国际肿瘤学杂志

国际肿瘤学杂志 ›› 2021, Vol. 48 ›› Issue (12): 705-710.doi: 10.3760/cma.j.cn371439-20210813-00140

• 论著 •    下一篇

沉默PD-L1表达对胃癌细胞生物学行为的影响

赵丽丽1, 赵文文2, 冯青青2, 赵文飞2, 张雪1, 井文君2, 魏红梅2   

  1. 1青岛大学医学部 266042
    2青岛大学附属青岛市中心医院肿瘤综合二科 266042
  • 收稿日期:2021-08-13 修回日期:2021-10-03 出版日期:2021-12-08 发布日期:2022-01-12

Effects of silencing PD-L1 expression on biological behaviors of gastric cancer cells

Zhao Lili1, Zhao Wenwen2, Feng Qingqing2, Zhao Wenfei2, Zhang Xue1, Jing Wenjun2, Wei Hongmei2   

  1. 1Medical College of Qingdao University, Qingdao 266042, China
    2Second Department of General Oncology, Affiliated Qingdao Central Hospital, Qingdao University, Qingdao 266042, China
  • Received:2021-08-13 Revised:2021-10-03 Online:2021-12-08 Published:2022-01-12

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摘要： 目的 探讨程序性死亡受体配体-1(PD-L1)的表达对胃癌细胞MKN45生物学行为的影响。 方法 采用RNA干扰技术,沉默胃癌细胞MKN45的PD-L1基因,将MKN45细胞分为空白对照组、si-NC组(转染siRNA-NC)及si-PD-L1组(转染siRNA-PD-L1)。采用实时定量PCR法检测各组MKN45细胞PD-L1及上皮间质转化(EMT)相关蛋白上皮钙黏素、波形蛋白、Snail的mRNA表达水平;采用蛋白质印记法检测各组MKN45细胞PD-L1蛋白的表达水平;采用Transwell小室迁移实验、Transwell小室侵袭实验及MTT实验检测MKN45细胞迁移、侵袭和黏附能力。 结果 PD-L1在空白对照组、si-NC组、si-PD-L1组细胞中的mRNA相对表达量分别为1.002±0.092、1.005±0.121、0.237±0.017,差异具有统计学意义(F=75.61,P<0.001);PD-L1在3组细胞中的蛋白表达量分别为0.944±0.028、1.008±0.088、0.269±0.015,差异具有统计学意义(F=172.99,P<0.001);si-PD-L1组PD-L1的mRNA和蛋白表达均低于其他两组(均P<0.001),而空白对照组和si-NC组之间差异均无统计学意义(均P>0.05)。空白对照组、si-NC组和si-PD-L1组3组细胞的迁移率分别为(1.000±0.020)%、(1.012±0.084)%、(0.488±0.050)%,差异具有统计学意义(F=80.73,P<0.001);3组细胞的侵袭率分别为(0.929±0.087)%、(0.924±0.208)%、(0.300±0.100)%,差异具有统计学意义(F=19.37,P<0.001);3组细胞的黏附率分别为(100.000±5.407)%、(99.280±4.845)%、(59.723±2.674)%,差异具有统计学意义(F=79.87,P<0.001);与空白对照组和si-NC组相比较,si-PD-L1组MKN45细胞的迁移能力、侵袭能力及黏附能力均显著下降(均P<0.001)。3组细胞上皮钙黏素mRNA相对表达量分别为1.000±0.023、0.981±0.051、3.618±0.201,波形蛋白mRNA相对表达量分别为1.000±0.043、1.108±0.150、0.328±0.011,Snail mRNA相对表达量分别为1.061±0.103、1.090±0.110、0.304±0.043,差异均具有统计学意义(F=477.17,P<0.001;F=65.97,P<0.001;F=72.70,P<0.001);与空白对照组和si-NC组相比较,si-PD-L1组MKN45细胞的波形蛋白及Snail的mRNA表达均降低,上皮钙黏素mRNA表达升高,差异均具有统计学意义(均P<0.001)。 结论 沉默PD-L1的表达可降低MKN45细胞的迁移、侵袭及黏附能力,其机制可能与PD-L1影响胃癌EMT通路相关。

关键词: 胃肿瘤, 细胞运动, 上皮间质转化, PD-L1

Abstract: Objective To investigate the effects of programmed death-ligand 1 (PD-L1) expression on the biological behaviors of gastric cancer cell line MKN45. Methods The PD-L1 gene of gastric cancer cell line MKN45 was silenced by RNA interference technique. MKN45 cells were divided into blank control group, si-NC group (transfected with siRNA-NC) and si-PD-L1 group (transfected with siRNA-PD-L1). Quantitative real-time PCR was used to detect the mRNA expressions of PD-L1 and epithelial-mesenchymal transformation (EMT)-related proteins E-cadherin, Vimentin and Snail in MKN45 cells, and Western blotting was used to detect the expression levels of PD-L1 protein in MKN45 cells of each group. Transwell migration test, Transwell invasion test and MTT test were used to detect the migration, invasion and adhesion abilities of MKN45 cells. Results The relative expression levels of PD-L1 mRNA in the blank control group, si-NC group and si-PD-L1 group were 1.002±0.092, 1.005±0.121 and 0.237±0.017, respectively, with a statistically significant difference (F=75.61, P<0.001). The protein expression levels of PD-L1 in the three groups were 0.944±0.028, 1.008±0.088 and 0.269±0.015, respectively, with a statistically significant difference (F=172.99, P<0.001). The mRNA and protein expression levels of PD-L1 in the si-PD-L1 group were lower than those in the other two groups (all P<0.001), but there were no statistically significant differences between the blank control group and si-NC group (all P>0.05). The cell migration rates of the blank control group, si-NC group and si-PD-L1 group were (1.000±0.020)%, (1.012±0.084)% and (0.488±0.050)%, respectively, with a statistically significant difference (F=80.73, P<0.001). The cell invasion rates of the three groups were (0.929±0.087)%, (0.924±0.208)% and (0.300±0.100)%, respectively, with a statistically significant difference (F=19.37, P<0.001), and the cell adhesion rates of the three groups were (100.000±5.407)%, (99.280±4.845)% and (59.723±2.674)%, respectively, with a statistically significant difference (F=79.87, P<0.001). Compared with the blank control group and si-NC group, the migration, invasion and adhesion abilities of MKN45 cells in the si-PD-L1 group decreased significantly (all P<0.001). The expression levels of E-cadherin mRNA of the three groups were 1.000±0.023, 0.981±0.051, 3.618±0.201, the expression levels of Vimentin mRNA were 1.000±0.043, 1.108±0.150, 0.328±0.011, the expression levels of Snail mRNA were 1.061±0.103, 1.090±0.110, 0.304±0.043, respectively, with statistically significant differences (F=477.17, P<0.001; F=65.97, P<0.001; F=72.70, P<0.001). Compared with the blank control group and si-NC group, the mRNA expression levels of Vimentin and Snail of MKN45 cells in the si-PD-L1 group decreased, while the expression level of E-cadherin mRNA increased, with statistically significant differences (all P<0.001). Conclusion Silencing the expression of PD-L1 can reduce the migration, invasion and adhesion abilities of MKN45 cells, and the mechanism may be related to the effect of PD-L1 on the EMT pathway of gastric cancer.

Key words: Stomach neoplasms, Cell movement, Epithelial-mesenchymal transition, PD-L1