August 1, 1997
View Full PaperAdenoviral Gene Transfer of Phospholamban in Isolated Rat Cardiomyocytes
Key Result
Adenovirus-mediated overexpression of phospholamban in rat cardiomyocytes prolonged the relaxation phase (710 vs 344 ms, P<0.01) and decreased peak intracellular calcium (630 vs 967 nmol/L, P<0.01).
Structured PICO
P
Population
Neonatal rat cardiomyocytes used to study the effects of phospholamban overexpression on intracellular calcium handling.
I
Intervention
Recombinant adenovirus designed to overexpress phospholamban (Ad.RSV.PL) at a multiplicity of infection (MOI) of 10.0 pfu/cell
C
Comparator
Uninfected or control cardiomyocytes
O
Outcome
Intracellular Ca2+ transients (relaxation phase, peak [Ca2+]i, and resting [Ca2+]i)surrogate
Overexpression of phospholamban in rat cardiomyocytes impairs intracellular Ca2+ handling, mimicking abnormalities seen in heart failure, which can be partially corrected by simultaneous SERCA2a overexpression.
Main Result
p-value: p=<.01
Abstract
Phospholamban forms an integral part of the cardiac sarcoplasmic reticulum (SR) and regulates the activity of SR Ca(2+)-ATPase (SERCA2a). A number of studies have suggested a decrease in SERCA2a relative to phospholamban in heart failure. To test the hypothesis that changes in the relative abundance of phospholamban to SERCA2a could account for the pathophysiological abnormalities in Ca2+ handling observed in failing myocardium, we created a recombinant adenovirus designed to overexpress phospholamban (Ad.RSV.PL). In neonatal rat cardiomyocytes, Ad.RSV.PL increased the expression of phospholamban in a concentration-dependent fashion, reaching 280 +/- 43% at a multiplicity of infection (MOI) of 10.0 plaque forming units (pfu)/cell at 48 hours. The relationship between Ca(2+)-ATPase activity and Ca2+ was shifted rightward in membrane preparations from cardiomyocytes infected with Ad.RSV.PL. Intracellular Ca2+ transients measured in the neonatal cells infected with Ad.RSV.PL (MOI, 10 pfu/cell) were characterized by (1) a significant prolongation of the relaxation phase (344 +/- 26 versus 710 +/- 56 milliseconds, P < .01), (2) a decrease in peak Ca2+i (967 +/- 43 versus 630 +/- 33 nmol/L, P < .01), and (3) an elevation in resting Ca2+i (143 +/- 14 versus 213 +/- 17 nmol/L, P < .05). Similarly, the time course of shortening was prolonged in myocytes infected with Ad.RSV.PL. These effects were partially restored by simultaneous transduction with an adenovirus carrying SERCA2a. Cardiomyocytes infected with Ad.RSV.PL had an abnormal frequency response: a decrease in peak Ca2+i and an increase in resting Ca2+i with increasing frequency. These findings indicate that adenovirus-mediated gene transfer of phospholamban modifies intracellular Ca2+ handling and the frequency response in cardiomyocytes. Our results suggest that alterations in the ratio of phospholamban to SERCA2a could account for the abnormalities in Ca2+ handling observed in heart failure and that overexpression of SERCA2a can largely correct these abnormalities.
Cite This Study
Hajjar et al. (Fri,) conducted a other in Heart failure. Adenoviral gene transfer of phospholamban (Ad.RSV.PL) vs. Uninfected control cells was evaluated on Intracellular Ca2+ transients (relaxation phase, peak [Ca2+]i, resting [Ca2+]i) (p=<.01). Adenovirus-mediated overexpression of phospholamban in rat cardiomyocytes prolonged the relaxation phase (710 vs 344 ms, P<0.01) and decreased peak intracellular calcium (630 vs 967 nmol/L, P<0.01).