摘要
目的研究缺氧微环境对人源雄激素非依赖性前列腺癌细胞系PC3和人源雄激素依赖性前列腺癌细胞系LNCaP恶性生长的影响。方法两种前列腺癌细胞分别进行常氧和缺氧培养,常氧组培养条件为21%O_(2)、5%CO_(2)和74%N 2,缺氧组培养条件为1%O_(2)、5%CO_(2)和94%N 2。使用细胞增殖检测试剂盒(CCK-8)检测培养不同时间点两种癌细胞的增殖情况;采用实时荧光定量PCR(qRT-PCR)和蛋白印迹(Western blotting)方法检测癌细胞上皮标志物E钙粘蛋白(E-cadherin)和间充质标志物波形蛋白(Vimentin)mRNA和蛋白表达水平的变化;通过细胞划痕实验和Transwell实验检测癌细胞的转移和侵袭能力。结果短时间缺氧状态下两种细胞增殖活性均显著增强,与其他时间组比较,PC3和LNCaP分别在4 h和16 h时相对常氧状态增殖活性最高(P<0.05);但长时间缺氧后,两种细胞增殖活性均下降,分别在20 h和72 h时达到最低(P<0.05)。qRT-PCR和Western blotting结果显示,两种前列腺癌细胞中,不同缺氧时间组E-cadherin的mRNA和蛋白表达水平均显著低于常氧组(P<0.05),并且随缺氧时间的增加总体上呈现逐渐降低的趋势;各缺氧组Vimentin的mRNA和蛋白表达水平均显著高于常氧组(P<0.05),并且随缺氧时间的增加总体上呈现逐渐升高的趋势,呈现出典型的上皮间充质表型转化。细胞划痕和Transwell实验结果显示,在不同缺氧阶段,与常氧组比较,两种前列腺癌细胞的细胞迁移率和侵袭能力均显著增高(P<0.01)。结论缺氧促进前列腺癌的恶性生长是一个动态变化的过程,缺氧可以促进前列腺癌细胞的恶性增殖与迁移。
Objective:To study the effects of hypoxia microenvironment on the malignant growth of human androgen independent prostate cancer cell line PC3 and human androgen dependent prostate cancer cell line LNCaP.Methods:Two kinds of prostate cancer cells were cultured in normoxic and anoxic culture conditions respectively.The culture conditions of normoxia group were 21%O_(2),5%CO_(2) and 74%N 2.The culture conditions of hypoxia group were 1%O_(2),5%CO_(2) and 94%N 2.After culture for different time,the proliferation ability of the two kinds of cancer cells was detected by CCK-8 test.The mRNA and protein expression levels of epithelial marker E-cadherin and mesenchyme marker Vimentin were detected by qRT-PCR and Western blot respectively.Cell scratch assay and Transwell assay were used to detect the metastatic and invasive ability of the two prostate cancer cells.Results:The proliferation activity of the two kinds of cells was significantly increased after culture with short-time hypoxia.The proliferative activity of PC3 and LNCaP cells was the highest at 4 hours and 16 hours,respectively(P<0.05).However,the proliferation activity of the two kinds of cells decreased after a long period of hypoxia,reaching the lowest at 20 hours and 72 hours,respectively(P<0.05).The results of qRT-PCR and Western blot showed that the mRNA and protein expression levels of E-cadherin in the two kinds of prostate cancer cells in the different hypoxia time groups were significantly lower than those in the normal oxygen control group(P<0.05),and the overall trend was gradually decreased with the increase of hypoxia time.The expression levels of mRNA and protein of Vimentin in each hypoxia group were significantly higher than those in the normoxic group(P<0.05).The overall trend was gradually increased with the increase of hypoxia time,showing typical epithelial mesenchymal phenotypic transformation.The results of cell scratch and Transwell assay showed that the cell migration rates of both prostate cancer cells were significantly increased at
作者
郭爱鑫
杨丽雅
周晓光
何佳璘
郭一鸣
王慧萍
宋黎明
GUO Ai-xin;YANG Li-ya;ZHOU Xiao-guang;HE Jia-lin;GUO Yi-ming;WANG Hui-ping;SONG Li-ming(Graduate School of Peking Union Medical College,Chinese Academy of Medical Sciences,Beijing 100730;National Research Institute for Family Planning,Beijing 100081;Department of Urology,Beijing Chaoyang Hospital,Capital Medical University,Beijing 100020)
出处
《生殖医学杂志》
CAS
2021年第5期658-665,共8页
Journal of Reproductive Medicine
基金
国家自然科学基金面上项目(81772700)。
关键词
缺氧
前列腺癌
增殖
上皮间充质转化
迁移
Hypoxia
Prostate cancer
Proliferation
Epithelial mesenchymal transformation
Migration
