微小RNA-15b-5p靶向犰狳重复X连锁蛋白2抑制膀胱尿路上皮癌细胞增殖的实验研究  

Proliferation inhibition of bladder urothelial carcinoma cells by microRNA-15b-5p targeting armadillo repeat containing,X-linked 2

作　　者：陈化磊[1] 卓小岸[2] 卓小丽[3] 车宪平[1] 胡鑫明[1] Chen Hualei;Zhuo Xiaoan;Zhuo Xiaoli;Che Xianping;Hu Xinming(Department of Urology,the Second Affiliated Hospital of Hainan Medical College,Haikou 570311,China;Department of Emergency,Hainan People′s Hospital(Hainan Hospital Affiliated to Hainan Medical College),Haikou 570100,China;Department of Nuclear Medicine,Hainan People′s Hospital(Hainan Hospital Affiliated to Hainan Medical College),Haikou 570100,China)

机构地区：[1]海南医学院第二附属医院泌尿外科,海口570311 [2]海南省人民医院(海南医学院附属海南医院)急诊外科,海口570100 [3]海南省人民医院(海南医学院附属海南医院)核医学科,海口570100

出　　处：《中华实验外科杂志》2021年第1期123-126,共4页Chinese Journal of Experimental Surgery

摘　　要：目的检测膀胱尿路上皮癌中微小RNA-15b-5p(miR-15b-5p)的表达,探讨其靶向犰狳重复X连锁蛋白2(ARMCX2)对细胞增殖的调控作用。方法选择69例2014年6月至2015年5月在海南医学院第二附属医院确诊并行手术治疗膀胱尿路上皮癌的患者,留取肿瘤组织。实时荧光定量PCR(qPCR)检测miR-15b-5p、免疫组织化学法检测ARMCX2和细胞核增殖抗原(Ki-67)的表达。选择尿路上皮癌T24细胞系,应用双荧光素酶报告基因实验验证miR-15b-5p和ARMCX2的靶向关系。建立无关序列组(NC组)、miR-15b-5p mimic组、miR-15b-5p inhibitor组、miR-15b-5p inhibitor和小干扰RNA(siRNA)-ARMCX2共转染组,应用蛋白质印迹法(Western blot)检测ARMCX2和Ki-67的表达,应用细胞计数试剂盒(CCK-8)检测细胞活性。两组间比较行独立样本的t检验,多组间比较行方差分析(SNK法行两两比较)。结果膀胱尿路上皮癌中miR-15b-5p的表达量范围为1.2~4.5,miR-15b-5p的表达量和ARMCX2的阳性率在不同病变级别(1.51±0.26比2.04±0.43,t=4.560,P<0.05;78.95%比19.35%,χ^2=24.290,P<0.05)、是否为浸润性生长(1.60±0.40比1.86±0.44,t=2.360,P<0.05;73.33%比35.90%,χ^2=9.520,P<0.05)的分组中差异有统计学意义。相关分析显示miR-15b-5p和ARMCX2(r=-0.620,P<0.05)、miR-15b-5p和Ki-67(r=-0.600,P<0.05)呈负相关,ARMCX2和Ki-67(r=0.660,P<0.05)呈正相关。生存分析显示miR-15b-5p和ARMCX2与生存时间有关(χ^2=5.010,P<0.05)。双荧光素酶报告基因实验显示miR-15b-5p和ARMCX2具有靶向关系。MiR-15b-5p mimic组中ARMCX2(1.15±0.40比1.82±0.28,t=3.430,P<0.05)和Ki-67(1.34±0.37比0.99±0.21,t=2.780,P<0.05)的表达高于NC组,细胞活性低于NC组,miR-15b-5p inhibitor组中ARMCX2(2.24±0.35比1.82±0.28,t=2.630,P<0.05)和Ki-67(1.65±0.26比0.99±0.21,t=5.760,P<0.05)的表达高于NC组,细胞活性高于NC组,miR-15b-5p inhibitor和siRNA-ARMCX2共转染组ARMCX2(1.66±0.25比1.82±0.28,t=1.130,P>0.05)、Ki-67(1.24±0.29比0.99±0.21,t=1.710,P>0.05)的表达差异Objective To detect the expression of microRNA(miRNA,miR)-15b-5p in urothelial carcinoma of bladder and analyze the regulatory effect of miR-15b-5p armadillo repeat-containing X-linked protein 2(ARMCX2)on cell proliferation.Methods A total of 69 cases of urothelial carcinoma of bladder were selected.The expression of miR-15b-5p was detected by quantitative polymerase chain reaction(qPCR),andARMCX2and Ki-67 were detected by immunohistochemistry(IHC)mehtod.T24 cell line of urothelial carcinoma was selected.The targeting relationship between miR-15b-5p and ARMCX2 was verified by double luciferase reporter gene assay.Unrelated sequence group(NC group),miR-15b-5p mimic group,miR-15b-5p inhibitor group,miR-15b-5pinhibitor and small interfering RNA(siRNA)-ARMCX2 co-transfection group were established.The expression of ARMCX2 and proliferation cell nuclear antigen(Ki-67)was detected by Western blotting.Cell activity was detected by cell counting kit-8(CCK-8)assay.Results Expression range of miR-15b-5p was 1.2-4.5 in urothelial carcinoma.The expressions of miR-15b-5p and ARMCX2 were statistically significant in pathological grades(1.51±0.26 vs.2.04±0.43,t=4.560,P<0.05;78.95%vs.19.35%,χ^2=24.290,P<0.01)and infiltration(1.60±0.40 vs.1.86±0.44,t=2.360,P<0.05;73.33%vs.35.90%,χ^2=9.520,P<0.01).Negative correlation was found between miR-15b-5p and ARMCX2(r=-0.620,P<0.05),miR-15b-5p and Ki-67(r=-0.600,P<0.05).Positive correlation was found between ARMCX2 and Ki-67(r=0.660,P<0.05).MiR-15b-5p and ARMCX2 were related with survival time(χ^2=5.010,P<0.05).Targeting relationship was found between miR-15b-5p and ARMCX2.Compared with NC group,the expressions of ARMCX2(1.15±0.40 vs.1.82±0.28,t=3.430,P<0.05)and Ki-67(1.34±0.37 vs.0.99±0.21,t=2.780,P<0.05),cell activity were down-regulated in miR-15b-5p mimic group,expressions of ARMCX2(2.24±0.35 vs.1.82±0.28,t=2.630,P<0.05)and Ki-67(1.65±0.26 vs.0.99±0.21,t=5.760,P<0.01),cell activity were up-regulated in miR-15b-5p inhibitor group,ARMCX2(1.66±0.25 vs.1.82±0.28,t=1.130,P>

关 键 词：尿路上皮癌 微小RNA 增殖 预后 

分 类 号：R737.14[医药卫生—肿瘤]