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Suppression of Very Early Stage Of Adipogenesis by Baicalein, a Plant-Derived Flavonoid through Reduced Akt-C/EBP α -GLUT4 Signaling-Mediated Glucose Uptake in 3T3-L1 Adipocytes

View Article: PubMed Central - PubMed

ABSTRACT

Baicalein has been used as a Chinese medicine, and is an abundant plant flavonoid present in fruits and vegetables. Here, we examined the effects of baicalein in adipogenesis and investigated its molecular mechanism in adipocytes. Baicalein lowered the intracellular lipid accumulation and decreased the transcription levels of the adipocyte-specific genes in mouse 3T3-L1 adipocytes. Glucose uptake mediated by glucose transporter 4 (GLUT4) was reduced, causing down-regulation of the intracellular lipid accumulation. These reductions were also observed even when baicalein was added in only early stage of adipogenesis (0–2 days) of 6-day-adipogenesis. Chromatin immunoprecipitation assay showed that baicalein decreased the binding level of C/EBPα protein to the promoter region of the GLUT4 gene. Phosphorylation of Akt at 1 h after the initiation of adipogenesis was inhibited by the treatment with baicalein. Inhibition during only the first 1.5 h after the initiation of adipogenesis by baicalein or an Akt inhibitor was enough to decrease the lipid contents in the cells undergoing adipocyte differentiation for 6 days. These results indicate that baicalein decreased the intracellular lipid accumulation by down-regulation of glucose uptake via repression of Akt-C/EBPα-GLUT4 signaling in the very early stage of adipogenesis of 3T3-L1 adipocytes.

No MeSH data available.


Differentiation stage-specific effects of baicalein during adipogenesis.A, Addition of baicalein at the indicated time-points during adipocyte differentiation of 3T3-L1 cells for 6 days. B, Oil Red O staining of intracellular lipids in 3T3-L1 cells in DMEM with stage-specific addition of baicalein. The cells (undifferentiated cells: U; white column) were differentiated into adipocytes (differentiated cells: D) for 6 days in DMEM without (gray column) or with (50 μM; black columns) baicalein during days 0–6, 0–2, 2–4, or 4–6. C, Intracellular triglyceride level. 3T3-L1 cells (undifferentiated cells: U) were differentiated into adipocytes (differentiated cells: D) as described in the legend of Fig 3B. Data are the means ± S.D. from three experiments. **p<0.01, as indicated by the brackets.
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pone.0163640.g004: Differentiation stage-specific effects of baicalein during adipogenesis.A, Addition of baicalein at the indicated time-points during adipocyte differentiation of 3T3-L1 cells for 6 days. B, Oil Red O staining of intracellular lipids in 3T3-L1 cells in DMEM with stage-specific addition of baicalein. The cells (undifferentiated cells: U; white column) were differentiated into adipocytes (differentiated cells: D) for 6 days in DMEM without (gray column) or with (50 μM; black columns) baicalein during days 0–6, 0–2, 2–4, or 4–6. C, Intracellular triglyceride level. 3T3-L1 cells (undifferentiated cells: U) were differentiated into adipocytes (differentiated cells: D) as described in the legend of Fig 3B. Data are the means ± S.D. from three experiments. **p<0.01, as indicated by the brackets.

Mentions: Next, we elucidated the molecular mechanism of baicalein-mediated inhibition of adipogenesis in 3T3-L1 adipocytes. At first, we investigated the effect of baicalein on adipogenesis at different stages of the differentiation process. Adipogenesis (for 6 days; days 0–6) was divided into 3 stages: early stage (day 0–2) at clonal expansion stage; middle stage (day 2–4) at initiation of adipogenesis, and late stage (after day 4) at maturation of adipocytes. 3T3-L1 cells were differentiated into adipose cells in medium containing baicalein at the indicated stages (Fig 4A). On day 6, the intracellular lipid-droplets were visualized by staining with Oil Red O, and the triglyceride levels in the cells were also measured. When baicalein was present during days 0–2 or days 0–6 of 6-day-adipogenesis, the accumulation of lipids in the cells was decreased as compared with the vehicle-treated differentiated cells (Fig 4B and 4C). In contrast, when baicalein was present in the medium during days 2–4 or days 4–6 in 6-day-adipogenesis, the intracellular lipid levels were almost the same as that for the vehicle-treated differentiated cells (Fig 4B and 4C). Therefore, these results reveal that baicalein repressed the intracellular lipid accumulation during the early stage of adipogenesis.


Suppression of Very Early Stage Of Adipogenesis by Baicalein, a Plant-Derived Flavonoid through Reduced Akt-C/EBP α -GLUT4 Signaling-Mediated Glucose Uptake in 3T3-L1 Adipocytes
Differentiation stage-specific effects of baicalein during adipogenesis.A, Addition of baicalein at the indicated time-points during adipocyte differentiation of 3T3-L1 cells for 6 days. B, Oil Red O staining of intracellular lipids in 3T3-L1 cells in DMEM with stage-specific addition of baicalein. The cells (undifferentiated cells: U; white column) were differentiated into adipocytes (differentiated cells: D) for 6 days in DMEM without (gray column) or with (50 μM; black columns) baicalein during days 0–6, 0–2, 2–4, or 4–6. C, Intracellular triglyceride level. 3T3-L1 cells (undifferentiated cells: U) were differentiated into adipocytes (differentiated cells: D) as described in the legend of Fig 3B. Data are the means ± S.D. from three experiments. **p<0.01, as indicated by the brackets.
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Related In: Results  -  Collection

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pone.0163640.g004: Differentiation stage-specific effects of baicalein during adipogenesis.A, Addition of baicalein at the indicated time-points during adipocyte differentiation of 3T3-L1 cells for 6 days. B, Oil Red O staining of intracellular lipids in 3T3-L1 cells in DMEM with stage-specific addition of baicalein. The cells (undifferentiated cells: U; white column) were differentiated into adipocytes (differentiated cells: D) for 6 days in DMEM without (gray column) or with (50 μM; black columns) baicalein during days 0–6, 0–2, 2–4, or 4–6. C, Intracellular triglyceride level. 3T3-L1 cells (undifferentiated cells: U) were differentiated into adipocytes (differentiated cells: D) as described in the legend of Fig 3B. Data are the means ± S.D. from three experiments. **p<0.01, as indicated by the brackets.
Mentions: Next, we elucidated the molecular mechanism of baicalein-mediated inhibition of adipogenesis in 3T3-L1 adipocytes. At first, we investigated the effect of baicalein on adipogenesis at different stages of the differentiation process. Adipogenesis (for 6 days; days 0–6) was divided into 3 stages: early stage (day 0–2) at clonal expansion stage; middle stage (day 2–4) at initiation of adipogenesis, and late stage (after day 4) at maturation of adipocytes. 3T3-L1 cells were differentiated into adipose cells in medium containing baicalein at the indicated stages (Fig 4A). On day 6, the intracellular lipid-droplets were visualized by staining with Oil Red O, and the triglyceride levels in the cells were also measured. When baicalein was present during days 0–2 or days 0–6 of 6-day-adipogenesis, the accumulation of lipids in the cells was decreased as compared with the vehicle-treated differentiated cells (Fig 4B and 4C). In contrast, when baicalein was present in the medium during days 2–4 or days 4–6 in 6-day-adipogenesis, the intracellular lipid levels were almost the same as that for the vehicle-treated differentiated cells (Fig 4B and 4C). Therefore, these results reveal that baicalein repressed the intracellular lipid accumulation during the early stage of adipogenesis.

View Article: PubMed Central - PubMed

ABSTRACT

Baicalein has been used as a Chinese medicine, and is an abundant plant flavonoid present in fruits and vegetables. Here, we examined the effects of baicalein in adipogenesis and investigated its molecular mechanism in adipocytes. Baicalein lowered the intracellular lipid accumulation and decreased the transcription levels of the adipocyte-specific genes in mouse 3T3-L1 adipocytes. Glucose uptake mediated by glucose transporter 4 (GLUT4) was reduced, causing down-regulation of the intracellular lipid accumulation. These reductions were also observed even when baicalein was added in only early stage of adipogenesis (0&ndash;2 days) of 6-day-adipogenesis. Chromatin immunoprecipitation assay showed that baicalein decreased the binding level of C/EBP&alpha; protein to the promoter region of the GLUT4 gene. Phosphorylation of Akt at 1 h after the initiation of adipogenesis was inhibited by the treatment with baicalein. Inhibition during only the first 1.5 h after the initiation of adipogenesis by baicalein or an Akt inhibitor was enough to decrease the lipid contents in the cells undergoing adipocyte differentiation for 6 days. These results indicate that baicalein decreased the intracellular lipid accumulation by down-regulation of glucose uptake via repression of Akt-C/EBP&alpha;-GLUT4 signaling in the very early stage of adipogenesis of 3T3-L1 adipocytes.

No MeSH data available.