Nutr Res Pract.  2021 Aug;15(4):444-455. 10.4162/nrp.2021.15.4.444.

Effects of quercetin on cell differentiation and adipogenesis in 3T3-L1 adipocytes

• Hong SY
• Ha AW
• Kim W

• KMID: 2518753
• DOI: http://doi.org/10.4162/nrp.2021.15.4.444

Abstract

BACKGROUND/OBJECTIVES
Adipocytes undergo angiogenesis to receive nutrients and oxygen needed for adipocyte' growth and differentiation. No study relating quercetin with angiogenesis in adipocytes exists. Therefore, this study investigated the role of quercetin on adipogenesis in 3T3-L1 cells, acting through matrix metalloproteinases (MMPs).
MATERIALS/METHODS
After proliferating preadipocytes into adipocytes, various quercetin concentrations were added to adipocytes, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays were performed to evaluate cell proliferation. Glycerol-3-phosphate dehydrogenase (GPDH) activity was investigated as an indicator of fat accumulation. The mRNA expressions of transcription factors related to adipocyte differentiation, CCAAT/enhancer-binding proteins (C/EBPs), peroxisomal proliferatoractivated receptors (PPAR)-γ, and adipocyte protein 2 (aP2), were investigated. The mRNA expressions of proteins related to angiogenesis, vascular endothelial growth factor (VEGF)-α, vascular endothelial growth factor receptor (VEGFR)-2, MMP-2, and MMP-9, were investigated. Enzyme activities and concentrations of MMP-2 and MMP-9 were also measured.
RESULTS
Quercetin treatment suppressed fat accumulation and the expressions of adipocyte differentiation-related genes (C/EBPα, C/EBPβ, PPAR-γ, and aP2) in a concentration-dependent manner in 3T3-L1 cells. Quercetin treatments reduced the mRNA expressions of VEGF-α, VEGFR-2, MMP-2, and MMP-9 in 3T3-L1 cells. The activities and concentrations of MMP-2 and MMP-9 were also decreased significantly as the concentration of quercetin increased.
CONCLUSIONS
The results confirm that quercetin inhibits adipose tissue differentiation and fat accumulation in 3T3-L1 cells, which could occur through inhibition of the angiogenesis process related to MMPs.

Keyword

Quercetin; adipocyte; adipogenesis; vascular endothelial growth factor

Figure

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  Fig. 1 Effect of quercetin on cell proliferation in 3T3-L1 cells.After differentiation induction, as described in Methods, a cell monolayer was incubated in post-differentiation medium with 0, 5, 10, or 20 μM quercetin. Viable cell numbers were estimated by MTT assay at 0, 24, 48, and 96 h after quercetin treatments. Each bar represents the mean ± SE. Comparisons among different concentrations of quercetin that produced significant differences (P < 0.05) are indicated by different letters above each bar.MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide.

• 

  Fig. 2 Effect of quercetin on GPDH activity in 3T3-L1 cells.After differentiation induction, the cell monolayer was incubated in post-differentiation medium with 0, 5, 10, or 20 μM quercetin for 24 h. GPDH activity was estimated by using a commercial GPDH activity assay kit. Each bar represents the mean ± SE. Comparisons among different concentrations of quercetin that produced significant differences (P < 0.05) are indicated by different letters above each bar.GPDH, glycerol-3-phosphate dehydrogenase.

• 

  Fig. 3 Effect of quercetin on C/EBPα, C/EBPβ, PPAR-γ, and aP2 mRNA expressions in 3T3-L1 cells.After differentiation induction, the cell monolayer was incubated in post-differentiation medium with 0, 5, 10, or 20 μM quercetin for 24 h. Total RNA was isolated, and real-time PCR was performed. β-actin levels were compared, acting as the equal loading control. Each bar represents the mean ± SE. Comparisons among different concentrations of quercetin that generated significant differences (P < 0.05) are indicated by different letters above each bar.C/EBP, CCAAT/enhancer-binding protein; PPAR, peroxisomal proliferator-activated receptors; aP2, adipocyte protein 2; PCR, polymerase chain reaction.

• 

  Fig. 4 Effect of quercetin on VEGF-α and VEGFR-2 mRNA expressions in 3T3-L1 cells.After differentiation induction, the cell monolayer was incubated in post-differentiation medium with 0, 5, 10, or 20 μM quercetin for 24 h. Total RNA was isolated, and real-time PCR was performed. β-actin levels were compared, acting as the equal loading control. Each bar represents the mean ± SE. Comparisons among different concentrations of quercetin that produced significant differences (P < 0.05) are indicated by different letters above each bar.VEGF, vascular endothelial growth factor; VEGFR, vascular endothelial growth factor receptor; PCR, polymerase chain reaction.

• 

  Fig. 5 Effect of quercetin on MMP-2 and MMP-9 mRNA expressions in 3T3-L1 cells.After differentiation induction, the cell monolayer was incubated in post-differentiation medium with 0, 5, 10, or 20 μM quercetin for 24 h. Total RNA was isolated, and real-time PCR was performed. β-actin levels were compared, acting as the equal loading control. Each bar represents the mean ± SE. Comparisons among different concentrations of quercetin that resulted in significant differences (P < 0.05) are indicated by different letters above each bar.MMP, matrix metalloproteinase; PCR, polymerase chain reaction.

• 

  Fig. 6 Effect of quercetin on activity of MMP-2 and MMP-9 in 3T3-L1 cells.After differentiation induction, the cell monolayer was incubated in post-differentiation medium with 0, 5, 10, or 20 μM quercetin for 24 h. The MMP-2 and MMP-9 activity levels were estimated by colorimetric assay. Each bar represents the mean ± SE. Comparisons among different concentrations of quercetin that produced significant differences (P < 0.05) are indicated by different letters above each bar.MMP, matrix metalloproteinase; PCR, polymerase chain reaction.

• 

  Fig. 7 Effect of quercetin on concentrations of MMP-2 and MMP-9 in 3T3-L1 cells.After differentiation induction, as described in Methods, the cell monolayer was incubated in post-differentiation medium with 0, 5, 10, or 20 μM quercetin for 24 h. The MMP-2 and MMP-9 concentrations were estimated by using ELISA kits. Each bar represents the mean ± SE. Comparisons among different concentrations of quercetin that resulted in significant differences (P < 0.05) are indicated by different letters above each bar.MMP, matrix metalloproteinase; PCR, polymerase chain reaction.

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