荷载精子相关抗原9基因短发卡RNA的溶瘤腺病毒ZD55-SPAG9联合放疗对前列腺癌LNCaP细胞的治疗作用  

Therapeutic effect of the combined use of oncolytic adenovirus carrying short hairpin RNA targeting sperm-associated antigen 9 with radiotherapy on prostate cancer LNCaP cells

作　　者：刘万启 孙方浩 魏晋 马赛 娄禄 高超 Liu Wanqi;Sun Fanghao;Wei Jin;Ma Sai;Lou Lu;Gao Chao(Department of Urology,the Affiliated Xuzhou Municipal Hospital of Xuzhou Medical University,Xuzhou 221002,China)

机构地区：[1]徐州医科大学附属市立医院泌尿外科,徐州221002

出　　处：《中华实验外科杂志》2023年第12期2446-2450,共5页Chinese Journal of Experimental Surgery

基　　金：徐州医科大学附属市立医院2022年拔尖人才项目。

摘　　要：目的观察荷载精子相关抗原9(SPAG9)基因短发卡RNA(shRNA)的溶瘤腺病毒ZD55-SPAG9联合放疗对前列腺癌LNCaP细胞的治疗作用并初步探讨其机制。方法将前列腺癌LNCaP细胞分为4组,分别为磷酸盐对照组(PBS组)、放疗组(10 GY)、病毒组[ZD55-SPAG9,10感染复数(MOI)]、联合组(ZD55-SPAG9+放疗,5 MOI+5 GY);细胞计数试剂盒(CCK-8)法检测各组LNCaP细胞增殖能力的变化;Hoechst-33258染色法检测各组细胞的凋亡;Transwell迁移及侵袭实验检测各组细胞的迁移和侵袭能力;蛋白质印迹法(Western blot)检测各组细胞SPAG9蛋白、凋亡相关蛋白及侵袭相关蛋白的表达。两组间差异的比较采用独立样本t检验,多组间差异比较采用One-way ANOVA分析。结果CCK-8结果显示,联合组LNCaP细胞的存活率为(46.02±2.20)%、病毒组为(56.83±2.40)%(t=12.855,P<0.01)、放疗组为(64.12±2.23)%(t=22.389,P<0.01)、PBS组为(99.63±2.72)%(t=59.407,P<0.01),各治疗组的细胞存活率均低于PBS组,差异有统计学意义(F=1407.384,P<0.01),联合组与单一治疗组比较,LNCaP细胞的存活率更低。Hoechst-33258染色实验结果显示,联合组LNCaP细胞的凋亡率为(55.80±1.49)%、病毒组为(36.42±1.78)%(t=-32.295,P<0.01)、放疗组为(30.01±1.80)%(t=-42.814,P<0.01)、PBS组为(9.30±1.53)%(t=-84.343,P<0.01),各治疗组的细胞凋亡率均高于PBS组,差异有统计学意义(F=2010.396,P<0.01),联合组与单一治疗组比较,LNCaP细胞的凋亡更加明显。Transwell迁移和侵袭实验结果显示,(1)迁移实验:联合组迁移细胞数为(102.60±16.03)个、病毒组为(163.67±11.00)个(t=12.166,P<0.01)、放疗组为(206.47±17.05)个(t=17.192,P<0.01)、PBS组为(304.73±17.16)个(t=33.345,P<0.01),各治疗组迁移细胞数均少于PBS组,差异有统计学意义(F=450.498,P<0.01),联合组迁移细胞数明显少于单一治疗组;(2)侵袭实验:联合组侵袭细胞数为(90.93±13.91)个、病毒组为(145.33±15.31)个(t=10.184,P<0.01)、放疗组为(191.33±1Objective To observe the therapeutic effect of the combined use of oncolytic adenovirus carrying short hairpin RNA targeting sperm-associated antigen 9(SPAG9)with radiotherapy on prostate cancer LNCaP cells and to initially explore its mechanism.Methods LNCaP prostate cancer cells were divided into 4 groups for intervention,namely:phosphate belanced solution control group(PBS group);radiotherapy group(10 GY);Oncolytic adenovirus group[ZD55-SPAG9,10 multiple infections(MOI)];Combination group(ZD55-SPAG9+radiotherapy,5 MOI+5 GY);cell counting kit-8(CCK-8)was used to detect the changes of LNCaP cell proliferation in each group;Apoptosis was detected by Hoechst-33258 staining;Transwell Migration and Invasion Test was used to detect the ability of cell migration and invasion in each group;The expression of SPAG9 protein,apoptosis-related protein and invasion-related protein were detected by Western blotting.The difference between the two groups was compared by independent sample t-test,and the difference between multiple groups was analyzed by One-way ANOVA.Results CCK-8 results showed that the survival rate of LNCaP cells in the combination group was(46.02±2.20)%,the oncolytic adenovirus group was(56.83±2.40)%(t=12.855,P<0.01),the radiotherapy group was(64.12±2.23)%(t=22.389,P<0.01),and the PBS group was(99.63±2.72)%(t=59.407,P<0.01).The survival rate of cells in each treatment group was lower than that in the PBS group,the difference was statistically significant F=1407.384,P<0.01),and the survival rate of LNCaP cells was lower in the combined group than in the single treatment group.The results of the Hoechst-33258 staining experiment showed that the apoptosis rate of LNCaP cells in the combination group was(55.80±1.49)%,the oncolytic adenovirus group was(36.42±1.78)%(t=-32.295,P<0.01),the radiotherapy group was(30.01±1.80)%(t=-42.814,P<0.01),and the PBS group(9.30±1.53)%(t=-84.343,P<0.01)%.The cell apoptosis rate in each treatment group was higher than PBS,with differences Statistical significance(F=2010.3

关 键 词：精子相关抗原9 前列腺癌 放疗 

分 类 号：R737.25[医药卫生—肿瘤]