The Effect Of N-3 Polyunsaturated Fatty Acids On Human Breast Cancer Cells And Mechanism

Objectives:To study the effect of n-3 Polyunsaturated fatty acid (PUFAs) on human breast cancer cells and the underlying mechanisms by in vivo and in vitro methods.Methods:(1) Cell death of human breast cancer cells MDA-MB-231 treated with DHA under conditions with and without glucose was detected by flow cytometry (FCM) with PI fluorescence staining. Meanwhile, the morphological changes of cells were observed using inverted microscope.(2)H2DCFDA probe was used to detect ROS level of breast cancer cells treated by DHA and N-acetyl-L-cystein (NAC) alone or in combination.(3)The effect of DHA and NAC on the expression of cleaved-Caspase-3 protein was examined by Western blot.(4) MTT assay was used to investigate the inhibitory effect of DHA and tamoxifen alone or in combination on breast cancer cells MDA-MB-231. (5)fat-1 transgenic/SCID mice was established to investigate the effect of endogenous n-3 PUFAs on MDA-MB-231 cells, the level of PUFAs in mice was analyzed by gas chromatography.Results:(1) DHA could increase the death rate of MDA-MB-231 cells in a concentration-dependent manner under glucose starvation, while DHA cannot do that when 5 mM glucose was added to medium. The FCM with the probe of DCF indicated that DHA dramatically promoted accumulation of ROS (P<0.005) in intracellular. However, after treatment with NAC, the death rate of cells and the ROS level were decreased. (2) Western blot analysis confirmed that co-treated with NAC could partly attenuate the cleavage of Caspase-3 induced by DHA. (3) Both DHA and tamoxifen could inhibit tumor cell proliferation; they also have a synergic effect. (4) The level of n-3 PUFAs in fat-1/SCID mice is higher than that in SCID mice, while the ratio of n-6/n-3 PUFAs in fat-1/SCID mice is lower than that in SCID mice. (5) Human breast cancer cells cannot grow in any of fat-1/SCID mice, but they can develop xenograft tumors in SCID mouse.Conclusions:(1) DHA could promote death rate of glucose-starved MDA-MB-231 cells. (2) DHA induced cells death by accumulation of intracellular ROS, and then activated Caspase-3 pathway. (3) DHA and tamoxifen inhibited MDA-MB-231 cells viability with synergic effect. (4) Breast tumor cells did not grow in fat-1/SCID mice for they are inhibited by endogenous n-3 PUFAs.