AimsTransforming growth factor β1 (TGF-β1) is a prosclerotic cytokine involved in cardiac remodelling leading to heart failure (HF). Acetylation/de-acetylation of specific lysine residues in Smad2/3 has been shown to regulate TGF-β signalling by altering its transcriptional activity. Recently, the lysine de-acetylase sirtuin 1 (SIRT1) has been shown to have a cardioprotective effect; however, SIRT1 expression and activity are paradoxically reduced in HF. Herein, we investigate whether pharmacological activation of SIRT1 would induce cardioprotection in a pressure overload model and assess the impact of SIRT1 activation on TGF-β signalling and the fibrotic response.Methods and resultsEight weeks old male C57BL/6 mice were randomized to undergo sham surgery or transverse aortic constriction (TAC) to induce pressure overload. Post-surgery, animals were further randomized to receive SRT1720 or vehicle treatment. Echocardiography, pressure-volume loops, and histological analysis revealed an impairment in cardiac function and deleterious left ventricular remodelling in TAC-operated animals that was improved with SRT1720 treatment. Genetic ablation and cell culture studies using a Smad-binding response element revealed SIRT1 to be a specific target of SRT1720 and identified Smad2/3 as a SIRT1 specific substrate.ConclusionOverall, our data demonstrate that Smad2/3 is a specific SIRT1 target and suggests that pharmacological activation of SIRT1 may be a novel therapeutic strategy to prevent/reverse HF via modifying Smad activity.

Cardiac fibrosis is a common finding in patients with chronic kidney disease. Here, we investigate the cardio-renal effects of theracurmin, a novel formulation of the polyphenolic compound curcumin, in a rat model of chronic kidney disease. Briefly, Sprague-Dawley rats were randomized to undergo sham or subtotal nephrectomy (SNx) surgery. At 3 weeks post surgery, SNx animals were further randomized to received theracurmin via once daily oral gavage or vehicle for 5 consecutive weeks. At 8 weeks post surgery, cardiac function was assessed via echocardiography and pressure volume loop analysis, followed by LV and renal tissue collection for analysis. SNx animals developed key hallmarks of renal injury including hypertension, proteinuria, elevated blood urea nitrogen, and glomerulosclerosis. Renal injury in SNx animals was also associated with significant diastolic dysfunction, macrophage infiltration, and cardiac NLRP3 inflammasome activation. Treatment of SNx animals with theracurmin improved structural and functional manifestations of cardiac injury associated with renal failure and also attenuated cardiac NLRP3 inflammasome activation and mature IL-1β release. Taken together, our findings suggest a significant role for the NLRP3 inflammasome in renal injury-induced cardiac dysfunction and presents inflammasome attenuation as a unique strategy to prevent adverse cardiac remodeling in the setting of chronic kidney disease.

Excessive cardiac interstitial fibrosis impairs normal cardiac function. We have shown that the α11β1 (α11) integrin mediates fibrotic responses to glycated collagen in rat myocardium by a pathway involving transforming growth factor-β. Little is known of the role of the α11 integrin in the developing mammalian heart. Therefore, we examined the impact of deletion of the α11 integrin in wild-type mice and in mice treated with streptozotocin (STZ) to elucidate the role of the α11 integrin in normal cardiac homeostasis and in the pathogenesis of diabetes-related fibrosis. As anticipated, cardiac fibrosis was reduced in α11 integrin knockout mice (α11 −/− ; C57BL/6 background) treated with STZ compared with STZ-treated wild-type mice ( P < 0.05). Unexpectedly, diastolic function was impaired in both vehicle and STZ-treated α11 −/− mice, as shown by the decreased minimum rate of pressure change and prolonged time constant of relaxation in association with increased end-diastolic pressure (all P < 0.05 compared with wild-type mice). Accordingly, we examined the phenotype of untreated α11 −/− mice, which demonstrated a reduced cardiomyocyte cross-sectional cell area and myofibril thickness (all P < 0.05 compared with wild-type mice) and impaired myofibril arrangement. Immunostaining for desmin and connexin 43 showed abnormal intermediate filament organization at intercalated disks and impaired gap-junction development. Overall, deletion of the α11 integrin attenuates cardiac fibrosis in the mammalian mouse heart and reduces ECM formation as a result of diabetes. Furthermore, α11 integrin deletion impairs cardiac function and alters cardiomyocyte morphology. These findings shed further light on the poorly understood interaction between the fibroblast–cardiomyocyte and the ECM.

Background: Theracurmin is a novel formulation of the polyphenolic compound curcumin, which has been shown to have anti-inflammatory properties in a wide range of diseases. In chronic kidney disease (CKD), NLRP3 and inflammasome-dependent cytokines IL-1β and IL-18 have been shown to contribute significantly to disease progression and ensuing cardiovascular complications. Here, we investigate the cardio-renal effects of theracurmin in a rodent model of CKD and assess its impact on NLRP3 inflammasome activation. Methods: Sprague-Dawley rats were randomized to undergo sham or subtotal nephrectomy (SNx) surgery at 8 weeks of age. At 3 weeks post surgery, SNx animals were randomized to received 100mg/kg/day of theracurmin via once daily oral gavage. At 8 weeks post surgery, cardiac function was assessed, followed by LV and renal tissue collection for analysis. Results: Renal dysfunction, evidenced by elevated blood pressure and proteinuria, was confirmed in SNx animals. Treatment with theracurmin had no significant effect on renal function; however, an improvement in systolic blood pressure was observed (p<0.05). Cardiac function of SNx animals was improved with theracurmin treatment. Chamber compliance, as measured by the slope of the end diastolic pressure volume relationship was improved (p<0.05). SNx-induced LV hypertrophy and fibrosis was attenuated with theracurmin treatment (p<0.05). Cardiac NLRP3 inflammasome activation, characterized by an overexpression of NLRP3 inflammasome components in SNx animals, was also attenuated in theracurmin treated animals (p<0.001). Conclusions: Theracurmin may be an effective compound to ameliorate cardiovascular events in the setting of CKD. The noted improvements to cardiac structure and function, albeit in the absence of significant changes to renal function, suggests a cardioprotective effect of the compound and raises the possibility of a dose-specific effect of theracurmin in the setting of CKD.

In addition to degrading glucagon‐like peptide‐1 (GLP‐1), dipeptidyl peptidase‐4 (DPP‐4) inactivates several chemokines, including stromal cell‐derived factor‐1α (SDF‐1α), a pro‐angiogenic and cardiomyocyte protective protein. We hypothesized that DPP‐4 inhibition may confer benefit following myocardial infarction (MI) in the diabetic setting as a consequence of enhanced SDF‐1α availability rather than potentiating GLP‐1. To test this we compared the effects of saxagliptin with those of liraglutide and used the SDF‐1α receptor (CXCR4) antagonist plerixafor.

Background: Transforming growth factor β1 ( TGFβ1) is a prosclerotic cytokine involved in cardiac remodeling leading to heart failure (CHF). In order to precisely regulate TGFβ1 signaling, acetylation/de-acetylation of lysine 19 within the MH1 domain of Smad2 has been shown to alter DNA binding and transcriptional activity. Recently the lysine de-acetylase Sirt1 has been shown to have a cardioprotective effect, however both Sirt1 expression and activity are reduced in CHF. We hypothesized that pharmacological activation of Sirt1 using SRT1720 would induce cardioprotection in a transverse aortic constriction (TAC) model through modulation of TGF-s/Smad signaling. Methods: Wildtype C57BL/6 mice were randomized to receive TAC or sham surgery at 8 weeks of age; each group was subsequently randomized to receive SRT1720 (100mg/kg/day) or placebo, administered once daily via oral gavage. Six weeks following TAC, cardiac function was assessed followed by left ventricular (LV) tissue collection for analysis. Results: Animals randomized to TAC demonstrated reduced systolic function, LV hypertrophy and increased fibrosis (all p Conclusion: Treatment of mice with SRT1720 improves cardiac function and reduces cardiac fibrosis and hypertrophy following TAC. Given that Sirt1 expression and activity are reduced in CHF, enhancing Sirt1 activity may represent a novel intervention to reduce TGF-s mediated fibrosis.

Integrins, transmembrane receptors, have two major functions: 1) cell attachment to the extracellular matrix (ECM) and 2) signal transduction from the ECM to the cell. In the heart, various pathological stimuli induce adverse ECM remodeling, including fibrosis and excess collagen deposition, contributing to heart failure (HF) in the vast majority of patients. Accordingly, collagen fibril-binding integrins expressed by cardiac fibroblasts may contribute to HF. Notably, the α11β1 integrin (α11) is mainly expressed by fibroblasts and binds preferentially to type I collagen fibers. Currently, the role of the α11 in pathological remodeling of the heart is not defined. We examined the association of α11 in human end-stage HF and experimentally-induced HF in mice. In human LV sections obtained from patients with end-stage idiopathic (n = 5) and ischemic (n = 5) HF, qRT-PCR and Western Blot analysis showed varied α11 mRNA and protein expression. α11 expression appeared upregulated and downregulated in ischemic and idiopathic HF relative to control (1.2 ± 0.34 vs. 0.83 ± 0.34, mRNA, 1.4 ± 0.28 vs. 0.87 ± 0.37, protein), respectively, suggesting a unique role in human end-stage HF (Figure 1). We examined α11 in experimental HF. Normal C57 mice were divided into two groups: 1) transverse aortic constriction (TAC; n = 13) and 2) sham (n = 14). At 6 weeks, mice underwent echocardiography and invasive pressure-volume (PV) loop analysis, followed by tissue collection. Six weeks post-operation, TAC animals demonstrated significant structural and functional differences, including LV and myocyte hypertrophy and myocardial fibrosis. Hearts from TAC animals demonstrated upregulated α11 mRNA and protein expression, consistent with the human data (Table 1). To determine the mechanism of α11 regulation, we examined isolated human cardiac fibroblasts. Applied tensile force increased α11 expression, which was regulated by TGF-β/Smad signaling. Analysis of the α11 promoter identified a novel Smad binding element at nt-881 and nt-890. Utilizing a specific inhibitor of Smad3 phosphorylation, SIS3, α11 promoter activity induced by tensile force, was reduced, which blocked the increased expression of α11 (all P < 0.05). α11 expression is increased in both human and experimental HF, and the increased expression is mediated in part by Smad3 signaling. These findings suggest that α11 is involved in a positive feedback loop whereby enhanced ECM production increases α11 expression, which in turn drives further matrix accumulation. This suggests that α11 may be a novel target to reduce cardiac fibrosis in the failing heart.View Large Image Figure ViewerDownload (PPT)

BACKGROUND: Integrins, transmembrane receptors, play crucial roles in diverse cellular and developmental processes due to critical interactions with the extracellular matrix (ECM). During fetal development and towards adulthood, heart growth and function is suggested to depend on forming and remodeling the ECM and its connection to the myocyte. Currently however, the role of integrins in cardiovascular development (CVD) is poorly defined. Thus, we hypothesized that the α11 integrin (α11), which is expressed by fibroblasts and binds preferentially to type I collagen fibers, plays a vital role in CVD. METHODS: α11 KO and wildtype littermate mice (both n = 8) were examined at 4 weeks and 8 weeks of age. Animals underwent function assessments, including echocardiography and invasive pressure volume (PV) loop analysis, and structural examination via histological and electron microscopy (EM) analysis. RESULTS: At 4 weeks, heart weight (HW) and HW indexed to tibial length were decreased in α11 KO mice (P < 0.05), which were normalized at 8 weeks. Echocardiography revealed reduced end-diastolic area (EDA) at 4 weeks (P < 0.05). Despite normalization of EDA at 8 weeks, PV loop revealed impaired diastolic function as evidence by increased EDP, prolonged Tau and steeper EDPVR (all P < 0.05). No differences in HR or systolic parameters were evident. α11 KO mice also demonstrated structural changes. WGA staining revealed evidence of myofibrillar disarray. Connexin 43 and desmin staining showed increased Z-disk and intermediate filament clustering, respectively. LV myocyte size was also reduced (P < 0.05). Similarly, EM analysis showed reduced cardiomyocyte thickness and distance between end plates (both P < 0.05). CONCLUSION: Loss of α11 resulted in progressively worsening diastolic function that was associated with myofibrillar disarray and impaired cardiomyocyte growth. These findings suggest that α11 is required for the development of normal heart structure and function.

The family of Integrins are transmembrane receptors that have two major functions; attachment of the cell to the extracellular matrix (ECM) and signal transduction from the ECM to the cell. As a result they play crucial roles in diverse cellular and developmental processes including cell growth, differentiation and survival. We have previously demonstrated that diabetes induces the cardiac expression of the α11 integrin that, via Smad dependent mechanisms mediates the formation of profibrotic fibroblasts and contributes to the development of a fibrotic interstitium. The exact role of α11 integrin in cardiovascular development however remains obscure. We therefore utilized the α11 integrin KO mouse to identify the role of endogenous α11 integrin in the myocardium.

Energy metabolism is complex, and is broadly categorized into 3 components: substrate utilization, oxidative phosphorylation and ATP transfer and utilization. Whilst an area of intense research, our understanding of energy metabolism in health and disease remains limited, primarily due to inherent difficulties in measuring the various processes involved. With the advent of hyperpolarization using the dynamic nuclear polarization (DNP) technique, magnetic resonance spectroscopy (MRS) utilizing 13C-labelled tracers allows for the non invasive visualization of energy metabolism in the intact organism from mouse to man. We hypothesized that long term diabetes would induce alterations in carbohydrate metabolism in the kidney and heart in a model of spontaneous non obese type 2 diabetes, the Goto Kakisaki (GK) rat.

Pulmonary hypertension (PH) is a rare and incurable disease with a poor prognosis. Characterized by sustained elevation of pulmonary arterial pressure, progressive vascular remodeling, right ventricular hypertrophy (RVH), PH ultimately culminates in right ventricular failure. Current therapies, primarily vasodilators, lack inhibitory effects on vascular remodeling, and are therefore inadequate to prevent disease progression. Hypoxic pulmonary vasoconstriction (HPV) is an early event occurring in PH due to chronic hypoxic exposure. Recent evidence suggests that the small GTPase RhoA and its downstream Rho-associated kinase (ROCK) play an important role in HPV, due to their effects on vasoconstriction and pulmonary cell proliferation.

Di-peptidyl peptidase 4 (DPP-4) is a membrane spanning exopeptidase that cleaves dipeptides from the N terminus of proteins/peptides. Recent studies demonstrate circulating DPP-4 activity is increased in diabetes, which may in part explain the poor prognosis of diabetic patients with ischemic heart disease. In addition to degrading glucagon-like peptide-1 (GLP-1), DPP4 is also known to degrade stromal derived factor-1 (SDF-1), a pro-angiogenic and cardiomyocyte protective protein currently in early phase clinical trials for patients with ischemic heart disease. We hypothesized that DPP-4 inhibition in the diabetic post myocardial infarction (MI) setting may confer a benefit as a result of enhanced SDF-1 availability rather than potentiating GLP-1. We therefore compared DPP-4 inhibition with GLP-1 agonism, and used the specific SDF-1 receptor (CXCR4) antagonist, AMD3100 to demonstrate SDF-1 specificity.