Stretch-induced hypertrophy of isolated adult rabbit cardiomyocytes.

Both mechanical and humoral triggers have been put forward to explain the hypertrophic response of the challenged cardiomyocyte. The aim of the present study was to investigate whether cyclic equibiaxial stretch is a direct stimulus for isolated adult rabbit cardiomyocytes to develop hypertrophy and to explore the potential involvement of the autocrine/paracrine factors ANG II, transforming growth factor (TGF)-beta(1), and IGF-I in this process. Isolated cardiomyocytes were exposed to 10% cyclic equibiaxial stretch (1 Hz) for up to 48 h or treated with ANG II (100 nM), TGF-beta(1) (5 ng/ml), IGF-I (100 ng/ml), ANG II type 1 (AT(1)) receptor blockers, or conditioned medium of stretched fibroblasts. Cyclic stretch significantly increased cell surface area (+3.1%), protein synthesis (+21%), and brain natriuretic peptide (BNP) mRNA expression (6-fold) in cardiomyocytes. TGF-beta(1) expression increased (+42%) transiently at 4 h, whereas cardiomyocyte IGF-I expression was not detectable under all experimental conditions. The AT(1) receptor blockers candesartan and irbesartan (100 nM) did not prevent the stretch-induced hypertrophic response. Direct exposure to ANG II, TGF-beta(1), or IGF-I did not enhance cardiomyocyte BNP expression. In cardiac fibroblasts, stretch elicited a significant approximately twofold increase in TGF-beta(1) and IGF-I expression. Conditioned medium of stretched fibroblasts increased BNP expression in cardiomyocytes ( approximately 2-fold, P = 0.07). This study clearly indicates that cyclic stretch is a strong, direct trigger to induce hypertrophy in fully differentiated rabbit cardiomyocytes. The present findings do not support the notion that stretch-mediated hypertrophy of adult rabbit cardiomyocytes involves autocrine/paracrine actions of ANG II, TGF-beta(1), or IGF-I.

Re-expression of alpha skeletal actin as a marker for dedifferentiation in cardiac pathologies.

Differentiation of foetal cardiomyocytes is accompanied by sequential actin isoform expression, i.e. down-regulation of the 'embryonic' alpha smooth muscle actin, followed by an up-regulation of alpha skeletal actin (alphaSKA) and a final predominant expression of alpha cardiac actin (alphaCA). Our objective was to detect whether re-expression of alphaSKA occurred during cardiomyocyte dedifferentiation, a phenomenon that has been observed in different pathologies characterized by myocardial dysfunction. Immunohistochemistry of alphaCA, alphaSKA and cardiotin was performed on left ventricle biopsies from human patients after coronary bypass surgery. Furthermore, actin isoform expression was investigated in left ventricle samples of rabbit hearts suffering from pressure- and volume-overload and in adult rabbit ventricular cardiomyocytes during dedifferentiation in vitro. Atrial goat samples up to 16 weeks of sustained atrial fibrillation (AF) were studied ultrastructurally and were immunostained for alphaCA and alphaSKA. Up-regulation of alphaSKA was observed in human ventricular cardiomyocytes showing down-regulation of alphaCA and cardiotin. A patchy re-expression pattern of alphaSKA was observed in rabbit left ventricular tissue subjected to pressure- and volume-overload. Dedifferentiating cardiomyocytes in vitro revealed a degradation of the contractile apparatus and local re-expression of alphaSKA. Comparable alphaSKA staining patterns were found in several areas of atrial goat tissue during 16 weeks of AF together with a progressive glycogen accumulation at the same time intervals. The expression of alphaSKA in adult dedifferentiating cardiomyocytes, in combination with PAS-positive glycogen and decreased cardiotin expression, offers an additional tool in the evaluation of myocardial dysfunction and indicates major changes in the contractile properties of these cells.

Pacing-induced dys-synchrony preconditions rabbit myocardium against ischemia/reperfusion injury.

BACKGROUND: Because increased mechanical load induces preconditioning (PC) and dys-synchrony increases loading in late-activated regions, we investigated whether dys-synchrony induced by ventricular pacing (VP) at normal heart rate leads to cardioprotection. METHODS AND RESULTS: Isolated working rabbit hearts were subjected to 35 minutes of global ischemia and 2 hours of reperfusion. Seven hearts underwent VP PC (3 periods of 5 minutes VP at the posterior left ventricular [LV] wall), 7 hearts underwent ischemic preconditioning (IPC) (3 periods of 5 minutes of global ischemia), and 9 hearts served as control (C). LV pressure and sonomicrometry were used to assess global hemodynamics and segment work (SW) and end-diastolic segment length (EDSL) in anterior and posterior LV myocardium. Myocardial release of lactate and expression of proBNP mRNA were determined to gain insight in molecular processes involved in VP PC (*P<0.05). Infarct size (triphenyl tetrazolium chloride staining) was 18.3+/-13.0% in group C, and was uniformly reduced in the VP PC and IPC groups (1.8+/-0.8%*, and 3.5+/-3.1%*, respectively; and not significant between VP PC and IPC). LV posterior wall pacing (VP PC group) increased EDSL (by 6.3+/-5.8%*) and SW (to 335+/-207%*) in the LV anterior wall, whereas posterior wall SW decreased to negative values (-23+/-63%*). LV pacing did not significantly change lactate release and coronary flow but significantly increased proBNP mRNA expression in both anterior and posterior myocardium as compared with controls. CONCLUSIONS: Intermittent dys-synchrony is equally cardioprotective as "classical" IPC. Stretch-mediated signaling is a more likely trigger for VP PC than ischemia. VP PC is potentially applicable in cardiac surgery.

Stretch-induced upregulation of connective tissue growth factor in rabbit cardiomyocytes.

Connective Tissue Growth Factor (CTGF, CCN2) is considered to play an important role in cardiac remodelling. We studied whether stretch is a primary stimulus to induce CTGF expression in vivo in rabbit heart, and in vitro in isolated cardiomyocytes and fibroblasts. Twenty weeks of combined volume and pressure overload resulted in eccentric left ventricular (LV) hypertrophy, with increased LV internal diameter (+36 %) and LV weight (+53 %). Myocardial CTGF mRNA and protein levels were substantially increased in the overloaded animals. In isolated adult rabbit cardiomyocytes, cyclic stretch strongly induced CTGF mRNA expression (2.9-fold at 48 h), whereas in cardiac fibroblasts CTGF-induction was transient and modest (1.4-fold after 4 h). Conditioned medium from stretched fibroblasts induced CTGF mRNA expression in non-stretched cardiomyocytes (2.3-fold at 48 h). Our findings indicate that stretch is an important primary trigger for CTGF-induction in the overloaded heart.