and whether metformin or neuregulin-1 (NRG-1) can correct disturbed titin modifications and increased titin-based stiffness. METHODS AND RESULTS: We used cardiac biopsies from human diabetic (n=23) and nondiabetic patients (n=19), cultured rat cardiomyocytes, left ventricular tissue from apolipoprotein E-deficient mice with streptozotocin-induced diabetes mellitus (n=12-22), and ZSF1 (obese diabetic Zucker fatty/spontaneously hypertensive heart failure F1 hybrid) rats (n=5-6) and analyzed insulin-dependent signaling pathways that modulate titin phosphorylation. Titin-based passive tension was measured using permeabilized cardiomyocytes. In human diabetic hearts, we detected titin hypophosphorylation at S4099 and hyperphosphorylation at S11878, suggesting altered activity of protein kinases; cardiomyocyte passive tension was significantly increased. When applied to cultured cardiomyocytes, insulin and metformin increased titin phosphorylation at S4010, S4099, and S11878 via enhanced ERK1/2 (extracellular signal regulated kinase 1/2) and PKCα (protein kinase Cα) activity; NRG-1 application enhanced ERK1/2 activity but reduced PKCα activity. In apolipoprotein E-deficient mice, chronic treatment of streptozotocin-induced diabetes mellitus with NRG-1 corrected titin phosphorylation via increased PKG (protein kinase G) and ERK1/2 activity and reduced PKCα activity, which reversed the diabetes mellitus-associated changes in titin-based passive tension. Acute application of NRG-1 to obese ZSF1 rats with type-2 diabetes mellitus reduced end-diastolic pressure. CONCLUSIONS: Mechanistically, we found that impaired cGMP-PKG signaling and elevated PKCα activity are key modulators of titin-based cardiomyocyte stiffening in diabetic hearts. We conclude that by restoring normal kinase activities of PKG, ERK1/2, and PKCα, and by reducing cardiomyocyte passive tension, chronic NRG-1 application is a promising approach to modulate titin properties in heart failure with preserved ejection fraction associated with type-2 diabetes mellitus.

Inhibitory actions of the NRG-1/ErbB4 pathway in macrophages during tissue fibrosis in the heart, skin, and lung.

The neuregulin-1 (NRG-1)/receptor tyrosine-protein kinase erbB (ErbB) system is an endothelium-controlled paracrine system modulating cardiac performance and adaptation. Recent studies have indicated that NRG-1 has antifibrotic effects in the left ventricle, which were explained by direct actions on cardiac fibroblasts. However, the NRG-1/ErbB system also regulates the function of macrophages. In this study, we hypothesized that the antifibrotic effect of NRG-1 in the heart is at least partially mediated through inhibitory effects on macrophages. We also hypothesized that the antifibrotic effect of NRG-1 may be active in other organs, such as the skin and lung. First, in a mouse model of angiotensin II (ANG II)-induced myocardial hypertrophy and fibrosis, NRG-1 treatment (20 µg·kg-1·day-1 ip) significantly attenuated myocardial hypertrophy and fibrosis and improved passive ventricular stiffness (4 wk). Interestingly, 1 wk after exposure to ANG II, NRG-1 already attenuated myocardial macrophage infiltration and cytokine expression. Furthermore, mice with myeloid-specific deletion of the ErbB4 gene (ErbB4F/FLysM-Cre+/-) showed an intensified myocardial fibrotic response to ANG II. Consistently, NRG-1 activated the ErbB4 receptor in isolated macrophages, inhibited phosphatidylinositide 3-kinase/Akt and STAT3 signaling pathways, and reduced the release of inflammatory cytokines. Further experiments showed that the antifibrotic and anti-inflammatory effects of NRG-1 were reproducible in mouse models of bleomycin-induced dermal and pulmonary fibrosis. Overall, this study demonstrates that the antifibrotic effect of NRG-1 in the heart is linked to anti-inflammatory activity NRG-1/ErbB4 signaling in macrophages. Second, this study shows that NRG-1 has antifibrotic and anti-inflammatory effects in organs other than the heart, such as the skin and lung.NEW & NOTEWORTHY Our study contributes to the understanding of the antifibrotic effect of neuregulin-1 during myocardial remodeling. Here, we show that the antifibrotic effect of neuregulin-1 is at least partially mediated through anti-inflammatory activity, linked to receptor tyrosine-protein kinase erbB-4 activation in macrophages. Furthermore, we show that this effect is also present outside the heart.

Neuregulin-1 attenuates development of nephropathy in a type 1 diabetes mouse model with high cardiovascular risk.

Neuregulin-1 (NRG-1) is an endothelium-derived growth factor with cardioprotective and antiatherosclerotic properties and is currently being tested in clinical trials as a treatment for systolic heart failure. In clinical practice, heart failure often coexists with renal failure, sharing an overlapping pathophysiological background. In this study, we hypothesized that NRG-1 might protect against cardiomyopathy, atherosclerosis, and nephropathy within one disease process. We tested this hypothesis in a hypercholesterolemic apolipoprotein E-deficient (apoE(-/-)) type 1 diabetes mouse model prone to the development of cardiomyopathy, atherosclerosis, and nephropathy and compared the effects of NRG-1 with insulin. Upon onset of hyperglycemia induced by streptozotocin, apoE(-/-)mice were treated with vehicle, insulin, or recombinant human (rh)NRG-1 for 14 wk and were compared with nondiabetic apoE(-/-)littermates. Vehicle-treated diabetic apoE(-/-)mice developed left ventricular (LV) dilatation and dysfunction, dense atherosclerotic plaques, and signs of nephropathy. Nephropathy was characterized by abnormalities including hyperfiltration, albuminuria, increased urinary neutrophil gelatinase-associated lipocalin (NGAL), upregulation of renal fibrotic markers, and glomerulosclerosis. rhNRG-1 treatment induced systemic activation of ErbB2 and ErbB4 receptors in both heart and kidneys and prevented LV dilatation, improved LV contractile function, and reduced atherosclerotic plaque size. rhNRG-1 also significantly reduced albuminuria, NGALuria, glomerular fibrosis, and expression of fibrotic markers. Regarding the renal effects of rhNRG-1, further analysis showed that rhNRG-1 inhibited collagen synthesis of glomerular mesangial cells in vitro but did not affect AngII-induced vasoconstriction of glomerular arterioles. In conclusion, systemic administration of rhNRG-1 in hypercholesterolemic type 1 diabetic mice simultaneously protects against complications in the heart, arteries and kidneys.

Left ventricular diastolic dysfunction and myocardial stiffness in diabetic mice is attenuated by inhibition of dipeptidyl peptidase 4.

AIMS: Obesity and Type 2 diabetes mellitus (DM) induce left ventricular (LV) diastolic dysfunction, which contributes to an increasing prevalence of heart failure with a preserved LV ejection fraction. We investigated the effects of sitagliptin (SITA), an inhibitor of dipeptidylpeptidase-4 (DPP-4) and anti-diabetic drug, on LV structure and function of obese mice with Type 2 DM. METHODS AND RESULTS: Obese Type 2 diabetic mice (Lepr(db/db), BKS.Cg-Dock7(m)+/+ Lepr(db)/J), displaying increased cardiomyocyte and LV stiffness at the age of 16 weeks, were treated with SITA (300 mg/kg/day) or vehicle for 8 weeks. SITA severely impaired serum DPP-4 activity, but had no effect on glycaemia. Invasive haemodynamic recordings showed that SITA reduced LV passive stiffness and increased LV stroke volume; LV end-systolic elastance remained unchanged. In addition, SITA reduced resting tension of isolated single cardiomyocytes and intensified phosphorylation of the sarcomeric protein titin. SITA also increased LV concentrations of cGMP and increased activity of protein kinase G (PKG). In vitro activation of PKG decreased resting tension of cardiomyocytes from vehicle-treated mice, but had no effect on resting tension of cardiomyocytes from SITA-treated mice. CONCLUSIONS: In obese Type 2 diabetic mice, in the absence of hypoglycaemic effects, inhibition of DPP-4 decreases LV passive stiffness and improves global LV performance. These effects seem at least partially mediated by stimulatory effects on the myocardial cGMP-PKG pathway and, hence, on the phosphorylation status of titin and the hereto coupled cardiomyocyte stiffness modulus.