沉默p21活化激酶1表达对人黑素瘤A375细胞凋亡及侵袭能力的影响  

Effect of silencing p21-activated kinase 1 expression on apoptosis and invasion of human melanoma A375 cells

作　　者：张娜[1] 李税琪[1] 李红文[1] ZHANG Na;LI Shuiqi;LI Hongwen(Department of Dermatology,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)

机构地区：[1]郑州大学第一附属医院皮肤科,郑州450052

出　　处：《中华实用诊断与治疗杂志》2020年第5期433-436,共4页Journal of Chinese Practical Diagnosis and Therapy

基　　金：国家自然科学基金青年基金(81502731);河南省科技厅项目(182300410305)。

摘　　要：目的探讨shRNA沉默p21活化激酶1(p21-activated kinase 1,PAK1)基因表达对人黑素瘤A375细胞凋亡、迁移及侵袭能力的影响.方法对数生长期人黑素瘤A375细胞随机分为转染组(转染PAK1特异性shRNA)、阴性对照组(转染NC-shRNA)、空白对照组(不进行转染).转染后培养72 h,采用实时荧光定量PCR法检测A375细胞PAK1mRNA相对表达量,采用Western blot法检测A375细胞PAK1及p-PAK1蛋白相对表达量,采用流式细胞术检测细胞凋亡率,采用Transwell小室实验检测细胞迁移、侵袭能力.结果转染后培养72 h,转染组PAK1 mRNA相对表达量(0.45±0.07)、PAK1蛋白相对表达量(0.53±0.01)、p-PAK1蛋白相对表达量(0.21±0.03)均低于阴性对照组(0.98±0.04、0.65±0.02、0.38±0.04)、空白对照组(1.00±0.08、0.67±0.51、0.36±0.02)(P<0.05),细胞凋亡率[(11.20±0.21)%]高于阴性对照组[(5.86±0.51)%]、空白对照组[(5.43±0.03)%](P<0.05),迁移细胞数[(47.00±2.23)个]、侵袭细胞数[(30.00±4.44)个]较阴性对照组[(76.00±4.85)、(54.00±3.67)个]、空白对照组[(77.90±4.97)、(56.00±4.12)个]少(P<0.05),阴性对照组上述各指标与空白对照组比较差异均无统计学意义(P>0.05).结论沉默PAK1基因表达可促进人黑素瘤A375细胞凋亡,降低其迁移及侵袭能力.Objective To investigate the effects of shRNA silencing p2l-activated kinases l(PAKl)gene on the apoptosis,migration and invasion of human melanoma A375cells.Methods The A375cells in logarithmic growth phase were randomly divided into transfection group(transfected with pG1.1-PAK1-shRNA),negative control group(transfected with pG1.1-NC-shRNA)and blank control group(without transfection).After 72hculture,the expression of fluorescence was observed under fluorescence microscope,the relative expression of PAK1mRNA was detected by realtime fluorescence quantitative PCR,the relative expressions of PAK1and p-PAK1protein were detected by Western blot,the apoptotic rate was detected by flow cytometry,and the migration and invasion abilities were evaluated by Transwell chamber experiment.Results After 72hculture,the relative expressions of PAK1mRNA,PAK1protein and p-PAK1 protein were lower in transfection group(0.45±0.07,0.53±0.01,0.21±0.03)than those in negative control group(0.98±0.04,0.65±0.02,0.38±0.04)and blank control group(1.00±0.08,0.67±0.51,0.36±0.02)(P<0.05),the apoptotic rate was higher in transfection group((11.20±0.21)%)than that in negative control group((5.86±0.51)%)and blank control group((5.43±0.03)%)(P<0.05),the counts of migrating cells and invading cells were lower in transfection group(47.00±2.23,30.00±4.44)than those in negative control group(76.00±4.85,54.00±3.67)and blank control group(77.90±4.97,56.00±4.12),and all the above showed no significant differences between negative control group and blank control group(P>0.05).Conclusion Silencing PAK1gene can promote the apoptosis of human melanoma A375cells and reduce its migration and invasion abilities.

关 键 词：黑素瘤 A375细胞 p21活化激酶1 

分 类 号：R739.5[医药卫生—肿瘤]