The Regulatory Effect And Mechanism Of Caveolin-1on The Cell Growth, Migration And Invasion, And17β-estradiol-mediated Autophagy And Apoptosis In Human Breast Cancer BT474Cells

ObjectiveTo investigate the effect and mechanism of caveolin-1in the regulation of cell growth, migration and invsion, and E2-mediated autophagy and apoptosis of breast cancer BT474cells, and to discuss if caveolin-1functions as a tumor promoter in breast cancer cells.MethodsPart1:We used realtime-PCR assay to detect the the expression of caveolin-1in breast cancer BT474, MCF7, SK-BR-3cell lines. Caveolin-1siRNA was used to knock down the caveolin-1expression in BT474and MCF-7cells. And then realtime-PCR, western blot and immunofluorescence staining were performed to examine the knockdown efficiency of caveolin-1siRNA in BT474and MCF-7cells. In addition, CCK-8assay, transwell assay, colony formation analysis and flow cytometry analysis were used, respectively, to test the effect of caveolin-1knockdown on the cell growth, resistance to DOX, migration and invasion ability, colony formation efficiency and cell cycle of BT474and MCF-7cells. Furthermore, western blot analysis was used to detect the effect of caveolin-1knockdown on the activation of ERK1/2signal pathway, the expression of MMP-1,-2,-9, and cyclin D1, c-Fos, β-catenin and E-cadherin of BT474cells.Part2:CCK-8assay and the transwell analysis were used to measure the impact of E2on the cell growth, migration and invasion ability of BT474cells, and the effect of cavoelin-1knockdown on the E2-mediated cell growth, migration and invasion. And then western blot assay was performed to detect the effect of E2on the expression caveolin-1, HMGB1and autophagy-related proteins (LC3-Ⅱ, Atg12/5, Beclin-1). Immnunofluorescence staining was used to directly detect the LC3punctate and MDC staining was used to test the formatin of autophagosome in BT474cells. Moreover, the effect of caveolin-1or HMGB1knockdown on the expression of autophagy-related proteins, LC3punctate and autophagosome formation was tested via western blot analysis, immunofluorescence analysis and MDC staining. In addition, flow cytometry for apoptotic rate, immnuofluorescence staining for cleaved-caspase3were used to examine the influence of E2, caveolin-1knockdown, or HMGB1knockdown on the apoptotic rate and cleaved-caspase3punctate formation. ResultsPart1:Caveolin-1was more highly expressed in BT474cells than MCF-7and SK-BR-3cells. The efficiency of caveolin-1knockdown in BT474and MCF-7cells using siRNA technique was over60%. Caveolin-1knockdown inhibited the cell growth, migration and invasion ability, and decreased the resistance to DOX treatment, and also attenuated the efficiency of colony formation in BT474cells. Caveolin-1knockdown also increased the population of G0/G1phase and decreased the population of S phase in BT474cells. However, caveolin-1knockdown increased the cell growth, migration and invasion ability, and promoted the colony formation in MCF-7cells. Moreover, caveolin-1knockdown decreased the population of GO/G1phase and increased the population of S phase in MCF-7cells. Furthermore, caveolin-1knockdown suppressed the activation of ERK1/2pathway, down-regulated the protein expression of MMP-1,-2,-9and cyclin D1, c-Fos and β-catenin, and promoted the protein expression of E-cadherin in BT474cells.Part2:Knockdown of caveolin-1inhibited the E2-indcued cell growth, migration and invasion of BT474cells. E2promoted the protein expression of caveolin-1, HMGB1and autophagy-related proteins (LC3-Ⅱ, Atg12/5, Beclin-1), and the LC3punctate and autophagosome formation in BT474cells. Caveolin-1or HMGB1knockdown inhibited the expression of E2-induced LC3-Ⅱ, as well as E2-induced formation of LC3punctate and autophagosome. E2inhitited the apoptotic rate and cleaved-caspase3punctate formation in BT474cells. Caveolin-1or HMGB1knockdown augmented the apoptotic rate and cleaved-caspase3punctate formation and affected the E2-inhibited apooptosis in BT474cells.ConclusionCaveolin-1is involved in the regulation of cell growth, migration and invasion ability of breast cancer cells. Caveolin-1acts as a tumor promoter in BT474cells, which is different from the response in MCF-7cells. Moreover, caveoin-1functions as an key regulator of E2-mediated autophagy and apoptosis in BT474cells, which also contributes to the E2-induced cell proliferation. Our results may provide more evidence for the understanding the mechanism of caveolin-1in the tumorgenesis of breast cancer caner cells.