Journal of Obesity & Metabolic Syndrome

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Korean J Obes 2012; 21(1): 29-36

Published online March 30, 2012

Copyright © Korean Society for the Study of Obesity.

The Effect of AMPK Activation on Wnt and sFRP5 in TNF-α Induced Adipocyte Metabolic Dysfunction in 3T3-L1 Cell

Seok-Woo Hong, Jin-Mi Lee, Ji-Hun Choi(1), Se-Eun Park(1), Eun-Jung Rhee(1), Cheol-Young Park(1), Ki-Won Oh(1), Sung-Woo Park(1), Sun-Woo Kim(1), Won-Young Lee(1)*

Institute of Medical Research; and Department of Endocrinology and Metabolism(1), Kangbuk Samsung Hospital, Sungkyunkwan University School of Medicine

Background: Chronic inflammation in adipose tissue is closely associated with metabolic diseases, such as, type 2 diabetes and obesity. In the present study, we investigated the Wnt signaling pathway and fat metabolism in the course of TNF-α-induced inflammation and recovery in 3T3-L1 adipocyte.
Methods: We stimulated the fully differentiated 3T3-L1 adipocyte with either TNF-α only or with both TNF-α and AICAR for 24 hrs to induce inflammation. We assessed the alteration of Wnt signaling pathway and the factors associated with fat metabolism with western blot assay and real-time PCR.
Results: The expression of inflammatory cytokines, IL-6 and MCP-1, was increased by TNF-α treatment in fully differentiated 3T3-L1 adipocyte, and the marked activation of Wnt5a, a noncanonical Wnt ligand, was observed. The expression of PPAR-γ was reduced, and the lipolysis measured by glycerol release, was markedly increased. The activation of AMPK by AICAR inhibited the TNF-α-induced inflammation, reversed the alteration in Wnt signaling pathway, and reversed fat metabolism induced by TNF-α.
AMPK activation stimulated the secretion of sFRP5, an anti-inflammatory and anti-Wnt signaling adipokine.
Conclusion: The activation of AMPK suppressed noncanonical Wnt signaling pathway and protected the adipoctye from inflammation and lipolysis, induced by TNF-α treatment, by sFRP5 stimulation. Based on these results, we suggest that noncanonical Wnt signaling pathway and sFRP may have important roles in metabolic diseases such as, obesity and diabetes.

Keywords: Adipocyte, Inflammation, Noncanonical Wnt signaling pathway, sFRP5, AMPK, TNF-α

Fig. 1.

AMPK suppressed TNF-α induced inflammatory gene expression. 3T3-L1 adipocyte was incubated with TNF-α (10 ng/mL) for 24 hrs either in the presence or absence of AICAR (1 mM). The gene transcript of IL-6 (A) and MCP-1 (B) was measured by real-time RT-PCR.

*P < 0.05 and **P < 0.01 compared with control group.

P < 0.05 compared with TNF-α group.


Fig. 2.

Noncanonical wnt signaling is specifically involved in inflammation and restoration in 3T3-L1 adipocyte. Expression of ligand proteins and signaling molecules of noncanonical (A) and canonical (B) Wnt signaling pathway was evaluated by western blot assay.

*P < 0.05 and **P < 0.01 compared with control group.

P < 0.05 and ††P < 0.01 compared with TNF-α group.


Fig. 3.

AMPK prevented TNF-α induced insulin signaling impairment in 3T3-L1 adipocyte. The key molecules of insulin signaling, pIRS-1 (A), pAkt (B) were detected by a specific antibody. Total protein and β-actin were used for quantitative analysis.

*P < 0.05 compared with control group.

††P < 0.01 compared with TNF-α group.


Fig. 4.

TNF-α and AMPK regulated PPAR-γ expression and lipolysis. The amount of PPAR-γ protein (A) and released glycerol (B) to medium was estimated by western blot and free glycerol reagent.

**P < 0.01 compared with control group.

P < 0.05 and ††P < 0.01 compared with TNF-α group.


Fig. 5.

AMPK stimulated sFRP5 secretion in 3T3-L1 adipocyte. The amount of sFRP5 in medium and cell lysate was measured by western blot (A). The secretion rate was described as a ratio of secreted sFRP5 / non-secreted sFRP5. (B)

*P < 0.05 compared with control group.

P < 0.05 compared with TNF-α group.


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