【摘要】：正Breast cancer is one of the most common cancers among women in the world.While early detection is continuing to improve the survival rate,the prognosis is poor after the tumor has metastasized.Hence,it is urgent to identify the genes that drive breast cancer metastasis and develop new therapies.Caveolin-1,a 21-24 kD membrane protein,is an essential structural molecule of Caveolae,participating in Caveolae formation,cell trafficking, cholesterol homeostasis,signal transduction,and Caveolin-1 could inhibit the anchorage-independent cell growth. To study the relationship between Caveolin-1 and metastasis in breast cancer,we developed MCF10A-ST1 and MCF10A~(CE) cell lines which relatively express 30%and 15%of Caveolin-1 compared with mammary epithelial cell (MCF10A) by means of gene trapping and RNAi technology.Then we examined the expression of ERαand associated proteins.ERα36 is a novel variation of ERα,predominantly localized on the plasma membrane which is beneficial to the transduction of estrogen- and antiestrogen-dependent membrane-initiated mitogenic signaling to mediate cell growth.The results indicated that the expression of ERα66 and ERα36 were up-regulated,and the phosphorylation of ERK1/2 was increased in MCF10A~(CE) cell.MCF10A,MCF10A~(CE) and MDA-MB-231 were tested by microarray assay of the estrogen receptor signaling,wounded healing assay,cell adhesion assay,and matrigel invasion assay.Immunocytochemistry assay was used to detect the location of MMP-2 in MCF10A~(CE) cell.Then, MCFIOA,MCF10A-ST1,MCF10A~(CE) and MCF-7 as well as estrogen treated group,estrogen recepter inhibitor (ICI182,780) treated group and association treated group were cultured in monolayer.Western blot,RT-PCR and gelatin zymography detection were used.The specific estrogen receptor signaling microarray assay showed the expression of adhesion related proteins were significantly down-regulated in MCF10A~(CE) cell.Wounded healing assay showed that migrant distance was different in the four cell lines,50%in MCF10A~(CE) and 30%in MCF10A-ST1 compared with MCF-7,while little in MCF10A.Both adhesion and invasion ability of MCF10A~(CE) cell were significantly higher than MCF10A cell,but lower than MDA-MB-231 cell.In MCF10A-ST1 and MCF10A~(CE) cell lines,the expression of MMP-2 in wounded healing group showed an increasing tendency,while the expression of E-cadherin decreased.With the gene silencing of Caveolin-1,the expression of ERα66 and ERα36 were up-regulated. The activity of MMP-2 and MMP-9 were high in MDA-MB-231,MCF10A-ST1 and MCF10A~(CE) cell lines,while almost no activity was detected in MCF10A cell,and the enzyme activity was almost suppressed with the treatment of ICI 182,780.RT-PCR showed that tomor metastasis related gene,VEGF,β-catenin,MMP-1, MMP-2,MMP-9,CD147 were sigificantly high in MCF10A~(CE) cells,while the expression of E-cadherin was low, ICI 182,780 was able to inhibit the expression of related genes.In conclusion,we found that the down-regulation of Caveolin-1 promoted cell adhesion,invasion and metastasis,while the inhibition of ERαsignal molecules attenuated the activity and expression of MMP-2,MMP-9 and other metastasis related proteins.Furthermore,Caveolin-1 increased the activation of ERK and Akt.Our data showed that Caveolin-1 might regulate MMP-2 and MMP-9 activity through ER signal pathway.These results indicate that Caveolin-1 plays an important role in mammary tumorigensis and may affect cell metastasis in vitro.Such evidences will greatly advance the progress in prevention and treatment of human breast cancer.