摘要
目的通过检测内皮素转化酶2(ECE2)在乳腺癌中的表达,探究其对乳腺癌细胞增殖、迁移的影响及潜在分子机制。方法选取20例临床乳腺癌组织及其对应癌旁正常组织样本,通过qRT-PCR检测ECE2基因在乳腺癌组织中的表达特征。通过siRNA介导敲低ECE2表达,利用细胞增殖实验、克隆形成实验、划痕迁移实验和细胞周期实验分别验证敲低ECE2表达对乳腺癌细胞增殖、克隆形成、迁移及凋亡的影响。通过分析与ECE2共表达基因参与的信号通路,探索ECE2在乳腺癌发展进程中的潜在分子机制,进一步通过细胞增殖回补实验进行验证。结果20例临床乳腺癌组织中ECE2表达较癌旁正常组织显著上调(30.342±6.564 vs 21.753±8.244),差异有统计学意义(t=3.645,P=0.000)。转染敲低ECE2表达后,siECE2#1组和siECE2#2组在24,48,72 h的吸光度值较对照组明显降低,差异有统计学意义(F=71.409~211.681,均P<0.01)。siECE2#1组(0.515±0.014)和siECE2#2组(0.511±0.006)细胞克隆形成速率明显低于对照组(3.473±0.147),差异有统计学意义(F=467.152,P<0.001)。siECE2#1组(56.423±1.137)和siECE2#2组(42.036±0.754)细胞迁移愈合速率显著低于对照组(78.124±1.352),差异有统计学意义(F=805.162,P<0.001)。siECE2#1组和siECE2#2组细胞凋亡率较对照组明显降低,细胞周期显著阻滞在G1期。siECE2#1组和siECE2#组细胞中E2F1 mRNA和蛋白表达水平明显低于对照组,差异均有统计学意义(F=703.020,136.301,均P<0.001);MYC mRNA和蛋白表达水平也明显低于对照组(F=613.395,102.573,均P<0.001)。在敲低ECE2表达的乳腺癌细胞中分别回补E2F1和MYC后,细胞增殖速率回归至正常水平。结论ECE2高表达诱导促癌基因E2F1和MYC的高表达进而促进乳腺癌细胞的增殖及迁移,阻碍细胞周期停滞在G1期,参与乳腺癌的发展进程。
Objective To detect the expression of ECE2 in breast cancer and explore its effect on proliferation and migration of breast cancer cells and its potential molecular mechanism.Methods Twenty clinical breast cancer tissues and corresponding adjacent normal tissue samples were selected to detect the expression characteristics of ECE2 gene in breast cancer tissues by qRTPCR.The effects of knockdown of ECE2 expression on proliferation,clone formation,migration and apoptosis of breast cancer cells were verified by cell proliferation assay,clone formation assay,scratch migration assay and cell cycle assay,respectively.The potential molecular mechanism of ECE2 in the development of breast cancer was explored by analyzing the signaling pathways involved in the genes co-expressed with ECE2,and further verified by cell proliferation and complement experiments.Results The expression of ECE2 in 20 clinical breast cancer tissues was significantly higher than that in adjacent normal tissues(30.342±6.564 vs 21.753±8.244),the difference was statistically significant(t=3.645,P=0.000).After knockdown of ECE2 expression by transfection,the absorbance of SIEC2#1 and SIEC2#2 groups at 24,48 and 72 h was significantly lower than that of the control group,the difference was statistically significant(F=71.409~211.681,all P<0.01).The cell clone formation rate in SiECE2#1 group(0.515±0.014)and SiECE2#2 group(0.511±0.006)was significantly lower than that in control group(3.473±0.147),the difference was statistically significant(F=467.152,P<0.001).Cell migration and healing rate in SiECE2#1 group(56.423±1.137)and SiECE2#2 group(42.036±0.754)was significantly lower than that in control group(78.124±1.352),the difference was statistically significant(F=805.162,P<0.001).The apoptosis rate of SiECE2#1 and SiECE2#2 groups was significantly lower than that of the control group,and the cell cycle was significantly arrested in the G1 phase.The expression levels of E2F1 mRNA and protein in SiEC2#1 and SiEC2#groups were significantly lower tha
作者
顾芯烨
吴旦平
江波
商江峰
蒋国勤
魏金荣
李华
GU Xin-ye;WU Dan-ping;JIANG Bo;SHANG Jiang-feng;JIANG Guo-qin;WEI Jin-rong;LI hua(Department of Breast and Thyroid Surgery,the First People’s Hospital of Changshu City,Jiangsu Changshu 215500,China;Department of General Surgery,the Second Affiliated Hospital of Soochow University,Jiangsu Suzhou 215004,China;Fugu County People’s Hospital,Shaanxi Yulin 719400,China)
出处
《现代检验医学杂志》
CAS
2021年第4期39-44,共6页
Journal of Modern Laboratory Medicine
基金
江苏省卫生计生委科研课题(Z201951)。
