Nutr Res Pract.  2009 Jun;3(2):77-83.

Effects of retinoic acid isomers on apoptosis and enzymatic antioxidant system in human breast cancer cells

• Hong TK
• Lee-Kim YC

Abstract

Retinoic acids (RAs) modulate growth, differentiation, and apoptosis in normal, pre-malignant & malignant cells. In the present study, the effects of RA isomers (all-trans RA, 13-cis RA, and 9-cis RA) on the cell signal transduction of human breast cancer cells have been studied. The relationship between RAs and an enzymatic antioxidant system was also determined. Estrogen-receptor (ER) positive MCF-7 and ER-negative MDA-MB-231 human breast cancer cells were treated with different doses of each RA isomers, all-trans RA, 13-cis RA, or 9-cis RA. Treatment of RA isomers inhibited cell viability and induced apoptosis of MCF-7 cells as a result of increased caspase activity in cytoplasm and cytochrome C released from mitochondria. All-trans RA was the most effective RA isomer in both cell growth inhibition and induction of apoptosis in MCF-7 cells. However, no significant effect of RA isomers was observed on the cell growth or apoptosis in ER-negative MDA-MB-231 cells. In addition, activities of antioxidant enzymes such as catalase and glutathione peroxidase were decreased effectively after treatment of RA in MCF-7 cells, whereas SOD activity was rarely affected. Thus, the present data suggest that all-trans RA is the most potential inducer of apoptosis and modulator of antioxidant enzymes among RA isomers in MCF-7 human breast cancer cells.

Keyword

Breast cancer cells; RA Isomers; apoptosis; antioxidant enzymes

MeSH Terms

Apoptosis
Breast
Breast Neoplasms
Catalase
Cell Survival
Cytochromes c
Cytoplasm
Glutathione Peroxidase
Humans
MCF-7 Cells
Mitochondria
Signal Transduction
Tretinoin
Catalase
Cytochromes c
Glutathione Peroxidase
Tretinoin

Figure

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  Fig. 1 Effect of retinoic acid (RA) isomer concentrations on the growth of human breast cancer MCF-7 cells (A) and MDA-MB-231 cells (B). MCF-7 cells and MDA-MB-231 cells were seeded at 3×104 cells and incubated with various concentrations of RA isomers for 4 days. The viability of Cells were measured by MTT assay. Values are means ± S.D. of three independent experiments. *P<0.05: compared to the control

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  Fig. 2 Effect of retinoic acid (RA) isomer concentrations on apoptosis of human breast cancer MCF-7 cells (A) and MDA-MB-231 cells (B). Apoptotic cells were detected the most at 1 µM all-trans RA. Results are expressed as relative % of control value. Values are means ± S.D. of three independent experiments. *p<0.05: compared to the control group

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  Fig. 3 Effect of retinoic acid (RA) isomers on caspase-3 activity in MCF-7 cells. MCF-7 cells were seeded at 3×104 cells and incubated with 1 µM RA isomers. Caspase-3 activity was measured by a Western blot using caspase cleavage specific antibody.

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  Fig. 4 Effect of all-trans retinoic acid (RA) on caspase inhibiton of MCF-7 cells. MCF-7 cells were seeded at 3×104 cells and incubated with 1 µM all-trans RA. Caspase inhibitor was used by Ac-DEVD-CHO. A: Caspase-3 activity by caspase inhibitor in ER-positive MCF-7 cells. B: Effect of caspase inhibitor on viability of MCF-7 cells. Results are expressed as relative % of control value. Values are means ± S.D. of three independent experiments. *p<0.05, **p<0.01: compared to the control group

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  Fig. 5 Effect of retinoic acid (RA) isomers on cytochrome C release from mitochondria in MCF-7 cells. MCF-7 cells were seeded at 3×104 cells and incubated with 1 µM RA isomers. Released cytochrome C was measured by a Western blot. Actin was a protein marker that sould not change between samples.

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  Fig. 6 Effect of retinoic acid (RA) isomers on caspase -8 (A) & -9 (B) activities in MCF-7 cells. MCF-7 cells were seeded at 3×104 cells and incubated with 1 µM RA isomers. All-trans RA was the most effector of RA isomers. The activities of caspase-8 &-9 were measured by using fluorogenic substrates Ac-IETD-AMC (caspase-8) and Ac-LEHD-PNA (caspase-9). Results are expressed as relative % of activities of caspase-8 &-9. Values are means ± S.D. of three independent experiments. *p<0.05: compared to the control group

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  Fig. 7 Effect of retinoic acid (RA) isomers on antioxidant defense enzymes of MCF-7 cells. MCF-7 cells were seeded at 3×104 cells and incubated with 1 µM RA isomers. SOD activites did not show a significant change, but catalase & GPx activities decreased the most by all-trans RA. Results are expressed as relative % of enzymes' activities. Values are means ± S.D. of three independent experiments. *p<0.05, **p<0.01: compared to the control group

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