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Cardiovascular Research 2001 49(2):288-297; doi:10.1016/S0008-6363(00)00234-0
© 2001 by European Society of Cardiology
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Copyright © 2000, European Society of Cardiology

Adenoviral SERCA1a gene transfer to adult rat ventricular myocytes induces physiological changes in calcium handling

Nathalie Chossata, Frank Griscellib, Philippe Jourdonc, Damien Logearta, Thierry Ragotb, Michèle Heimburgera, Michel Perricaudetb, Anne-Marie Lompréc, Stéphane Hatema and Jean-Jacques Mercadiera,*

aINSERM U 460, Faculté de Médecine Xavier Bichat, 16 rue Henri Huchard, 75018 Paris, France
bCNRS UMR 1582, Institut Gustave Roussy, Villejuif, France
cCNRS URA 1131, Université de Paris XI, Orsay, France

* Corresponding author. Tel.: +33-1-4485-6158; fax: +33-1-4485-6157 jjmercadier{at}wanadoo.fr

Objective: We examined the functional consequences of expressing adult rabbit fast skeletal sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA1a) in isolated adult rat ventricular myocytes. Methods: Myocytes were infected with a recombinant adenovirus harboring SERCA1a. Then 2 days after myocyte infection, protein expression was estimated using Western blot and SDS–PAGE analysis. We also measured the ATP-dependent oxalate-facilitated Ca2+ uptake of myocyte homogenates and monitored Ca2+ transient in myocytes loaded with the Ca2+ dye, indo-1. Results: SERCA1a gene expression resulted in a 36% increase in the total SERCA protein level in infected myocytes compared to controls (P<0.01), while SERCA2 and phospholamban levels did not change. This increase was associated with a 42% rise in SR Ca2+ uptake (P<0.01), while {tau} (the time constant of Ca2+ transient decay), and the time to peak fell by 32% (P<0.01) and 38% (P<0.001), respectively. Increasing the frequency of stimulation from 0.2 to 2 Hz decreased {tau} in both cell types (P<0.01). However, the decrease was much smaller in infected (P<0.01) than in uninfected cells (P<0.001). Isoproterenol (1 µM) further decreased {tau} in infected myocytes by 23% (P<0.05). In these cells, the diastolic [Ca2+]i decreased by 50% (P<0.05) while the systolic [Ca2+]i increased by 19% (P<0.05). No difference was found in the speed of SR Ca2+ reloading after caffeine washout between the two cell types. Conclusion: Adenovirus-mediated SERCA1a gene transfer to adult rat ventricular myocytes enhances SR Ca2+ handling to a degree similar to that observed following physiological stimulation.

KEYWORDS Gene therapy; Myocytes; Ca-pump; SR (function); Cell culture/isolation


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