Fluctuation Of ROS Influences Proliferation And Mediates Inhibition Of Migration By Reducing The Interaction Between DLC1 And CAV-1 In Breast Cancer Cells

Cellular oncogenesis and the development of cancer are very complex life processes. Life activities of intracellular are regulated by the large and complex net of cell signaling pathways. Many underlying mechanisms of these signaling pathways are unclear, further investigation is necessary. The results had been indicated that reactive oxygen species（ROS） maintain the oncogenic phenotype of cancer cells, however, ROS can also cause cell senescence and apoptosis, therefore can function as anti-tumourigenic species. This function of ROS always was associated with oxidative stress. Previous results indicated that it is required to maintain physiological level of ROS stability. The up-regulation of ROS induced DNA-damage and the low level of intracellular ROS means hypoxia. These two conditions both influenced the expression of related proteins and affected on relevant signaling pathway. Besides,ROS can also function as the second messenger of signal pathway. Its modification in experiment in vitro and antioxidant enzymes which including CAT, GSH, SOD play a crucial role in maintain the balance of intracellular ROS in vivo. In our study, H₂O₂ and catalase（CAT） were used to induce the fluctuation of intracellular ROS level.p53 is tumor suppressor gene. This gene mutates or deletes in variety of human cancer cells and it plays a crucial part in regulating cell proliferation and apoptosis. p53 inhibited cell cycle transition by activating p21 which function as CDK kinase inhibitor and induced cycle arrest or apoptosis. Previous experiments showed that the up-regulation of ROS promoted p53 expression, but the effects of ROS fluctuations in cells is unclear. Our study examined the fluctuation of ROS, proliferation, apoptosis and cell cycle distribution, further, we explored the related mechanism.DLC1 which belongs to Rho-GAP family is considered to play a crucial part in cancer. DLC1 inhibited migration of cancer cells by regulating the activity of RhoA. Caveolin-1（CAV-1） belongs to Caveolin family and it is the primary structure protein of the caveolin. Studies revealed that CAV-1 was related with the cancer migration and invasion in several of cancers. But its effects on cell migration have not reached a conclusion. Prior reports indicated that function of DLC1 and CAV-1 was associated with ROS, and the interaction between DLC1 and CAV-1 was related with cell migration, but the mechanism is still indistinct. Therefore, our experiments explored effects of ROS on DLC1 and CAV-1 expression and on the interaction of DLC1 and CAV-1, and explored the relationship between cell migration and the interaction.The results as fellowing:1. The treatment by exogenous H₂O₂ and CAT for 24 h induced fluctuation of intracellular ROS level in cells MCF-7 and MDA-MB-231.2. Up-regulation of intracellular ROS induced by H₂O₂ inhibited the proliferation rate in a dose-dependent manner, and it also induced cell apoptosis in breast cancer cells. Down-regulation of intracellular ROS induced by CAT did not significantly influence cell proliferation and apoptosis.3. Up-regulation of intracellular ROS level incerased expression of p53 and induced G1 phase cycle-arrest. Down-regulation of ROS did not significantly regulate p53 expression and cell cycle distribution.4. Fluctuation of intracellular ROS level reduced migration index of breast cancer cells significantly.5. ROS fluctuations effected on expression of DLC1 and CAV-1 and inhibited the interaction between DLC1 and CAV-1.The results showed that Up-regulation of intracellular ROS inhibited the expression of p53, induced cell cycle arrest and further inhibited cell proliferation. Down-regulation of ROS did not decrease the cell proliferation rate. Up-regulation and down-regulation of intracellular ROS both decreased migration index significantly by regulating expression of DLC1 and CAV-1, and inhibiting the interaction between DLC1 and CAV-1 in breast cancer cells.