Effects Of Blocking Estrogen And Its α Signal Receptor For BCG Treatment In Bladder Cancer

Objective: To detect the expression levels in different superficial bladder cancer tissues of estrogen receptors ERα. Explore the mechanism of the inhibition of BCG induced by estrogen in superficial bladder cancer recurrence. Established model of superficial bladder cancer, to investigate the effects of estrogen in BCG-inhibition of bladder tumor in vivo.Methods:1. Using immunohistochemical staining to detect the expression of ERα in the same stage and grade with recurrent or non-recurrent superficial bladder tumor tissues.2. In vitro,T24 cell lines with stable expression of ERα and wild type human bladder transitional carcinoma 253 J cells lines were used. After 48 h incubation, Detected the expression of ERα in both cell lines. After treated with estradiol, estradiol antagonists ICI182.780 and TAM,the cells continued treating with BCG,then detect the expression of BCG and the integrin α5β1. Verify whether estrogen affects BCG adhesion process.Another part of the cells following the above process, human monocyte-macrophage cell line THP-1 were invasive experiment to test whether estrogen influence the recruitment of THP-1 cell to kill tumor cells.3.In vivo,using N-butyl-N-(4-hydroxybutyl) nitrosamine(OH-BBN) induced tumors in FVB female mice. Mice were divided into control group, BCG group, estrogen and estrogen antagonists and BCG antagonist group, the experimental group to each group of mice were instillation BCG, the experimental group and the control group of mice survival, tumor tissue taken after pathological section, HE staining, detection of Brd U proliferation clarify its case, the detection F4/80 to clarify its macrophage infiltration.Results:1. Expression of estrogen receptor α in the recurrence of superficial bladder cancer is higher than the postoperative recurrence of superficial bladder cancer.2. T24ERα cells expressing ERα protein compared with wild-type T24 cells, the expression of ERα in 253 J cells is low; estradiol suppress BCG attachment/internalization into the bladder cancer cell; estradiol suppression of bladder integrin-α5β1m RNA expression in cancer cells, estrogen antagonist ICI182.780 make the reverse efforts; In the two cell lines confirmed the antagonist of integrin-α5β1 can inhibit BCG attachment/internalization induced by ICI182.780; The two cell lines co-cultured with THP-1 cells proved BCG can promote the migration of THP-1 cells,and estradiol inhibit the progression; BCG,E2+BCG,E2+ICI182.780+BCG,add to the two cell lines,conditioned medium and the cells obtained. The IL-6 m RNA levels and IL-6 protein levels were measured, BCG can enhance bladder cancer cells to secrete more IL-6; conditioned medium of the four groups were treated THP-1 cells, ELISA show expression of TNF-α is highest in ICI182.780+BCG group,BCG group is the second; The two cell lines cultured with four conditions media above, the results show THP-1+BCG+ ICI182.780 group exhibits strongest inhibition of bladder cancer.3. HE staining showed that the control group and the group ICI182.780 appears papilloma, BCG group had urothelial dysplasia, BCG+ICI182.780 urothelial is normal;BCG+ICI182.780 group has the minimal number of Brd U positive cells,BCG group is the second,control group and ICI182.780 group are similar has the most. BCG+ICI182.780 group has the most of F4/80 positive cells,BCG group is the second, followed by the control group and the ICI group; After treatment, BCG+ICI182.780 mice survival the longest time, the control group and the ICI182.780 group mice has the shortest survival time.Conclusion: The high expression of estrogen receptor α in the recurrence of superficial bladder cancer, BCG and estradiol can inhibit tumor cell adhesion and enter the cell thus affecting the efficacy of BCG. In addition, estradiol can inhibit the role of BCG on recruitment of macrophages. Thus, estrogen-estrogen receptor signaling pathway may be one of the factors leading to reduced efficacy of BCG.