Doxycycline Inducible Expression of SERCA2a Improves Calcium Handling and Reverts Cardiac Dysfunction in Pressure Overload-Induced Cardiac Hypertrophy (original) (raw)
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Cardiovascular Research, 2003
Objective: The activity of sarcoplasmic reticulum Ca -ATPase (SERCA) is reduced in the failing myocardium. Therefore, transfer of SERCA2a cDNA is considered as a therapeutical approach. The aim of this study was analysis of the long-term effect of SERCA2a overexpression in normal as well as pressure overload challenged myocardium of transgenic rats. Methods: Independent transgenic rat lines were established expressing the rat SERCA2a cDNA specifically in the myocardium resulting in increased SERCA2a protein levels by 30-70%. Simultaneous measurements of isometric contraction and calcium transients were carried out in right ventricular papillary muscle preparations. Hemodynamic parameters were measured in hearts of unchallenged rats as well as 10 weeks after pressure overload induced by abdominal aortic banding. Results: Analysis of calcium handling and contractile parameters in isolated right ventricular papillary muscles revealed significant shortening of intracellular calcium transients and half maximal relaxation times (RT ). Assessing 50 myocardial contractility in working heart preparations, both transgenic rat lines revealed elevated left ventricular pressure, improved systolic and diastolic parameters, attenuated negative force-frequency relation, and a dose-dependent b-adrenergic effect. Aortic banding resulted in reduction of left ventricular pressure and worsening of contraction and relaxation parameters with no differences in mortality in both transgenic (1dP/dt 3084696 vs. 39386250 mmHg / s; RT 47.061.2 vs. 36.761.4 ms) and wild-type rats (1dP/dt 2695686 vs. 50 32976122 mmHg / s; RT 53.061.6 vs. 44.161.4). SERCA2a overexpressing hearts revealed improved hemodynamic parameters 50 compared to wild-type controls. Acceleration of isovolumetric relaxation characterized by the index Tau was directly correlated to SERCA2a protein concentrations. Conclusion: Overexpression of SERCA2a protein results in a positive inotropic effect under baseline conditions remaining preserved under pressure overload without affecting mortality. Therefore therapeutic transfer of SERCA2a may become a potential approach for gene therapy of congestive heart failure. Moreover, transgenic SERCA2a rats will be useful for studies of long-term SERCA2a overexpression in further cardiovascular disease models.
SERCA2a: its role in the development of heart failure and as a potential therapeutic target
Research Reports in Clinical Cardiology, 2014
The complexity of heart physiology has delayed the implementation of efficient, feasible, and safe therapies to fight against heart diseases for many years. As knowledge of the precise mechanisms governing cardiac hypertrophy and heart failure development increases, the availability of new therapeutic alternatives also grows. Since the cardiomyocyte physiology deeply depends on the correct calcium handling, many efforts to describe accurately the excitation-contraction coupling process in the heart and the proteins involved have been made. Among the proteins participating in calcium handling, sarco/endoplasmic reticulum Ca 2+ adenosine triphosphatase-2a (SERCA2a), whose expression and function is decreased in heart failure, stands out because of its critical role regulating Ca 2+ concentration in the cardiomyocyte. The importance of SERCA2a has been reflected in numerous studies aimed to describe its expression and function. Recently, gene therapy to deliver SERCA2a has shown promising results in human clinical trials. This paper reviews the current literature knowledge exploring diverse approaches to rescue SERCA2a expression in heart failure. It also discusses some data suggesting other possible therapies that could improve SERCA2a expression and function in cardiac diseases.
Moderate heart dysfunction in mice with inducible cardiomyocyte-specific excision of the Serca2 gene
Journal of Molecular and Cellular Cardiology, 2009
The sarco(endo)plasmic reticulum calcium ATPase 2 (SERCA2) transports Ca 2+ from cytosol into the sarcoplasmic reticulum (SR) of cardiomyocytes, thereby maintaining the store of releasable Ca 2+ necessary for contraction. Reduced SERCA function has been linked to heart failure, and loss of SERCA2 in the adult mammalian heart would be expected to cause immediate severe myocardial contractile dysfunction and death. We investigated heart function in adult mice with an inducible cardiomyocyte-specific excision of the Atp2a2 (Serca2) gene (SERCA2 KO). Seven weeks after induction of Serca2 gene excision, the mice displayed a substantial reduction in diastolic function with a 5-fold increase in the time constant of isovolumetric pressure decay (tau). However, already at 4 weeks following gene excision less than 5% SERCA2 protein was found in myocardial tissue. Surprisingly, heart function was only moderately impaired at this time point. Tissue Doppler imaging showed slightly reduced peak systolic tissue velocity and a less than 2-fold increase in tau was observed. The SR Ca 2+ content was dramatically reduced in cardiomyocytes from 4-week SERCA2 KO mice, and Ca 2+ transients were predominantly generated by enhanced Ca 2+ flux through L-type Ca 2+ channels and the Na +-Ca 2+ exchanger. Moreover, equivalent increases in cytosolic [Ca 2+ ] in control and SERCA2 KO myocytes induced greater cell shortening in SERCA2 KO, suggesting enhanced myofilament responsiveness. Our data demonstrate that SR-independent Ca 2+ transport mechanisms temporarily can prevent major cardiac dysfunction despite a major reduction of SERCA2 in cardiomyocytes.
Journal of Biological Chemistry, 2000
The sarcoplasmic reticulum calcium ATPase SERCA2b is an alternate isoform encoded by the SERCA2 gene. SERCA2b is expressed ubiquitously and has a higher Ca 2؉ affinity compared with SERCA2a. We made transgenic mice that overexpress the rat SERCA2b cDNA in the heart. SERCA2b mRNA level was approximately ϳ20-fold higher than endogenous SERCA2b mRNA in transgenic hearts. SERCA2b protein was increased 8-10-fold in the heart, whereas SERCA2a mRNA/protein level remained unchanged. Confocal microscopy showed that SERCA2b is localized preferentially around the T-tubules of the SR, whereas SERCA2a isoform is distributed both transversely and longitudinally in the SR membrane. Calciumdependent calcium uptake measurements showed that the maximal velocity of Ca 2؉ uptake was not changed, but the apparent pump affinity for Ca 2؉ (K 0.5) was increased in SERCA2b transgenic mice (0.199 ؎ 0.011 M) compared with wild-type control mice (0.269 ؎ 0.012 M, p < 0.01). Work-performing heart preparations showed that SERCA2b transgenic hearts had a higher rates of contraction and relaxation, shorter time to peak pressure and half-time for relaxation than wild-type hearts. These data show that SERCA2b is associated in a subcompartment within the sarcoplasmic reticulum of cardiac myocytes. Overexpression of SERCA2b leads to an increase in SR calcium transport function and increased cardiac contractility, suggesting that SERCA2b plays a highly specialized role in regulating the beat-to-beat contraction of the heart.
Transcriptional changes following restoration of SERCA2a levels in failing rat hearts
The FASEB Journal, 2004
Heart failure is characterized at the cellular level by impaired contractility and abnormal Ca 2+ homeostasis. We have previously shown that restoration of a key enzyme that controls intracellular Ca 2+ handling, the sarcoplasmic reticulum Ca 2+ ATPase (SERCA2a), induces functional improvement in heart failure. We used high-density oligonucleotide arrays to explore the effects of gene transfer of SERCA2a on genetic reprogramming in a model of heart failure. A total of 1,300 transcripts were identified to be unmodified by the effect of virus alone. Of those, 251 transcripts were found to be up-or down-regulated upon failure. A total of 51 transcripts which were either up-(27) or down-(24) regulated in heart failure were normalized to the nonfailing levels by the restoration of SERCA2a by gene transfer. The microarray analysis identified new genes following SERCA2a restoration in heart failure, which will give us insights into their role in the normalization of multiple pathways within the failing cell.
Enhanced L-type calcium currents in cardiomyocytes from transgenic rats overexpressing SERCA2a
Experimental and Clinical Cardiology, 2010
Previous research reported that transgenic rats overexpressing the sarco(endo)plasmic reticulum Ca(2+)-ATPase SERCA2a exhibit improved contractile function of the myocardium. Furthermore, impaired Ca(2+) uptake and reduced relaxation rates in rats with diabetic cardiomyopathy were partially rescued by transgenic expression of SERCA2a in the heart. To explore whether enhanced Ca(2+) cycling in the cardiomyocytes of SERCA2a transgenic rats is associated with changes in L-type Ca(2+) (I(Ca-L)) currents. The patch-clamp technique was used to measure whole-cell currents in cardiomyocytes from transgenic rats overexpressing SERCA2a and from wild-type (nontransgenic) animals. The amplitudes of I(Ca-L) currents at depolarizing pulses ranging from -45 mV to 0 mV (350 ms duration, 1 Hz) were significantly higher in cardiomyocytes of SERCA2a transgenic rats than in nontransgenic rats (1985±48 pA [n=32] versus 1612±55 pA [n=28], respectively). The inactivation kinetics of I(Ca-L) showed subtle ...
Circulation. Arrhythmia and electrophysiology, 2011
Sarcoplasmic reticulum calcium ATPase 2a (SERCA2a) gene therapy improves mechanical function in heart failure and is under evaluation in a clinical trial. A critical question is whether SERCA2a gene therapy predisposes to increased sarcoplasmic reticulum calcium (SR Ca(2+)) leak, cellular triggered activity, and ventricular arrhythmias in the failing heart. We studied the influence of SERCA2a gene therapy on ventricular arrhythmogenesis in a rat chronic heart failure model. ECG telemetry studies revealed a significant antiarrhythmic effect of SERCA2a gene therapy with reduction of both spontaneous and catecholamine-induced arrhythmias in vivo. SERCA2a gene therapy also reduced susceptibility to reentry arrhythmias in ex vivo programmed electrical stimulation studies. Subcellular Ca(2+) homeostasis and spontaneous SR Ca(2+) leak characteristics were measured in failing cardiomyocytes transfected in vivo with a novel AAV9.SERCA2a vector. SR Ca(2+) leak was reduced after SERCA2a gene t...
Journal of molecular and …, 2006
Abnormal Ca 2+ cycling in the failing heart might be corrected by enhancing the activity of the cardiac Ca 2+ pump, the sarco(endo)plasmic reticulum Ca 2+ -ATPase 2a (SERCA2a) isoform. This can be obtained by increasing the pump's affinity for Ca 2+ by suppressing phospholamban (PLB) activity, the in vivo inhibitor of SERCA2a. In SKO mice, gene-targeted replacement of SERCA2a by SERCA2b, a pump with a higher Ca 2+ affinity, results in cardiac hypertrophy and dysfunction. The stronger PLB inhibition on cardiac morphology and performance observed in SKO was investigated here in DKO mice, which were obtained by crossing SKO with PLB −/− mice. The affinity for Ca 2+ of SERCA2 was found to be further increased in these DKO mice. Relative to wild-type and SKO mice, DKO mice were much less spontaneously active and showed a reduced life span. The DKO mice also displayed a severe cardiac phenotype characterized by a more pronounced concentric hypertrophy, diastolic dysfunction and increased ventricular stiffness. Strikingly, beta-adrenergic or forced exercise stress induced acute heart failure and death in DKO mice. Therefore, the increased PLB inhibition represents a compensation for the imposed high Ca 2+ -affinity of SERCA2b in the SKO heart. Limiting SERCA2's affinity for Ca 2+ is physiologically important for normal cardiac function. An improved Ca 2+ transport in the sarcoplasmic reticulum may correct Ca 2+ mishandling in heart failure, but a SERCA pump with a much higher Ca 2+ affinity may be detrimental.