| 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)2D3 72 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。 |
| 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 |
| DOI：10.3724/SP.J.1008.2012.00252 |
| 投稿时间：2012-01-07修订日期：2012-02-20 |
| 基金项目: |
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| COX-2/PGE2 pathway is involved in 1,25-dihydroxyvitamin D3-induced downregulation of CYP1B1 in breast cancer cell line MCF-7 |
| YUAN Lei,DENG Hua-yu*,JIANG Rong,DING Song-tao |
(Department of Pathophysiology, Laboratory for Stem Cell and Tissue Engineering, Chongqing Medical University, Chongqing 400016, China
*Corresponding author.) |
| Abstract: |
| 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(P<0.05).1,25(OH)2D3 inhibited the proliferation of MCF-7 cells in a time- and dose-dependent manner (P<0.05). Treatment with 100 nmol/L 1,25(OH)2D3 for 72 h significantly arrested cell cycle in G0/G1 phase (P<0.05), decreased the expression of COX-2 mRNA in MCF-7 cells (P<0.05), decreased PGE2 level in the cell culture supernatant (P<0.01), and down-regulated p-ERK, p-ERα and CYP1B1 protein expression(P<0.05). 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. |
| Key words: cyclooxygenase 2 1,25-dihydroxyvitamin D3 cytochrome P450 1B1 breast neoplasms |
