Nutr Res Pract.  2012 Aug;6(4):294-300.

Effect of resveratrol on the metastasis of 4T1 mouse breast cancer cells in vitro and in vivo

• Lee HS
• Ha AW
• Kim WK

Abstract

We investigated the effects of resveratrol on metastasis in in vitro and in vivo systems. 4T1 cells were cultured in the presence of various concentrations (0-30 micromol/L) of resveratrol. For experimental metastasis, BALB/c mice were injected intravenously with 4T1 cells in the tail vein, and were orally administered various concentrations (0, 100, or 200 mg/kg Body weight) of resveratrol for 21 days. After resveratrol treatment, cell adhesion, wound migration, invasion, and MMP-9 activity were significantly decreased in a dose-dependent manner in 4T1 cells (P < 0.05). The numbers of pulmonary nodules were significantly decreased in mice fed the resveratrol (P < 0.05). The plasma MMP-9 activity was decreased in response to treatment with resveratrol in mice (P < 0.05). We conclude that resveratrol inhibits cancer metastasis both in vitro and in vivo, and this inhibition is likely due to the decrease in MMP-9 activity caused by resveratrol.

Keyword

Resveratrol; metastasis; matrix metalloproteinase-9; 4T1 breast cancer cell; BALB/c mice

MeSH Terms

Animals
Breast
Breast Neoplasms
Cell Adhesion
Matrix Metalloproteinase 9
Mice
Neoplasm Metastasis
Plasma
Stilbenes
Veins
Matrix Metalloproteinase 9
Stilbenes

Figure

• 

  Fig. 1 The effects of resveratrol on the adhesion, motility, and invasion in 4T1 cells. (A) After 8 × 105 cells/mL suspended in DMEM/F12 medium containing 0, 10, 20, or 30 µmol/L resveratrol were plated in each well of a 96 well fibronectin-coated plate for 1 h, the medium was gently removed and the attached cells were subjected to an MTT assay. (B) Cells were plated in a 12-well plate at a density 5 × 105 cells/well in DMEM/F12 supplemented with 10% FBS. Confluent monolayers were then wounded and subsequently incubated in serum free medium in the presence of 0, 10, 20, or 30 µmol/L resveratrol. The cells were then photographed under a phase contrast microscope at 0, 12, 24, and 48 h after being wounded. (C) Cells were cultured in the presence of various concentrations of resveratrol for 8 h in a Boyden chamber. a) Microphotography of cells treated with resveratrol. b) Quantitative analysis of the cell motility assay. (D) Cells were cultured in the presence of various concentrations of resveratrol for 8 h in an invasion chamber a) Microphotography of cells treated with resveratrol. b) Quantitative analysis of the invasion assay. Each bar represents the mean ± SD of three independent experiments. Significant differences (P < 0.05) among groups are indicated by different letters above each bar.

• 

  Fig. 2 The effects of resveratrol on MMP-9 activity and mRNA expression in 4T1 cells. Resveratrol decreases activity and mRNA expression of MMP-9. (A) The 4T1 cells were plated in 6 well plates at a density 1 × 106 cells/well, the monolayers were then incubated in serum-free medium containing 0, 10, 20, and 30 µmol/L resveratrol for 16 h. Next, the medium was collected and concentrated for zymography. a) Photographs of MMP-9 bands. b) Densitometric analysis of western blots. (B) For RT-PCR, 4T1 cells were treated with resveratrol and the samples were then separated on a 1% agarose gel. a) Photographs of chemiluminiscent evaluation of the RT-PCR bands. b) Quantitative analysis of RT-PCR. The relative abundance of each band was estimated by densitomertric analysis. Each bar represents the mean ± SD of three independent experiments. Significant differences (P < 0.05) among groups are indicated by different letters above each bar.

• 

  Fig. 3 Resveratrol inhibits experimental metastasis in Balb/c mice. 4T1 cells (2 × 105 cells/0.1 mL) were injected into the lateral tail vein of 40 Balb/c mice. Resveratrol was then administered orally each day at a dose of 100 mg/kg BW (CanR100) or 200 mg/kg BW (CanR200). The colony numbers decreased with the amount of resveratrol increased. With no resveratrol, there were 42.6 ± 11.5 colonies. The number of colony were 29.3 ± 6.1 with CanR100 and 15.2 ± 8.3 with CanR200 (A). MMP-9 activity was measured in plasma (B). a) Photographs of MMP-9 bands. b) Densitometric analysis of western blots. Each bar represents the mean ± SD of three independent experiments. Significant differences (P < 0.05) among groups are indicated by different letters above each bar.

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