Tumor necrosis factor-alpha decreases sarcoplasmic reticulum Ca2+-ATPase expressions via the promoter methylation in cardiomyocytes.
OBJECTIVE
Sarcoplasmic reticulum Ca-ATPases (SERCA2a) plays an essential role in the Ca homeostasis and cardiac functions. Tumor necrosis factor-alpha (TNF-alpha) decreases the SERCA2a, which may underlie cardiac dysfunction during sepsis and heart failure. Because the promoter region of SERCA2a contains CpG islands, gene methylation should be critical in regulating SERCA2a. The present study was to evaluate whether TNF-alpha can modulate SERCA2a via enhancing methylation and to investigate the underlying mechanisms.


METHODS
Controlled laboratory experiment.


METHODS
University research laboratory.


METHODS
HL-1 cardiomyocytes.


METHODS
TNF-alpha (1-50 ng/mL) was administered in HL-1 cardiomyocytes with and without co-administration of an NF-kappaB inhibitor (SN-50, 50 microg/mL), antioxidant agents (ascorbic acid, 100 microM, or coenzyme Q10, 10 microM), or methylation inhibitor (5-aza-2'-deoxycytidine, 0.1, 1 microM).


RESULTS
TNF-alpha (50 ng/mL) decreased the SERCA2a RNA and protein by quantitative polymerase chain reaction and immunoblot. Furthermore, TNF-alpha (50 ng/mL) increased the methylation in the SERCA2a promoter region, which was not influenced by the co-administration of SN-50, ascorbic acid, or coenzyme Q10, but was attenuated by 5-aza-2'-deoxycytidine (0.1 microM). Additionally, TNF-alpha (50 ng/mL) increased the expression of DNA methyltransferase 1.


CONCLUSIONS
TNF-alpha increased DNA methyltransferase levels, thus enhancing the methylation in the SERCA2a promoter region with a result of reducing SERCA2a. These findings suggest that inhibition of hypermethylation may be a novel treatment strategy for cardiac dysfunction.
Kao
Yu-Hsun
YH
Graduate Institute of Clinical Medicine, Taipei Medical University, Taipei, Taiwan.
Chen
Yao-Chang
YC
Cheng
Chen-Chuan
CC
Lee
Ting-I
TI
Chen
Yi-Jen
YJ
Chen
Shih-Ann
SA
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D015152:Blotting, Northern; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D019175:DNA Methylation; D005786:Gene Expression Regulation; D006325:Heart Atria / Q000166:cytology; D051379:Mice; D032383:Myocytes, Cardiac / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects; D011401:Promoter Regions, Genetic; D012313:RNA / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D012680:Sensitivity and Specificity; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D014409:Tumor Necrosis Factor-alpha; D012313:RNA; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases; D002118:Calcium