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  • Title: Tight control of exogenous SERCA expression is required to obtain acceleration of calcium transients with minimal cytotoxic effects in cardiac myocytes.
    Author: O'Donnell JM, Sumbilla CM, Ma H, Farrance IK, Cavagna M, Klein MG, Inesi G.
    Journal: Circ Res; 2001 Mar 02; 88(4):415-21. PubMed ID: 11230109.
    Abstract:
    Collateral effects of exogenous sarcoendoplasmic reticulum Ca(2+) ATPase (SERCA) expression were characterized in neonatal rat and chicken embryo cardiac myocytes, and the conditions required to produce acceleration of Ca(2+) transients with minimal toxicity were established. Cultured myocytes were infected with adenovirus vector carrying the cDNA of wild-type SERCA1, an inactive SERCA1 mutant, or enhanced green fluorescence protein under control of the cytomegalovirus promoter. Controls were exposed to empty virus vector. Each group was tested with and without phenylephrine (PHE) treatment. Under conditions of limited calf-serum exposure, the infected rat myocytes manifested a more rapid increase in size, protein content, and rate of protein synthesis relative to noninfected controls. These changes were not accompanied by reversal to fetal transcriptional pattern (as observed in hypertrophy triggered by PHE) and may be attributable to facilitated exchange with serum factors. SERCA virus titers >5 to 6 plaque-forming units per cell produced overcrowding of ATPase molecules on intracellular membranes, followed by apoptotic death of a significant number of rat but not chicken myocytes. Enhanced green fluorescence protein virus and empty virus also produced cytotoxic effects but at higher titers than SERCA. Expression of exogenous SERCA and enhancement of Ca(2+) transient kinetics could be obtained with minimal cell damage in rat myocytes if the SERCA virus titer were maintained within 1 to 4 plaque-forming units per cell. Expression of endogenous SERCA was unchanged, but expression of exogenous SERCA was higher in myocytes rendered hypertrophic by treatment with PHE than in nontreated controls.
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