Phorbol myristate acetate-induced hypertrophy of neonatal rat cardiac myocytes is associated with decreased sarcoplasmic reticulum Ca2+ ATPase (SERCA2) gene expression and calcium reuptake.
The decreased expression of the sarcoplasmic reticulum Ca(2+)-ATPase associated with cardiac hypertrophy was investigated in cultured neonatal rat cardiac myocytes. Northern blot analysis indicated a significant 55-60% decrease in Ca(2+)-ATPase mRNA levels and after 12 and 24 h of treatment with the phorbol ester phorbol myristate acetate (PMA). Myocytes treated with the phorbol ester for 80 h showed a significant 34% decrease (relative to vehicle-treated control cells) in the levels of Ca(2+)-ATPase protein, and a significant 38% increase in the levels of alpha-sarcomeric actin, as assessed by Western blot analysis using specific antibodies. Immunocytochemistry of myocytes treated for 72 h with the phorbol ester revealed a hypertrophied cell morphology, and showed a marked decrease in Ca(2+)-ATPase staining intensity. Contractile calcium transients were evaluated through the use of indo-1. It was found that the t1/2 for the decline of calcium transient was significantly prolonged by PMA treatment (0.51 +/- 0.15) when compared to controls (0.38 +/- 0.17, P < 0.001). Treatment of myocytes with endothelin-1 also led to a 35% decrease in sarcoplasmic reticulum Ca(2+)-ATPase mRNA levels. It is concluded that phorbol ester treatment of neonatal rat cardiac myocytes induces similar changes in Ca(2+)-ATPase mRNA levels. It is concluded that phorbol ester treatment of neonatal rat cardiac myocytes induces similar changes in Ca(2+)-ATPase gene expression as observed in vivo in the hypertrophied and failing heart. The observed prolongation in t1/2 for [Ca2+]i decline might be due to the observed depressed levels for sarcoplasmic reticulum Ca(2+)-ATPase in PMA treated cells.
Hartong
R
R
Department of Medicine, University of California San Diego, La Jolla 92103-0618, USA.
Villarreal
F J
FJ
Giordano
F
F
Hilal-Dandan
R
R
McDonough
P M
PM
Dillmann
W H
WH
D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D016232:Endothelins / Q000494:pharmacology; D015870:Gene Expression; D006352:Heart Ventricles / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism; D012333:RNA, Messenger; D011817:Rabbits; D051381:Rats; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors
D016232:Endothelins; D012333:RNA, Messenger; D000252:Calcium-Transporting ATPases; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium