受体型蛋白酪氨酸磷酸酶O基因甲基化与乳腺癌细胞株化疗敏感性的关系  

Correlation of methylation of protein tyrosine phosphatase receptor-type O gene with the chemosensitivity in breast cancer cells

作　　者：李少英[1] 吴华聪[1] 王辉林[1] 王维[1] 陈静[1] 

机构地区：[1]深圳市宝安区妇幼保健院乳腺科,518133

出　　处：《中华乳腺病杂志（电子版）》2014年第5期28-33,共6页Chinese Journal of Breast Disease(Electronic Edition)

摘　　要：目的探讨受体型蛋白酪氨酸磷酸酶O(PTPRO)基因不同甲基化状态的乳腺癌细胞株对紫杉醇的敏感性。方法应用甲基化特异性PCR,检测MCF-7、MDA-MB-231和Hs578t乳腺癌细胞株中PTPRO基因启动子Cp G岛甲基化状态,并用RT-PCR法检测PTPRO mRNA表达。采用MTT法检测MCF-7、MDA-MB-231和Hs578t乳腺癌细胞株对0.000 6、0.003 0、0.015 0、0.075 0、0.375 0、1.875 0、9.350 0、46.750 0、234.000 0 mmol/L紫杉醇的敏感性,以及对细胞株进行去甲基化实验后紫杉醇抑制率的改变。两组间均数比较采用配对t检验;在检验方差齐性的前提下,多组间均数比较用单因素方差分析,组间两两比较采用LSD检验。结果乳腺癌MCF-7、MDA-MB-231细胞株中PTPRO基因甲基化,而乳腺癌Hs578t细胞株中PTPRO基因无甲基化。用紫杉醇处理细胞株后再检测PTPRO启动子甲基化情况,结果显示MCF-7、MDA-MB-231细胞株PTPRO基因甲基化率明显降低[(0.861±0.109)比(0.037±0.019),t=23.326,P=0.000;(0.758±0.114)比(0.086±0.010),t=16.109,P=0.000]。并且,MCF-7、MDA-MB-231细胞株经5-杂氮脱氧胞嘧啶(DAC)处理后,PTPRO mRNA表达水平明显高于DAC处理前[(0.033±0.006)比(0.166±0.016),t=28.338,P=0.000;(0.052±0.006)比(0.587±0.087),t=17.257,P=0.000],其表达水平与启动子Cp G岛甲基化状态有关。紫杉醇对MDA-MB-231、MCF-7及Hs578t细胞株的半数抑制浓度(IC50)分别为8.52、5.87和4.52 mmol/L。不同浓度紫杉醇对Hs578t细胞株的抑制率均比MCF-7和MDA-MB-231细胞株高(F=129.93、182.40、46.26、49.26、33.60、80.31、160.05、69.96、79.98,P均=0.000)。去甲基化实验显示:DAC处理MCF-7和MDA-MB-231乳腺癌细胞株后,不同浓度紫杉醇对细胞株的抑制率与处理前相比,差异均有统计学意义(MCF-7:t=14.195、8.328、7.042、6.385、5.450、8.557、4.788、13.351、11.887,P均<0.050;MDA-MB-231:t=16.324、30.443、9.177、12.694、9.528、11.912、10.260、18.109、3.754,P均<0.050)。结论 PTPRO基因甲基化是乳腺Objective To determine the role of protein tyrosine phosphatase receptor-type O（PTPRO） gene methylation as a potential molecular biomarker in the treatment of breast cancer and its association with the sensitivity to paclitaxel in the breast cancer cell lines. Methods Methylation-specific PCR was used to detect the methylation status of PTPRO gene promoter CpG island in three breast cancer cell lines MCF-7, MDA-MB- 231 and Hs578t. RT-PCR was used to measure PTPRO mRNA expression. MTT assay was applied to evaluate the sensitivity of MCF-7, MDA-MB-231 and Hs578 cells to paclitaxel under different concentrations, i. e. , 0. 000 6,0. 003 0,0. 015 0,0. 075 0,0. 375 0,1. 875 0,9. 350 0,46.750 0, 234.000 0 mmol/L. The changes of inhibition rate of paclitaxel in cell lines after demethylation were analyzed as well. The mean comparison between two groups was performed using paired t test; after determining the homogeneity of variance, mean comparison among multiple groups were performed using One-Way ANOVA; pairwise comparison using LSD test. Results PTPRO gene was methylated in MCF-7 and MDA-MB-231 cell lines, but unmethylated in Hs578t cell lines. After treated with paclitaxel, the methylation rate of PTPRO in MCF-7 and MDA-MB-231 cell lines was significantly decreased [ （ 0. 861 ±0. 109 ） vs （ 0. 037 ±0. 019 ）, t = 23. 326, P = 0. 000 ; （ 0. 75± 0. 114）vs（0. 086±0. 010） ,t= 16. 109,P=0. 000]. Moreover, PTPRO mRNA expression level in MCF-7 and MDA-MB-231 cell lines was significantly higher after DAC treatment [ （0. 033±0. 006） vs （0. 166±0. 016）, t=28. 338, P=0. 000; （0. 052±0. 006） vs （0. 587±0. 087）, t= 17. 257, P=0. 000], which was correlated with the methylation status of CpG islands. IC50 of paclitaxel on MCF-7, MDA-MB-231 and Hs578 breast cell lines was 8.52, 5.87 and 4. 52 mmol/L, respectively. Hs578t cell line was significantly more sensitive to paclitaxel at different concentrations compared with the other two cell lines （F = 129.93, 182.40, 46. 26, 49.26, 33.60, 80

关 键 词：编码膜结合受体型蛋白酪氨酸磷酸酶O 甲基化 乳腺肿瘤 化疗敏感性 

分 类 号：R737.9[医药卫生—肿瘤]