1,25-二羟基维生素D_3对乳腺癌MCF-7细胞CYP1B1表达的作用和COX-2/PGE2通路机制  

COX-2/PGE2 pathway is involved in 1,25-dihydroxyvitamin D_3-induced downregulation of CYP1B1 in breast cancer cell line MCF-7

作　　者：袁磊[1] 邓华瑜[1] 姜蓉[1] 丁嵩涛[1] 

机构地区：[1]重庆医科大学病理生理学教研室,干细胞与组织工程研究室,重庆400016

出　　处：《第二军医大学学报》2012年第3期252-258,共7页Academic Journal of Second Military Medical University

摘　　要：目的分析COX-2、p-ERα、CYP1B1在人乳腺癌组织和ERα阳性表达人乳腺癌细胞系MCF-7中的表达规律,探讨1,25-二羟基维生素D3[1,25(OH)2D3]在人乳腺癌细胞MCF-7中通过COX-2/PGE2通路对CYP1B1表达和细胞增殖、细胞周期转化的影响及作用机制。方法用免疫组织化学法检测42例人乳腺癌组织中COX-2、p-ERα、CYP1B1的表达,并分析其相关性;MTT法检测1,25(OH)2D3对MCF-7细胞增殖的影响,并确定后续实验药物浓度;流式细胞术检测细胞周期;RT-PCR检测MCF-7细胞COX-2 mRNA水平;ELISA法检测细胞培养上清液中PGE2水平;蛋白质印迹法检测MCF-7细胞COX-2、p-ERK、p-ERα、CYP1B1蛋白水平;免疫细胞荧光检测COX-2、p-ERα、CYP1B1蛋白在MCF-7中的原位表达。结果在人乳腺癌组织中COX-2、p-ERα、CYP1B1蛋白呈阳性表达,两两之间均呈正相关性(P<0.05)。1,25(OH)2D3对MCF-7细胞增殖具有抑制作用,且呈时间和剂量依赖性(P<0.05)。应用100 nmol/L 1,25(OH)2D372 h后,MCF-7细胞周期阻滞在G0/G1期(P<0.05);细胞COX-2 mRNA表达减少(P<0.05);细胞培养上清中PGE2水平降低(P<0.01);COX-2、p-ERK、p-ERα及CYP1B1蛋白表达均减少(P<0.05)。结论在乳腺癌中,COX-2/PGE2通路对CYP1B1的表达具有正向调控作用。1,25(OH)2D3可通过抑制COX-2/PGE2通路减少CYP1B1的表达,对乳腺癌细胞MCF-7的增殖产生抑制作用。Objective To analyze the expressions of COX-2,p-ERα and CYP1B1 in human breast cancer tissues and ERα-positive human breast cancer cell line MCF-7,and to investigate the influence of 1,25-dihydroxyvitamin D3 [1,25（OH）2D3] on proliferation,cell cycle transformation,and CYP1B1 protein expression in MCF-7 cells.Methods Immunohistochemical method was applied to examine the expressions of COX-2,p-ERα and CYP1B1 protein in 42 breast cancer tissues,and their association was analyzed.The effects of 1,25（OH）2D3 on MCF-7 cell proliferation was investigated by MTT assay and the optimal concentration of 1,25（OH）2D3 was determined for the following experiment.The cell cycle was analyzed by flow cytometry and COX-2 mRNA expression in MCF-7 cells was measured by RT-PCR.PGE2 level was detected by ELISA in the culture supernatant.The expression of COX-2,p-ERK,p-ERα and CYP1B1 protein was determined by Western blotting analysis and the distribution of COX-2,p-ERα and CYP1B1 expression in MCF-7 cells was examined by immune cell fluorescence.Results COX-2,p-ERα and CYP1B1 protein were positive in human breast cancer tissues and their expressions were positively correlated with each other（P0.05）.1,25（OH）2D3 inhibited the proliferation of MCF-7 cells in a time-and dose-dependent manner（P0.05）.Treatment with 100 nmol/L 1,25（OH）2D3 for 72 h significantly arrested cell cycle in G0/G1 phase（P0.05）,decreased the expression of COX-2 mRNA in MCF-7 cells（P0.05）,decreased PGE2 level in the cell culture supernatant（P0.01）,and down-regulated p-ERK,p-ERα and CYP1B1 protein expression（P0.05）.Conclusion COX-2/PGE2 pathway plays a positive role in regulating CYP1B1 expression in breast cancer.1,25（OH）2D3 may inhibit the growth of MCF-7 cells and down-regulate CYP1B1 through COX-2/PGE2 pathway.

关 键 词：环氧化酶2 1 25-二羟基维生素D3 细胞色素P4501B1 乳腺肿瘤 

分 类 号：R737.9[医药卫生—肿瘤]