Inhibitory Effect of (−)‐Epigallocatechin‐3‐Gallate on Lipid Accumulation of 3T3‐L1 Cells

Abstract

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. Research Methods and Procedures: 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 γ2 (PPARγ2) and liver X receptor (LXR)‐α. 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. Discussion: 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 PPARγ2‐induced transcriptional activity. Furthermore, the inhibition of adipocyte differentiation by EGCG involved generation of ROS and activation of AMPK.