Mohammed Shakil Ahmed

Myocardial Connective Tissue Growth Factor (CCN2/CTGF) Attenuates Left Ventricular Remodeling after Myocardial Infarction

Aims Myocardial CCN2/CTGF is induced in heart failure of various etiologies. However, its role in the pathophysiology of left ventricular (LV) remodeling after myocardial infarction (MI) remains unresolved. The current study explores the role of CTGF in infarct healing and LV remodeling in an animal model and in patients admitted for acute ST-elevation MI. Methods and Results Transgenic mice with cardiac-restricted overexpression of CTGF (Tg-CTGF) and non-transgenic littermate controls (NLC) were subjected to permanent ligation of the left anterior descending coronary artery. Despite similar infarct size (area of infarction relative to area at risk) 24 hours after ligation of the coronary artery in Tg-CTGF and NLC mice, Tg-CTGF mice disclosed smaller area of scar tissue, smaller increase of cardiac hypertrophy, and less LV dilatation and deterioration of LV function 4 weeks after MI. Tg-CTGF mice also revealed substantially reduced mortality after MI. Remote/peri-infarct tissue of Tg-CTGF mice contained reduced numbers of leucocytes, macrophages, and cells undergoing apoptosis as compared with NLC mice. In a cohort of patients with acute ST-elevation MI (n = 42) admitted to hospital for percutaneous coronary intervention (PCI) serum-CTGF levels (s-CTGF) were monitored and related to infarct size and LV function assessed by cardiac MRI. Increase in s-CTGF levels after MI was associated with reduced infarct size and improved LV ejection fraction one year after MI, as well as attenuated levels of CRP and GDF-15. Conclusion Increased myocardial CTGF activities after MI are associated with attenuation of LV remodeling and improved LV function mediated by attenuation of inflammatory responses and inhibition of apoptosis.

Substrate Specificities of G Protein-Coupled Receptor Kinase-2 and -3 at Cardiac Myocyte Receptors Provide Basis for Distinct Roles in Regulation of Myocardial Function

The closely related G protein-coupled receptor kinases GRK2 and GRK3 are both expressed in cardiac myocytes. Although GRK2 has been extensively investigated in terms of regulation of cardiac β-adrenergic receptors, the substrate specificities of the two GRK isoforms at G protein-coupled receptors (GPCR) are poorly understood. In this study, the substrate specificities of GRK2 and GRK3 at GPCRs that control cardiac myocyte function were determined in fully differentiated adult cardiac myocytes. Concentration-effect relationships of GRK2, GRK3, and their respective competitive inhibitors, GRK2ct and GRK3ct, at endogenous endothelin, α1-adrenergic, and β1-adrenergic receptor-generated responses in cardiac myocytes were achieved by adenovirus gene transduction. GRK3 and GRK3ct were highly potent and efficient at the endothelin receptors (IC50 for GRK3, 5 ± 0.7 pmol/mg of protein; EC50 for GRK3ct, 2 ± 0.2 pmol/mg of protein). The α1-adrenergic receptor was also a preferred substrate of GRK3 (IC50,7 ± 0.4 pmol/mg of protein). GRK2 lacked efficacy at both endothelin and α1-adrenergic receptors despite massive overexpression. On the contrary, both GRK2ct and GRK3ct enhanced β1-adrenergic receptor-induced cAMP production with comparable potencies. However, the potency of GRK3ct at β1-adrenergic receptors was at least 20-fold lower than that at endothelin receptors. In conclusion, this study demonstrates distinct substrate specificities of GRK2 and GRK3 at different GPCRs in fully differentiated adult cardiac myocytes. As inferred from the above findings, GRK2 may play its primary role in regulation of cardiac contractility and chronotropy by controlling β1-adrenergic receptors, whereas GRK3 may play important roles in regulation of cardiac growth and hypertrophy by selectively controlling endothelin and α1-adrenergic receptors.

CCN2/CTGF attenuates myocardial hypertrophy and cardiac dysfunction upon chronic pressure-overload

BACKGROUND Myocardial CCN2/CTGF (connective tissue growth factor) is strongly induced in heart failure (HF) and acts as a cardioprotective factor in ischemia/reperfusion injury. However, its functional role in myocardial hypertrophy remains unresolved. METHODS AND RESULTS Transgenic mice with cardiac-restricted overexpression of CTGF (Tg-CTGF) and non-transgenic littermate control (NLC) mice were subjected to chronic pressure-overload by abdominal aortic banding. After 4weeks of persistent pressure-overload, a time point at which compensatory hypertrophy of the left ventricle (LV) prevails, Tg-CTGF mice displayed diminished increase of LV mass compared with NLC. At study end-point after 12 weeks of sustained aortic constriction, the mice displayed LV dilatation and reduced cardiac function. Repeated transthoracic echocardiography during the 12 weeks of chronic pressure-overload, revealed attenuation of LV dilatation and virtually sustained systolic function in Tg-CTGF mice compared with NLC mice. Also, increase of LV mass was blunted in Tg-CTGF versus NLC mice at study end-point. Consistently, increases of myocardial ANP, BNP and skeletal α-actin mRNA levels were blunted in Tg-CTGF mice subjected to chronic pressure-overload. Furthermore, cardiac myocytes from Tg-CTGF mice displayed increased phospho-NFATc2 levels and attenuated hypertrophic response upon stimulation with α1-adrenoceptor agonist, indicating that CTGF attenuates hypertrophic signaling in cardiac myocytes. Increase of myocardial collagen contents in mice subjected to aortic banding was similar in Tg-CTGF and NLC mice, indicating that CTGF have minimal impact on myocardial collagen deposition. CONCLUSION This study provides novel evidence that CTGF attenuates cardiac hypertrophy upon chronic pressure-overload due to inhibition of signaling mechanisms that promote pathologic myocardial hypertrophy.

Cardiomyocyte-restricted inhibition of G protein-coupled receptor kinase-3 attenuates cardiac dysfunction after chronic pressure overload

Transgenic mice with cardiac-specific expression of a peptide inhibitor of G protein-coupled receptor kinase (GRK)3 [transgenic COOH-terminal GRK3 (GRK3ct) mice] display myocardial hypercontractility without hypertrophy and enhanced α(1)-adrenergic receptor signaling. A role for GRK3 in the pathogenesis of heart failure (HF) has not been investigated, but inhibition of its isozyme, GRK2, has been beneficial in several HF models. Here, we tested whether inhibition of GRK3 modulated evolving cardiac hypertrophy and dysfunction after pressure overload. Weight-matched male GRK3ct transgenic and nontransgenic littermate control (NLC) mice subjected to chronic pressure overload by abdominal aortic banding (AB) were compared with sham-operated (SH) mice. At 6 wk after AB, a significant increase of cardiac mass consistent with induction of hypertrophy was found, but no differences between GRK3ct-AB and NLC-AB mice were discerned. Simultaneous left ventricular (LV) pressure-volume analysis of electrically paced, ex vivo perfused working hearts revealed substantially reduced systolic and diastolic function in NLC-AB mice (n = 7), which was completely preserved in GRK3ct-AB mice (n = 7). An additional cohort was subjected to in vivo cardiac catheterization and LV pressure-volume analysis at 12 wk after AB. NLC-AB mice (n = 11) displayed elevated end-diastolic pressure (8.5 ± 3.1 vs. 2.9 ± 1.2 mmHg, P < 0.05), reduced cardiac output (3,448 ± 323 vs. 4,488 ± 342 μl/min, P < 0.05), and reduced dP/dt(max) and dP/dt(min) (both P < 0.05) compared with GRK3ct-AB mice (n = 16), corroborating the preserved cardiac structure and function observed in GRK3ct-AB hearts assessed ex vivo. Increased cardiac mass and myocardial mRNA expression of β-myosin heavy chain confirmed the similar induction of cardiac hypertrophy in both AB groups, but only NLC-AB hearts displayed significantly elevated mRNA levels of brain natriuretic peptide and myocardial collagen contents as well as reduced β(1)-adrenergic receptor responsiveness to isoproterenol, indicating increased LV wall stress and the transition to HF. Inhibition of cardiac GRK3 in mice does not alter the hypertrophic response but attenuates cardiac dysfunction and HF after chronic pressure overload.

The homeostatic chemokine CCL21 predicts mortality in aortic stenosis patients and modulates left ventricular remodeling.

Background CCL21 acting through CCR7, is termed a homeostatic chemokine. Based on its role in concerting immunological responses and its proposed involvement in tissue remodeling, we hypothesized that this chemokine could play a role in myocardial remodeling during left ventricular (LV) pressure overload. Methods and Results Our main findings were: (i) Serum levels of CCL21 were markedly raised in patients with symptomatic aortic stenosis (AS, n = 136) as compared with healthy controls (n = 20). (ii) A CCL21 level in the highest tertile was independently associated with all-cause mortality in these patients. (iii) Immunostaining suggested the presence of CCR7 on macrophages, endothelial cells and fibroblasts within calcified human aortic valves. (iv). Mice exposed to LV pressure overload showed enhanced myocardial expression of CCL21 and CCR7 mRNA, and increased CCL21 protein levels. (v) CCR7−/− mice subjected to three weeks of LV pressure overload had similar heart weights compared to wild type mice, but increased LV dilatation and reduced wall thickness. Conclusions Our studies, combining experiments in clinical and experimental LV pressure overload, suggest that CCL21/CCR7 interactions might be involved in the response to pressure overload secondary to AS.