内皮素3对恶性黑素瘤A375细胞上皮基质转化的影响  

Effects of endothelin-3 on epithelial-to-mesenchymal transition in a malignant melanoma cell line A375

作　　者：李艳秋[1] 朱里[1] 曾山鹰 王翠彦[1] 孙兰[1] 林云[2] 陈宏翔[1] 黄长征[1] 陈思远[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院皮肤科,武汉430030 [2]华中科技大学同济医学院附属协和医院麻醉科,武汉430030 [3]湖北省中山医院皮肤科

出　　处：《中华皮肤科杂志》2012年第7期501-504,共4页Chinese Journal of Dermatology

基　　金：国家自然科学基金（30671891,30901984,81072232）

摘　　要：目的研究内皮素-3（ET-3）对人恶性黑素瘤（MM）A375细胞上皮基质转化（EMT）的影响。方法体外培养A375细胞，分别设立3组：空白对照组、100nmol／LET-3组、100nmol／LET-3和100txmol／LBQ788（内皮素受体B阻断剂）组。采用Transwell小室检测细胞转移，细胞爬片技术检测细胞形态变化，实时PCR和Western印迹检测上皮基质转化相关分子上皮细胞钙黏蛋白、波形蛋白及转录因子（Twist、Slug）表达情况，使用方差分析及Scheffe法对结果进行分析。结果各干预条件中，与空白对照组比较。ET～3可以促进A375细胞的转移，BQ788可阻断该效应（3组穿膜细胞数分别为4．200±0．837、9．400±0．548、3．400±0．894．F=88．44，P〈0．01）；ET-3可以促进A375细胞由上皮型向成纤维细胞样形态转变，促进A375上皮细胞钙黏蛋白表达下调（3组分别为0．330±0．002、0．280±0．007、0．420±0．008，F=329．98，P〈0．01），波形蛋白表达上调（0．830±0．014、1．160±0．003、0．750±0．030，F=262．94，P〈0．01），而BQ788可阻断这种效应。ET-3可以促进上皮基质转化相关转录因子SlugmRNA（F=376．94，P〈0．01）及TwistmRNA（F=215．62，P〈0．01）及其蛋白水平（FSlug=288．87，P〈0．01；FTwist=156．96，P〈0．05）上的表达上调。结论ET-3／ETRB通过上调波形蛋白，下调上皮细胞钙黏蛋白的表达，并上调转录因子（Twist、Slug）的表达，促进黑素瘤A375细胞上皮基质转化。Objective To explore the role of endothelin-3 （ET-3） on epithelial-to-mesenchymal transition in a malignant melanoma cell line A375. Methods A375 cells were cultured in vitro and classified into 3 groups to be treated with ET-3 at 100 nmol/L （ET-3 group）, co-cultured with ET-3 at 100 nmol/L and endothelin receptor B （ETRB） antagonist BQ788 at 100 μmol/L （ETRB antagonist group）, or to remain untreated （blank control group）. After additional 24-hour culture, Transwell chamber assay was used to detect the invasive capability of A375 cells, real time-PCR to measure the mRNA expressions of Twist and Slug, and Western blot to determine the protein expression of E-cadherin, vimentin, Twist and Slug. The changes in the morphology of A375 cells were observed. Data were assessed by analysis of variance and Scheffe＇s method. Results In the Transwell assay, the number of A431 cells permeating through the basement membrane was 4.200±0.837, 9.400±0.548 and 3.400±0.894 respectively in the blank control group, ET-3 group and ETRB antagonist group （F = 88.44, P 〈 0.01 ）, suggesting that ET-3 could promote the metastasis of A375 cells, while BQ788 could block the promotive effect of ET-3. The epithelial-to-mesenchymal transition was obvious in cells treated with ET-3 alone, but was inapparent in cells treated with ET-3 and BQ788. The ET-3 at 100 nmol/L significantly decreased the protein expression of E-cadherin from 0.33±0.002 （blank control group） to 0.28 ±0.007,but increased that of vimentin from 0.83± 0.014 （blank control group） to 1.16±0.003, while BQ788 upregulated the E-cadherin expression to 0.42±0.008 and downregulated the vimentin expression to 0.75±0.030, and significant differences were observed in the E-cadherin expression and vimentin expression among the ET-3 group, ETRB antagonist group and blank control group （F = 329.98,262.94,respectively, both P 〈 0.01 ）. A significant increase was observed in the mRNA and protein expression of Slug （F= 376.94, 288.87, both

关 键 词：黑色素瘤 内皮缩血管肽3 上皮基质转化 

分 类 号：R739.5[医药卫生—肿瘤]