[Effects of tumor necrosis factor alpha on expression of phospholamban and intracellular calcium in cardiomyocytes].
OBJECTIVE
To explore the effects of tumor necrosis factor alpha (TNFalpha) on the expression of phospholamban (PLB) and sarco (endo) plasmic reticulum Ca(2+)-ATPase (SERCA2a) and concentration of intracellular free calcium in myocardiocytes.


METHODS
The neonatal rat myocardiocytes were randomly divided into 6 groups: treatment with different concentrations of TNFalpha (1,10,30,50,and 70 microg/L, respectively) and without TNFalpha (control). The mRNA and protein expression of PLB and SERCA2a were detected with one-step reverse transcription-polymerase chain reaction and Western blotting. The changes of intracellular free calcium concentration ([Ca2+]i) in cultured single neonatal rat cardiomyocyte were determined with Fluo-3/AM loading by laser scanning confocal microscopy. RESULTS TNFalpha significantly increased the expression of PLB mRNA and protein in a dose-dependent fashion. The ratio of PLB/beta-actin mRNA in myocardiocytes incubated with 10,30,50, and 70 microg/L TNFalpha significantly increased by 66%, 106%, 141%, and 189% compared with control (P < 0.05), and protein levels significantly increased by 30%, 48%, 73%, and 114% respectively compared with control (P < 0.001), but there was no significant difference in PLB mRNA expression between the group treated with 1 microg/L TNFalpha and control group. TNFalpha had no effect on the expression of mRNA and protein of SERCA2a. TNFalpha (50 microg/L) incubated with cell for 24 hours diminished delta[Ca2+]i of single neonatal rat cardiomyocyte about 33% stimulated by isoproterenol (P < 0.01), but had no effect on delta [Ca2+]i of cardiomyocyte without isoproterenol stimulation.


CONCLUSIONS
TNFalpha can increase the expression of PLB and decrease delta[Ca2+]i in cardiomyocytes, which may be related with its negative inotropic effects on cardiomyocytes.
Yao
Yu-mei
YM
Department of Cardiology, the First Affiliated Hospital, Zhejiang University Medical College, Hangzhou 310003, China.
Hu
Shen-jiang
SJ
Huang
Yuan-wei
YW
Yang
Chun-hu
CH
Sun
Jian
J
Zhu
Zhao-hui
ZH
Wu
Tao
T
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics; D002478:Cells, Cultured; D005260:Female; D008297:Male; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000096:biosynthesis* / Q000235:genetics; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D000099211:Phospholamban
D002118:Calcium; D002135:Calcium-Binding Proteins; D012333:RNA, Messenger; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D014409:Tumor Necrosis Factor-alpha; D000099211:Phospholamban