摘要
目的:探讨分化抑制因子1基因(inhibitor of differentiation-1, Id1)和Id3是否协同促进结肠癌SW620细胞EMT和细胞的侵袭迁移能力及其可能的机制。方法:利用慢病毒载体构建Idl和Id3单或双基因敲减的结肠癌SW620细胞,实验分为4组:SW620-Sh-Id1组(转染shRNA-Id1)、SW620-Sh-Id3组(转染shRNA-Id3)、SW620-Sh-Id1-Id3组(共转染shRNA-Id1和shRNAId3)、SW620-NC组(转染阴性空载慢病毒)。用qPCR法和Western blotting法分别检测敲减效果,显微镜下观察稳定下调Idl和Id3后SW620细胞形态的变化,划痕愈合实验和Transwell小室法检测SW620细胞迁移和侵袭的变化,Western blotting检测EMT标志分子及迁移、侵袭相关蛋白的表达。结果:成功构建了Idl和Id3单基因敲减与双基因敲减的结肠癌SW620细胞株。(1)Idl和Id3的敲除诱导SW620细胞形态由上皮样向间质样转变;(2)与对照组相比,SW620-Sh-Id1组和SW620-Sh-Id3组SW620细胞侵袭和迁移的能力显著下降(均P<0.05);SW620-Sh-Id1-Id3组细胞侵袭和迁移的能力较SW620-Sh-Id1组和SW620-Sh-Id3组也明显下降(均P<0.05);(3)与对照组相比,SW620-Sh-Id1组和SW620-Sh-Id3组β-catenin、snail1及MMP2蛋白表达降低,上皮黏蛋白及TIMP2蛋白表达增高(均P<0.05);SW620-Sh-Id1-Id3组与SW620-Sh-Id1组和SW620-Sh-Id3组相比,β-catenin、snail1及MMP2蛋白表达也下降,同时上皮钙黏蛋白及TIMP2蛋白表达也增高(均P<0.05)。结论:Id1和Id3可能通过诱导EMT协同影响结肠癌SW620细胞侵袭与迁移的能力。
Objective： To investigate whether inhibitor of differentiation 1 gene （Id1） and Id3 gene can synergistically promote epithelial-mesenchymal transition （EMT）, invasion and migration of colon cancer SW620 cells and to explore its underlying mechanisms.Methods： The SW620 cell strain with Idl or Id3 gene knockdown and the SW620 cell strain with Id1/Id3 gene double-knockdown were constructed by lentiviral vectors transfection. The SW620 cells were divided into four groups, which included SW620-Sh-Id1 group （transfected with shRNA-Id1）, SW620-Sh-Id3 group （transfected with shRNA-Id3）, SW620-Sh-Id1-Id3 group （transfected with shRNAId1 plus shRNA-Id3） and SW620-NC group （transfected with negative lentivirus）. The efficiency of knockdown was detected by Realtime qPCR and Western blotting. The influence of stable knockdown of Idl or Id3 on cell morphological change was observed under a microscope. The changes of migration and invasion abilities of the SW620 cells were determined by wound healing assay and Transwell assay. EMT, invasion and migration related proteins were measured by Western blotting. Results： The SW620 cell strains with Idl and/or Id3 gene knockdown were successfully constructed. Idl and Id3 knockdown induced the epithelial-like to the mesenchymanl-like transformation of SW620 cells. （1） Compared with the control group, the invasion and migration abilities of the SW620 cells were significantly decreased in the SW620-Sh-Id1 group and SW620-Sh-Id3 group （all P〈0.05）. （2） Meanwhile, the invasion and migration abilities in the SW620-Sh-Id1-Id3 group were obviously weaker than the SW620-Sh-Id1 group and SW620-Sh-Id3 group （all P〈0.05）. （3）Compared with the control group, the SW620-Sh-Id1 group and SW620-Sh-Id3 group had a reduction in the protein expressions of β-catenin, snail1 and MMP2, and an increase in the protein expressions of E-cadherin and TIMP2 （all P〈0.05）. （4） Compared with the SW620-Sh-Id1 group and SW620-Sh-Id3 group , the protein expressions of β-catenin, snail1 and MMP2 were reduced, and the protein expressions of E-cadherin and TIMP2 were increased in the SW620-Sh-Id1-Id3 group （all P〈0.05）. Conclusion： Id1 and Id3 could synergistically influence invasion and migration of SW620 cells, possibly through inducing EMT.
作者
陈韡
于悦
孙艳霞
林万松
叶韵斌
CHEN Wei;YU Yue;SUN Yanxia;LIN Wansong;YE Ytmbin(a.College of Basic Medicine/1b.Tumor Immunology Laboratory,Teaching Hospital of Fujian Medical University ＆ Fujian Province Tumor Hospital,Fuzhou 350014,Fujian,China;Fujian Key Laboratory of Translational Medicine,Fuzhou 350014,Fujian,China)
出处
《中国肿瘤生物治疗杂志》
CAS
CSCD
北大核心
2018年第10期987-993,共7页
Chinese Journal of Cancer Biotherapy
基金
国家自然科学基金资助项目(No.81472720)~~
关键词
分化抑制因子1基因
分化抑制因子3
上皮间质转化
结肠癌
SW620细胞
侵袭
迁移
inhibitor of differentiation 1 gene （Id1）
inhibitor of differentiation 3 gene （Id3）
epithelial-mesenchymal transition（EMT）
colon cancer
SW620 cell
invasion
migration
