Inhibitory effect of (-)-epigallocatechin-3-gallate on lipid accumulation of 3T3-L1 cells.
OBJECTIVE
The objective of this study was to investigate the molecular mechanisms underlying the attenuating effect of (-)-epigallocatechin-3-gallate (EGCG) on proliferation and lipid accumulation of 3T3-L1 cells, with a focus on the duration of EGCG treatment.


METHODS
Cell viability was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium assay and diamidino-2-phenylindole staining. The anti-adipogenic effect of EGCG on 3T3-L1 cells was analyzed by glycerol-3-phosphate dehydrogenase activity and Oil red O staining. Western blot analysis was used to detect adenosine monophosphate-activated protein kinase (AMPK) activation and phosphorylation of its substrate, acetyl-CoA carboxylase (ACC), and expression of insulin (INS) receptor, INS receptor substrate-1 (IRS-1), and adipocyte marker proteins.


RESULTS
Exposure to EGCG during the early period of adipogenesis (7 days) was sufficient to prevent lipid accumulation. During this period, EGCG greatly decreased expression of the adipocyte marker proteins peroxisome proliferator-activated receptor gamma2 (PPARgamma2) and liver X receptor (LXR)-alpha. Furthermore, EGCG significantly induced generation of reactive oxygen species (ROS), which led to AMPK activation, and these effects were eliminated by N-acetylcysteine (NAC) treatment. Also, EGCG increased the tyrosine phosphorylation of INS receptor and INS-1 with increasing incubation time. In contrast, EGCG treatment did not alter glycerol release in the presence or absence of 2',5'-dideoxyadenosine (DDA), indicating that EGCG had no effect on lipolysis.


CONCLUSIONS
Our data demonstrate that EGCG decreased cell viability and inhibited differentiation of 3T3-L1 cells in a manner dependent on the duration of treatment. Also, we showed that inhibition of adipocyte differentiation by EGCG was associated with decreased glycerol-3-phosphate dehydrogenase (GPDH) activity accompanied by a strong inhibition of PPARgamma2-induced transcriptional activity. Furthermore, the inhibition of adipocyte differentiation by EGCG involved generation of ROS and activation of AMPK.
Moon
Hyun-Seuk
HS
School of Agricultural Biotechnology, Seoul National University, San 56-1, Sillim-dong, Gwanak-gu, 151-921 Seoul, South Korea.
Chung
Chung-Soo
CS
Lee
Hong-Gu
HG
Kim
Tae-Gyu
TG
Choi
Yun-Jaie
YJ
Cho
Chong-Su
CS
D041721:3T3-L1 Cells / Q000187:drug effects* / Q000378:metabolism* / Q000473:pathology; D000091162:AMP-Activated Protein Kinase Kinases; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D002392:Catechin / Q000031:analogs & derivatives* / Q000494:pharmacology; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000037:antagonists & inhibitors / Q000378:metabolism; D055504:Insulin Receptor Substrate Proteins; D050356:Lipid Metabolism / Q000187:drug effects*; D000071518:Liver X Receptors; D051379:Mice; D057093:Orphan Nuclear Receptors; D047495:PPAR gamma / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000378:metabolism; D013997:Time Factors
D048868:Adaptor Proteins, Signal Transducing; D000975:Antioxidants; D004268:DNA-Binding Proteins; D055504:Insulin Receptor Substrate Proteins; C526295:Irs1 protein, mouse; D000071518:Liver X Receptors; C000605214:Nr1h3 protein, mouse; D057093:Orphan Nuclear Receptors; D047495:PPAR gamma; D017382:Reactive Oxygen Species; D018160:Receptors, Cytoplasmic and Nuclear; D002392:Catechin; C045651:epigallocatechin gallate; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D011494:Protein Kinases; D000091162:AMP-Activated Protein Kinase Kinases