Function of the sarcoplasmic reticulum and expression of its Ca2(+)-ATPase gene in pressure overload-induced cardiac hypertrophy in the rat (original) (raw)
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Japanese Circulation Journal, 2001
The cellular mechanisms of abnormal calcium regulation and excitation -contraction coupling in relation to glucose metabolism in the hypertrophied heart are not well understood. The present study evaluated the myocardial mechanics of 6-7-week-old pressure overload hypertrophied rabbit hearts in response to dobutamine by (1) serial echocardiograms in vivo and (2) isolated Langendorff perfusion. Cytosolic Ca 2+ ([Ca 2+ ]i) and sarcoplasmic reticulum Ca 2+ -ATPase (SERCA2) expression were measured by fluorescence spectroscopy and Western immunoblotting, respectively. The effect of glycolytic inhibition by 2-deoxy-D-glucose ± pyruvate was also evaluated. Both systolic and diastolic [Ca 2+ ]i tended to be higher and diastolic calcium removal ( Ca) significantly slower in the hypertrophied heart. The myocardial response to dobutamine was blunted and dobutamine insignificantly improved Ca. The SERCA2 protein level was higher in early hypertrophy, but was significantly reduced by 6 weeks of age, with progressive contractile failure. Inhibition of glycolysis or SERCA2 caused an increase in [Ca 2+ ]i as well as a slower Ca. Pyruvate completely preserved myocardial function and [Ca 2+ ]i handling during glycolytic inhibition. It was concluded that in this model of advanced pressure overload hypertrophy, contractile failure and inotrope insensitivity are associated with increased [Ca 2+ ]i, slower Ca and reduced sensitivity of the contractile proteins to Ca 2+ . These changes occur in association with downregulation of the SERCA2, probably caused by impaired glucose metabolism. (Jpn Circ J 2001; 65: 1064 -1070
Basic Research in Cardiology, 2002
Biochemical changes and their relationship with morphological and functional findings in pig heart subjected to lasting volume overload: a possible role of acylphosphatase in the regulation of sarcoplasmic reticulum calcium pump Abstract We evaluated the changes in sarcoplasmic reticulum (SR) function and the parallel hemodynamic and morphological modifications in a heart subjected to volume overload. We also determined the levels of acylphosphatase, a cytosolic enzyme, that could play a regulatory effect on SR Ca 2+ pump by hydrolyzing the phosphorylated intermediate of this transport system. For this, swine hearts were subjected to volume overload by aortacava shunt for 1, 2, or 3 months. Changes in heart contractility reflected modifications of SR function, whose reduction after 1 month of overload was followed by a gradual recovery. A decrease in SERCA2a protein and mRNA content was shown from 1 month and remained for the following 2 months. Phospholamban content and its phosphorylation status were not modified. Acylphosphatase was unchanged at 1 month, but at 2 months this enzyme exhibited an increased activity, protein and mRNA expression. Morphological alterations consisting of the cytoskeletal architectures, intermyofibrillar oedema, swollen mithochondria and abnormality of the membrane system (T-tubule and SR cisternae) were particularly evident after 1 month but almost disappeared after 3 months. These results suggest that our overloaded hearts underwent a substantial recovery of their structural and biochemical properties at 3 months after surgery. A possible involvement of acylphosphatase in the modification of SR function is discussed.
Serine Phosphorylation of the Sarcoplasmic Reticulum Ca2+-ATPase in the Intact Beating Rabbit Heart
Biochemical and Biophysical Research Communications, 1999
Recent studies have demonstrated that Ca 2؉ / calmodulin-dependent protein kinase phosphorylates the Ca 2؉-pumping ATPase of cardiac sarcoplasmic reticulum (SR) in vitro. Also, evidence from in vitro studies suggested that this phosphorylation, occurring at Ser 38 , results in stimulation of Ca 2؉ transport. In the present study, we investigated whether serine phosphorylation of the SR Ca 2؉-ATPase occurs in the intact functioning heart. Hearts removed from anesthetized rabbits were subjected to retrograde aortic perfusion of the coronary arteries with oxygenated mammalian Ringer solution containing 32 P i and contractions were monitored by recording systolic left ventricular pressure development. Following 45-50 min of 32 P perfusion, the hearts were freeze-clamped, SR isolated, and analyzed for protein phosphorylation. SDS-polyacrylamide gel electrophoresis and autoradiography showed phosphorylation of several peptides including the Ca 2؉-ATPase and Ca 2؉ release channel (ryanodine receptor). The identity of Ca 2؉-ATPase as a phosphorylated substrate was confirmed by Western immunoblotting as well as immunoprecipitation using a cardiac SR Ca 2؉-ATPase-specific monoclonal antibody. The Ca 2؉-ATPase showed immunoreactivity with a phosphoserine monoclonal antibody indicating that the in situ phosphorylation occurred at the serine residue. Quantification of Ca 2؉-ATPase phosphorylation in situ yielded a value of 208 ؎ 12 pmol 32 P/mg SR protein which corresponded to the phosphorylation of ϳ20% of the Ca 2؉ pump units in the SR membrane. Since this phosphorylation occurred under basal conditions (i.e., in the absence of any inotropic intervention), a considerable steady-state pool of serine-phosphorylated Ca 2؉-ATPase likely exists in the normally beating heart. These findings demonstrate that serine phosphorylation of the Ca 2؉-ATPase is a physiological event which may be important in the regulation of SR function.
Subcellular Remodeling and Heart Dysfunction in Cardiac Hypertrophy due to Pressure Overloada
Annals of the New York Academy of Sciences, 1999
Rats were treated with etomoxir, an inhibitor of palmitoyltransferase-1, to examine the role of a shift in myocardial metabolism in cardiac hypertrophy. Pressure overload was induced by abdominal aorta banding for 8 weeks. Sham-operated animals served as control. Left ventricular dysfunction, as reflected by decreased LVDP, +dP/dt, −dP/dt, and elevated LVEDP in the pressure overloaded animals, was improved by treatment with etomoxir. Cardiac hypertrophy in pressure-overload rats decreased the sarcoplasmic reticular (SR) Ca 2+ uptake and Ca 2+ release as well as myofibrillar Ca 2+-stimulated ATPase and myosin Ca 2+-ATPase activities; these changes were attenuated by treatment with etomoxir. Steady-state mRNA levels for =and >-myosin heavy chains, SR Ca 2+-pump, and protein content of SR Ca 2+-pump were reduced in hypertrophied hearts; these alterations were prevented by etomoxir treatment. The results indicate that modification of changes in myocardial metabolism by etomoxir may prevent remodeling of myofibrils and SR membrane and thereby improve cardiac function in hypertrophied heart.
Journal of Clinical Investigation, 1990
A decrease in the myocardial level of the mRNA encoding the Ca2+-ATPase of the sarcoplasmic reticulum (SR) has been recently reported during experimental cardiac hypertrophy and failure. To determine if such a deficit occurs in human endstage heart failure, we compared the SR Ca2+-ATPase mRNA levels in left (LV) and right ventricular (RV) specimens from 13 patients undergoing cardiac transplantation (6 idiopathic dilated cardiomyopathies; 4 coronary artery diseases with myocardial infarctions; 3 diverse etiologies) with control heart samples using a rat cardiac SR Ca2+-ATPase cDNA probe. We observed a marked decrease in the mRNA for the Ca2+-ATPase relative to both the 18S ribosomal RNA and the myosin heavy chain mRNA in LV specimens of patients with heart failure compared to controls (-48%, P < 0.01 and -47%, P < 0.05, respectively). The LV ratio of Ca2+-ATPase mRNA to 18S RNA positively correlated with cardiac index (P < 0.02).
Basic Research in Cardiology, 1999
It is unknown whether the transmural heterogeneity of sarcoplasmic reticulum (SR) Ca 2+-ATPase gene expression is present within the left ventricular (LV) wall. Moreover, the changes of transmural distribution have not been examined in the failing hearts. We thus quantified steady-state mRNA abundance of SR Ca 2+ regulatory proteins by Northern blot analysis in both subendocardial and subepicardial LV layers from normal and rapid pacing-induced heart failure (HF) dog hearts. For normal LV, Ca 2+-ATPase mRNA abundance (normalized to glyceraldehyde-3-phosphate dehydrogenase [GAPDH] mRNA) was significantly reduced in the subendocardium, whereas calsequestrin mRNA abundance was comparable between the two layers. For HF LV, Ca 2+-ATPase mRNA abundance in the subendocardium was also reduced compared to the subepicardium. However, the endocardium to epicardium ratio was comparable between control and HF (0.62 ± 0.08 vs. 0.65 ± 0.07; p = NS). Therefore, the transmural gradient of this gene was constant in both control and HF. Even though the data on the transmural heterogeneity of protein level is not available, the subendocardium contained significantly less Ca 2+-ATPase mRNA, which might contribute, at least in part, to the transmural gradients of biochemical and mechanical function.
European Journal of Heart Failure, 2007
Background: Reduced myocardial contractility is often attributed to altered Ca 2+ transients and expression of Ca 2+ -ATPase of the SR (SERCA) and Na+/Ca 2+ exchanger (NCX) genes. Aims: To assess myocardial expression of SERCA and NCX protein levels in left ventricular (LV) remodelling due to chronic severe mitral regurgitation (MR). Methods: Myocardial expression of SERCA/NCX in biopsy specimens obtained during mitral surgery was assessed in 36 MR patients with LV remodelling and plasma neurohumoral/cytokine activation and in four non-failing hearts (NFH). Results: Myocardial protein levels of SERCA were significantly (20%) lower in the MR group than in NFH group (p = 0.016). No significant changes in NCX were observed. However, a lack of homogeneity with regard to SERCA/NCX proteins was observed. Moreover, SERCA was negatively correlated with BNP (r = −0.49, p = 0.02), TNFα (r = − 0.68, p = 0.0005) and IL-6 (r = −0.52, p = 0.02), whereas NCX was only negatively correlated with TNFα (r = −0.62, p = 0.002). Conclusions: MR patients showed wide variations in SERCA/NCX protein expression. Myocardial protein levels of SERCAwere significantly lower in the MR population. Moreover, a correlation between BNP, cytokines (IL-6, TNFα) and the expression of SERCA/NCX proteins was observed.
FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 2002
Slowed relaxation in diabetic cardiomyopathy (CM) is partially related to diminished expression of the sarcoplasmic reticulum (SR) Ca2+-ATPase SERCA2a. To evaluate the impact of SERCA2a overexpression on SR Ca2+ handling in diabetic CM, we 1) generated transgenic rats harboring a human cytomegalovirus enhancer/chicken beta-actin promotor-controlled rat SERCA2 transgene (SERCA2-TGR), 2) characterized their SR phenotype, and 3) examined whether transgene expression may rescue SR Ca2+ transport in streptozotocin-induced diabetes. The transgene was expressed in all heart chambers. Compared to wild-type (WT) rats, a heterozygous line exhibited increased SERCA2 mRNA (1.5-fold), SERCA2 protein (+26%) and SR Ca2+ uptake (+37%). Phospholamban expression was not altered. In SERCA2-TGR, contraction amplitude (+48%) and rates of contraction (+34%) and relaxation (+35%) of isolated papillary muscles (PM) were increased (P2+ uptake and SERCA2 protein of SERCA2-TGR were 1.3-fold higher (P2+ uptake...
AJP: Heart and Circulatory Physiology, 2006
Early cardiovascular changes evoked by pressure overload (PO) may reveal adaptive strategies that allow immediate survival to the increased hemodynamic load. In this study, systolic and diastolic Ca2+ cycling was analyzed in left ventricular rat myocytes before ( day 2, PO-2d group) and after ( day 7, PO-7d group) development of hypertrophy subsequent to aortic constriction, as well as in myocytes from time-matched sham-operated rats (sham group). Ca2+ transient amplitude was significantly augmented in the PO-2d group. In the PO-7d group, intracellular Ca2+ concentration ([Ca2+]i) was reduced during diastole, and mechanical twitch relaxation (but not [Ca2+]i decline) was slowed. In PO groups, fractional sarcoplasmic reticulum (SR) Ca2+ release at a twitch, SR Ca2+ content, SR Ca2+ loss during diastole, and SR-dependent integrated Ca2+ flux during twitch relaxation were significantly greater than in sham-operated groups, whereas the relaxation-associated Ca2+ flux carried by the Na+/...