Actin isoform synthesis by cultured cardiac myocytes. Effects of doxorubicin.
Cultured neonatal rat myocardial cells (CMC) were incubated with 10(-10) M to 10(-5) M doxorubicin (adriamycin, ADR) and [35S]methionine to determine incorporation of radiolabeled methionine into myocardial contractile proteins. Cells were harvested after 24 hours homogenized, and subjected to centrifugation. Equivalent amounts of extracted protein were applied to 8 to 15% gradient sodium dodecylsulfate polyacrylamide gel electrophoresis. Similar aliquots were subjected to isoelectric focusing and to 2-dimensional gel electrophoresis. Electrophoretic gels were autoradiographed. Polypeptide bands on sodium dodecylsulfate polyacrylamide gel electrophoresis and autoradiograms were quantitated densitometrically. No effect of ADR on CMC actin or protein synthesis was seen from 10(-10) M to 10(-7) M ADR. ADR decreased protein synthesis in CMC by 31% at 10(-6) M and by 59% at 10(-5) M ADR. Autoradiograms of two-dimensional gels showed decreased radiolabeling of alpha-actin at 10(-6) M ADR compared to beta and gamma. Decreased CMC actin synthesis initiated at 10(-6) M ADR resulted in selective decrease in synthesis of the alpha-isoform. This in vitro observation may relate to poor contractility in ADR heart muscle disease.
D000199:Actins / Q000096:biosynthesis*; D000818:Animals; D000831:Animals, Newborn; D001345:Autoradiography; D002478:Cells, Cultured; D003720:Densitometry; D004317:Doxorubicin / Q000633:toxicity*; D004591:Electrophoresis, Polyacrylamide Gel; D006321:Heart / Q000187:drug effects*; D008715:Methionine / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism; D014176:Protein Biosynthesis; D051381:Rats
D000199:Actins; D004317:Doxorubicin; D008715:Methionine