CDK5RAP1通过Wnt/β-Catenin信号通路调控结直肠癌发生和进展的研究  

CDK5RAP1's regulation of occurrence and development of colorectal cancer through Wnt/β-Catenin signaling pathway

作　　者：蒋漫琦 何师茜 杨洪[1] 周晓倩[1] JIANG Manqi;HE Shiqian;YANG Hong;ZHOU Xiaoqian(Department of Gastroenterology,Guiyang First People's Hospital,Guiyang 550000,China)

机构地区：[1]贵阳市第一人民医院消化内科,550000

出　　处：《国际消化病杂志》2023年第4期246-256,共11页International Journal of Digestive Diseases

基　　金：贵阳市科技计划项目(筑科合同[2021]43-24号);2016年西部地区人才培养特别项目(201608525032)。

摘　　要：目的探讨周期素依赖性激酶5激活结合蛋白1(CDK5RAP1)通过Wnt/β-连环蛋白(β-Catenin)信号通路对结直肠癌(CRC)发生和进展的影响。方法选取2020年6月至2022年9月在贵阳市第一人民医院被首诊为CRC的72例患者的CRC组织和癌旁组织,检测组织中CDK5RAP1的表达水平。选取人CRC细胞系HCT-116、SW480、HT29、SW620、Caco-2及人正常结直肠上皮细胞系NCM460,检测各细胞中CDK5RAP1的表达水平。分别将si-NC和si-CDK5RAP1转染至HCT-116细胞中,分别将pc-NC和pc-CDK5RAP1转染至SW480细胞中,将SW480细胞分为SW480-CON组(正常培养细胞)、SW480-pc-NC组(阴性对照细胞)、SW480-pc-CDK5RAP1组(CDK5RAP1过表达细胞)和SW480-HLY78组(SW480-pc-CDK5RAP1+20μmol/L Wnt激活剂HLY78),将HCT-116细胞分为HCT-116-CON组(正常培养细胞)、HCT-116-si-NC组(阴性对照细胞)、HCT-116-si-CDK5RAP1组(CDK5RAP1低表达细胞)和HCT-116-HLY78组(HCT-116-si-CDK5RAP1+20μmol/L HLY78)。检测各组CRC细胞中CDK5RAP1的表达水平、CRC细胞存活率(增殖能力)、克隆、侵袭、迁移情况,以及CRC细胞成球率(细胞干性)。检测各组CRC细胞中β-Catenin、Axin1、c-Myc、CD133、CD44、SOX2蛋白的表达水平。选取18只BALB/C裸鼠随机分为CON组、si-NC组和si-CDK5RAP1组,每组6只,分别在各组裸鼠右前肢腋下注射对应的HCT-116细胞悬液,5周后处死裸鼠,剥离移植瘤并称重比较。结果CRC组织及细胞中CDK5RAP1的表达水平均显著高于癌旁组织和人正常结直肠上皮细胞(P均<0.05),且CDK5RAP1在HCT-116细胞中表达水平最高,在SW480细胞中表达水平最低,故分别使用HCT-116细胞和SW480细胞构建CDK5RAP1敲低和过表达的慢病毒载体转染细胞株。与SW480-CON组和SW480-pc-NC组比较,SW480-pc-CDK5RAP1组细胞的CDK5RAP1表达水平、细胞存活率、细胞克隆率、细胞划痕愈合率、细胞侵袭数量、细胞成球率及β-Catenin、c-Myc、CD133、CD44、SOX2蛋白表达水平均显著升高,Axin1蛋白表达水平Objective An attempt is made in this paper to investigate the influences of cyclindependent kinase 5 regulatory subunit associated protein 1(CDK5RAP1)on the occurrence and development of colorectal cancer(CRC)through Wnt/β-Catenin signaling pathway.Methods The CRC tissues and paracancerous tissues of 72 patients who were first diagnosed as CRC in Guiyang First People's Hospital from June 2020 to September 2022 were selected to detect the expression level of CDK5RAP1 in the tissues.Human CRC cell lines HCT-116,SW480,HT29,SW620,Caco-2,and human normal colorectal epithelial cell line NCM460 were selected to detect the expression level of CDK5RAP1 in each cell.Si-NC and si-CDK5RAP1 were transfected into HCT-116 cells,and pc-NC and pc-CDK5RAP1 were transfected into SW480 cells,respectively.SW480 cells were divided into the SW480-CON group(normal cultured cells),the SW480-pc-NC group(negative control cells),the SW480-pc-CDK5RAP1 group(CDK5RAP1 overexpressing cells),and the SW480-HLY78 group(SW480-pc-CDK5RAP1+20μmol/L Wnt activator HLY78).HCT-116 cells were divided into the HCT-116-CON group(normal cultured cells),the HCT-116-si-NC group(negative control cells),the HCT-116-si-CDK5RAP1 group(CDK5RAP1 low expression cells),and the HCT-116-HLY78 group(HCT-116-si-CDK5RAP1+20μmol/L HLY78).The expression level of CDK5RAP1,CRC cell survival rate(proliferation ability),cloning,invasion,migration status,and CRC cell sphericity(cell dryness)were detected in each group of CRC cells.The expression levels ofβ-Catenin,Axin1,c-Myc,CD133,CD44,and SOX2 proteins were detected in each group of CRC cells.Eighteen BALB/C nude mice were randomly divided into the CON group,the si-NC group,and the si-CDK5RAP1 group,with 6 mice in each group.HCT-116 cell suspensions were injected into the right forearm armpit of each group of nude mice,and after 5 weeks,the nude mice were killed.The transplanted tumors were dissected and weighed for comparison.Results The expression levels of CDK5RAP1 in CRC tissue and cells are significantly higher than th

关 键 词：周期素依赖性激酶5激活结合蛋白1 WNT/Β-CATENIN信号通路 结直肠癌 

分 类 号：R735.3[医药卫生—肿瘤]