Estrogen Regulate E-cadherin And A-actinin-4to Increase The Metastasis Of ShRNA ERa/β SKOV3via The Activation Of PI3K/Akt Signaling Transduction Pathway

Ovarian cancers, the third most common gynecological malignancy, and the leading cause of death from gynecological cancer, are highly lethal because of the occurrence of occult metastasis within the peritoneal cavity and the fact that they are often detected at an advanced stage, when curative therapy is ineffective. Although operation, radiotherapy and chemotherapy are ever improving, the5year survival rate is still30-50%. Therefore, great efforts have been made to improve the prognosis of this disease.The ovary produces sex steroids and is also the target organ. More and more researchers committed to investigating the correlation between estrogen and ovarian cancer metastasis. Previous research suggests that estrogen could regulate ovarian cancer cell migration and invasion by affecting metastasis-related genes. Our previous study demonstrated estrogen stimulated the migrations of ovarian cancer cell via PI3K/AKT pathway. The purpose of the current study was to explore the role of PI3K/AKT pathway on ovarian cancer cell migration initiated by estrogen after knockdown of ER a/(3.These experiments were divided into4parts, as follows:1. In vitro experiment:using the SKOV3and shRNA ERa/β SKOV3ovarian cancer cell lines, we sought to understand the mechanisms associated with estrogen, and the PI3K/AKT pathway on metastasis, and the mechanism involved in the regulation of tumor metastasis-related genes (E-cadherin and a-actinin-4) by steroids via the signal transduction pathway;2. In vivo experiment:using the athymic nude mice inoculated i.p. with shRNA ERa/β SKOV3, We evaluated effects of estrogen and LY294002, a potent inhibitor of PI3K, on tumor growth.To sum up, it helps us well understanding of the relationship between estrogen induction of cell metastasis and the PI3K/AKT signaling pathway to find new approaches for ovarian cancer treatment.Ⅰ. In vitro experimentEstrogen to increase the metastasis of shRNA ERa/β SKOV3via the activation of PI3K/Akt signaling transduction pathway.ObjectiveTo explore the impact of estrogen on PI3K/Akt signaling transduction pathway in shRNA ERa/β SKOV3cells, and the relationship between the pathway and the expression of E-cadherin and a-actinin-4.Methods Changes of pAKT protein level of shRNA ERa/β SKOV3after treated with estrogen were detected by Western blot. Transwell chambers were used in the evaluation of the invasion ability of cancer cells after treated with estrogen and LY294002. The expression of E-cadherin and a-actinin-4were detected by Western blot after treated with estrogen and LY294002.Results17β-estrogen increased the expression of pAKT but didnot change the level of AKT in shRNA ERa/β SKOV3. When the PI3K/Akt pathway was suppressed, estrogen’s impact on promoting the metastasis and invasion of the cancer cells, downregulation on E-cadherin, upregulation on a-actinin-4in cytoplasm was repressed.ConclusionThe effect of estrogen on metastasis in shRNA ERa/β SKOV3was regulated by the PI3K/Akt pathway and targeting genes. E-cadherin and a-actinin-4are the potential targeting genes of PI3K/Akt pathway.Ⅱ. In vivo experimentThe effect of estrogen and LY294002on transplanted tumors of nude mice.ObjectiveTo investigate the regulation of estrogen and LY294002on transplanted tumors of nude mice.MethodsShRNA ERa/β SKOV3were cultured and inoculated i.p. into20nude mice. The nude mice were divided into4groups after two weeks:control (physiological saline), estrogen, LY294002, estrogen+LY294002. Compare the weight of nude mice and the transplanted tumors, the expression of pAKT、AKT、E-cadherin and a-actinin-4in transplanted tumors in each group.ResultsThere was no difference on the weight of nude mice in4groups. Compared with control, the tumor weights in estrogen, estrogen+LY294002group were much heavier(P<0.05), while much lighter in LY294002group(P<0.05). Compared with LY294002group, the tumor weights in estrogen+LY294002group were much heavier(P<0.05). In estrogen group, E-cadherin was downregulated and cytoplasmic a-actinin-4upregulated. While in LY294002group, E-cadherin and nucleus a-actinin-4were upregulated.ConclusionEstrogen can promote the growth and proliferation of transplanted tumors in nude mice meanwhile LY294002can inhibit the growth and proliferation of transplanted tumors in nude mice. To sum up,17β-estrogen activated the PI3K/Akt signaling transduction pathway to regulate the expression of E-cadherin and a-actinin-4, which promoted metastasis and invasion of tumor cells. Meanwhile estrogen can promote the growth and proliferation of transplanted tumors in nude mice, while LY294002can repress it.