ERβ增加胃癌细胞氟尿嘧啶化疗敏感性的机制研究

Estrogen Receptor β enhanced the chemotherapy sensitivity of fluorouracil in gastric cancer cells by arresting cell cycle and inducing thymidine phosphorylase expression

目的:探索雌激素受体β(ERβ)表达与胃癌细胞氟尿嘧啶(5-FU)化疗敏感性的相关性及可能的机制。方法:IHC观察胃癌及癌旁组织中ERα、ERβ表达,q PCR和WB观察胃癌细胞及正常肠上皮细胞ERβ表达。以生理盐水为对照组,不同浓度的5-FU为实验组处理ERβ表达水平不同的胃癌细胞,WST-1观察ERβ表达对氟尿嘧啶抑制胃癌细胞增殖的影响。以转染空白质粒为对照组,转染ERβ表达载体为实验组,流式细胞术和免疫印迹观察ERβ表达对胃癌细胞周期的影响。结果:胃癌组织中ERβ高表达,ERα不表达;胃癌细胞KatoⅢ、AGS中ERβ高表达,且以KatoⅢ表达更高。5-FU对AGS和KatoⅢ细胞增殖的抑制作用呈浓度依赖性,KatoⅢ细胞对5-FU更为敏感(P <0. 05)。过表达ERβ使AGS细胞对5-FU的敏感性升高(P <0. 05);敲低ERβ可降低KatoⅢ对5-FU的敏感性(P <0. 05)。过表达ERβ可抑制周期蛋白D1、E的表达,促进胸苷磷酸化酶的表达,阻滞细胞于G1期。结论:ERβ可通过调节细胞周期、促进TP来改变胃癌细胞对5-FU化疗敏感性。

Objective: To access the relationship between Estrogen Receptor β(ERβ) expression and the chemotherapy sensitivity of fluorouracil (5-FU) in gastric cancer (GC). Methods: Immunohistochemical (IHC) was performed to observe the expression of Estrogen Receptor α(ERα) and ERβ in GC patient with surgery (GC tissue and adjacent normal tissue), qPCR and western blotting were used to investigate expression of ERβ in GC and normal epithelial cells. Superoxide Dismutase (SOD) assay (WST-1) was performed to assess the proliferation of different ERβ expression GC cells (KatoⅢ and AGS) with different dosage of 5-FU treatment, compared with normal saline as negative control. Flow cytometry and western blotting were used to detect cell cycle in different ERβ expression GC cells. Results: ERβ was highly expressed in the GC tissue and cell lines, while ERα expression was negative in GC tissue. 5-FU inhibited the proliferation of AGS and KatoⅢ as dose-dependent manner, KatoⅢ was more sensitive to 5-FU compared with AGS ( P < 0.05 ). Overexpression of ERβ in AGS increased the sensitivity of 5-FU compared with control plasmid ( P <0.05), knock-down of ERβ by siRNA in KatoⅢ decreased the sensitivity of 5-FU compared with control siRNA ( P <0.05). ERβ overexpression inhibited the expression of cyclin D1 and cyclin E, leading to the cell cycle arrested in G1 phase. Furthermore, ERβ induced expression of thymidine phosphorylase (TP) and enhanced the sensitivity of 5-FU in AGS. Conclusions: ERβ increased the chemosensitivity of 5-FU in GC cell lines by inducing cell cycle arrest and TP expression .