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High-dose Resveratrol Inhibits Insulin Signaling Pathway in 3T3-L1 Adipocytes.

Lee H, Kim JW - J Lifestyle Med (2013)

Bottom Line: We treated differentiated 3T3-L1 adipocytes with resveratrol to observe whether resveratrol is effective at reducing lipid accumulation.Resveratrol treatment after mitotic clonal expansion resulted in decreased lipid accumulation accompanied by reduced fatty acid synthase expression.The results also provide information about in vivo administration dosages and may explain the discrepancy between in vitro and in vivo effects of resveratrol.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Institute of Genetic Science ; Brain Korea 21 Project for Medical Science.

ABSTRACT

Background: Insulin resistance is a major factor in the development of metabolic syndrome and is associated with central obesity and glucose intolerance. Resveratrol, a polyphenol found in fruits, has been shown to improve metabolic conditions. Although it has been widely studied how resveratrol affects metabolism, little is known about how resveratrol regulates lipogenesis with insulin signaling in 3T3-L1 adipocytes.

Methods: We treated differentiated 3T3-L1 adipocytes with resveratrol to observe whether resveratrol is effective at reducing lipid accumulation.

Results: Resveratrol treatment after mitotic clonal expansion resulted in decreased lipid accumulation accompanied by reduced fatty acid synthase expression. Decreased glucose uptake was observed with inhibited GLUT4 translocation in cells treated with 100 μM resveratrol, suggesting that high doses of resveratrol block insulin signaling in adipocytes. Insulin-stimulated Akt phosphorylation is also dose-dependently reduced with resveratrol treatment. Interestingly, Akt phosphorylation is upregulated when cells are treated with long-term low doses of resveratrol, suggesting that only low doses of resveratrol improve metabolic conditions.

Conclusion: High doses of resveratrol block the insulin signaling pathway, thereby reducing glucose uptake and lipid accumulation in vitro. The results also provide information about in vivo administration dosages and may explain the discrepancy between in vitro and in vivo effects of resveratrol.

No MeSH data available.


Related in: MedlinePlus

Resveratrol inhibits lipogenesis in 3T3-L1 adipocytes. (A) Confluent 3T3-L1 preadipocytes were induced to differentiate as described in Materials and Methods. After 2 days of differentiation, 3T3-L1 cells were treated with various amounts of resveratrol (a, 0.1% DMSO; b, 50 μM; c, 100 μM) for 3 days. After 8 days, cells were fixed in formaldehyde, and stained with oil Red-O to determine the degree of adipocyte differentiation. (B) Resveratrol represses the expression of adipogenic gene in 3T3-L1 adipocytes. Confluent 3T3-L1 preadipocytes were induced to differentiate as described. After 2 days of differentiation, 3T3-L1 cells were treated with various amount of resveratrol for 2 or 3 days and total cell extracts were prepared at the indicated time for Western blot analyses.
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f1-jlm-03-41: Resveratrol inhibits lipogenesis in 3T3-L1 adipocytes. (A) Confluent 3T3-L1 preadipocytes were induced to differentiate as described in Materials and Methods. After 2 days of differentiation, 3T3-L1 cells were treated with various amounts of resveratrol (a, 0.1% DMSO; b, 50 μM; c, 100 μM) for 3 days. After 8 days, cells were fixed in formaldehyde, and stained with oil Red-O to determine the degree of adipocyte differentiation. (B) Resveratrol represses the expression of adipogenic gene in 3T3-L1 adipocytes. Confluent 3T3-L1 preadipocytes were induced to differentiate as described. After 2 days of differentiation, 3T3-L1 cells were treated with various amount of resveratrol for 2 or 3 days and total cell extracts were prepared at the indicated time for Western blot analyses.

Mentions: Upon hormonal stimuli, growth-arrested 3T3-L1 preadipocytes rapidly reenter the cell cycle and undergo two or more rounds of mitotic clonal expansion in 2 days of differentiation induction [5]. In this period, cells begin to express C/EBPβ, C/EBPδ, and other transcription factors to activate the gene expression of two pleotropic transcription factors, C/EBPα and PPARγ, which are responsible for the acquisition of mature phenotype of adipocytes [17]. We previously reported that resveratrol inhibits adipogenesis by modulating the amount of reactive oxygen species, which blocks the C/EBPβ activation process [18]. In order to investigate the effect of resveratrol in lipid accumulation of adipocytes, the differentiating 3T3-L1 cells were treated with resveratrol after two days of differentiation induction, which may exclude the effect of resveratrol on mitotic clonal expansion (Fig. 1A). As a result, treatment of 50 μM or 100 μM resveratrol markedly reduced lipid accumulation in cells in a dose-dependent manner as shown by oil-red O staining (Fig. 1B). Consistent with these morphological changes, the expression of adipogenic genes including acetyl CoA carboxylase (ACC) α, fatty acid synthase (FAS), and PPARγ was significantly decreased with high doses (100 μM) of resveratrol over the course of 2–3 days (Fig. 1C). Because the expression of C/EBPα and PPARγ begins after approximately 36 h of differentiation induction, it is assumed that the resveratrol treatment protocol in this study does not affect differentiation per se, but instead it appears that the treatment may reduce the accumulation of lipid after differentiation. Moreover, reduced expression of lipogenic genes such as ACCα and FAS indicates that resveratrol inhibits lipid accumulation by inhibiting of genes involved in the lipogenesis process.


High-dose Resveratrol Inhibits Insulin Signaling Pathway in 3T3-L1 Adipocytes.

Lee H, Kim JW - J Lifestyle Med (2013)

Resveratrol inhibits lipogenesis in 3T3-L1 adipocytes. (A) Confluent 3T3-L1 preadipocytes were induced to differentiate as described in Materials and Methods. After 2 days of differentiation, 3T3-L1 cells were treated with various amounts of resveratrol (a, 0.1% DMSO; b, 50 μM; c, 100 μM) for 3 days. After 8 days, cells were fixed in formaldehyde, and stained with oil Red-O to determine the degree of adipocyte differentiation. (B) Resveratrol represses the expression of adipogenic gene in 3T3-L1 adipocytes. Confluent 3T3-L1 preadipocytes were induced to differentiate as described. After 2 days of differentiation, 3T3-L1 cells were treated with various amount of resveratrol for 2 or 3 days and total cell extracts were prepared at the indicated time for Western blot analyses.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4390751&req=5

f1-jlm-03-41: Resveratrol inhibits lipogenesis in 3T3-L1 adipocytes. (A) Confluent 3T3-L1 preadipocytes were induced to differentiate as described in Materials and Methods. After 2 days of differentiation, 3T3-L1 cells were treated with various amounts of resveratrol (a, 0.1% DMSO; b, 50 μM; c, 100 μM) for 3 days. After 8 days, cells were fixed in formaldehyde, and stained with oil Red-O to determine the degree of adipocyte differentiation. (B) Resveratrol represses the expression of adipogenic gene in 3T3-L1 adipocytes. Confluent 3T3-L1 preadipocytes were induced to differentiate as described. After 2 days of differentiation, 3T3-L1 cells were treated with various amount of resveratrol for 2 or 3 days and total cell extracts were prepared at the indicated time for Western blot analyses.
Mentions: Upon hormonal stimuli, growth-arrested 3T3-L1 preadipocytes rapidly reenter the cell cycle and undergo two or more rounds of mitotic clonal expansion in 2 days of differentiation induction [5]. In this period, cells begin to express C/EBPβ, C/EBPδ, and other transcription factors to activate the gene expression of two pleotropic transcription factors, C/EBPα and PPARγ, which are responsible for the acquisition of mature phenotype of adipocytes [17]. We previously reported that resveratrol inhibits adipogenesis by modulating the amount of reactive oxygen species, which blocks the C/EBPβ activation process [18]. In order to investigate the effect of resveratrol in lipid accumulation of adipocytes, the differentiating 3T3-L1 cells were treated with resveratrol after two days of differentiation induction, which may exclude the effect of resveratrol on mitotic clonal expansion (Fig. 1A). As a result, treatment of 50 μM or 100 μM resveratrol markedly reduced lipid accumulation in cells in a dose-dependent manner as shown by oil-red O staining (Fig. 1B). Consistent with these morphological changes, the expression of adipogenic genes including acetyl CoA carboxylase (ACC) α, fatty acid synthase (FAS), and PPARγ was significantly decreased with high doses (100 μM) of resveratrol over the course of 2–3 days (Fig. 1C). Because the expression of C/EBPα and PPARγ begins after approximately 36 h of differentiation induction, it is assumed that the resveratrol treatment protocol in this study does not affect differentiation per se, but instead it appears that the treatment may reduce the accumulation of lipid after differentiation. Moreover, reduced expression of lipogenic genes such as ACCα and FAS indicates that resveratrol inhibits lipid accumulation by inhibiting of genes involved in the lipogenesis process.

Bottom Line: We treated differentiated 3T3-L1 adipocytes with resveratrol to observe whether resveratrol is effective at reducing lipid accumulation.Resveratrol treatment after mitotic clonal expansion resulted in decreased lipid accumulation accompanied by reduced fatty acid synthase expression.The results also provide information about in vivo administration dosages and may explain the discrepancy between in vitro and in vivo effects of resveratrol.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Institute of Genetic Science ; Brain Korea 21 Project for Medical Science.

ABSTRACT

Background: Insulin resistance is a major factor in the development of metabolic syndrome and is associated with central obesity and glucose intolerance. Resveratrol, a polyphenol found in fruits, has been shown to improve metabolic conditions. Although it has been widely studied how resveratrol affects metabolism, little is known about how resveratrol regulates lipogenesis with insulin signaling in 3T3-L1 adipocytes.

Methods: We treated differentiated 3T3-L1 adipocytes with resveratrol to observe whether resveratrol is effective at reducing lipid accumulation.

Results: Resveratrol treatment after mitotic clonal expansion resulted in decreased lipid accumulation accompanied by reduced fatty acid synthase expression. Decreased glucose uptake was observed with inhibited GLUT4 translocation in cells treated with 100 μM resveratrol, suggesting that high doses of resveratrol block insulin signaling in adipocytes. Insulin-stimulated Akt phosphorylation is also dose-dependently reduced with resveratrol treatment. Interestingly, Akt phosphorylation is upregulated when cells are treated with long-term low doses of resveratrol, suggesting that only low doses of resveratrol improve metabolic conditions.

Conclusion: High doses of resveratrol block the insulin signaling pathway, thereby reducing glucose uptake and lipid accumulation in vitro. The results also provide information about in vivo administration dosages and may explain the discrepancy between in vitro and in vivo effects of resveratrol.

No MeSH data available.


Related in: MedlinePlus