干扰PD-L1表达对小鼠B细胞淋巴瘤的抑制作用  

Inhibitory Effect of Interference with PD-L1 Expression on B-cell lymphoma in Mice

作　　者：施青青[1] 朱淼[1] 孙幸[1] 倪军[1] 孙梅[1] 顾健[1] 吴蔚[1] SHI Qing-Qing;ZHU Miao;SUN Xin;NI Jun;SUN Mei;GU Jian;WU Wei(Laboratory of Hematonosis,Northern Jiangsu People's Hospital,Yangzhou Institute of Hematology,Yangzhou 225001,Jiangsu Province,China)

机构地区：[1]江苏省苏北人民医院血液病实验室,扬州市血液学研究所,江苏扬州225001

出　　处：《中国实验血液学杂志》2022年第3期771-777,共7页Journal of Experimental Hematology

基　　金：江苏省苏北人民医院院级课题(yzucms 2018901)。

摘　　要：目的:探讨干扰PD-L1(programmed death 1-ligad)的表达对抑制小鼠B细胞淋巴瘤的作用。方法:针对C D274(PD-L1)靶基因序列,设计3个RNA干扰靶点序列,连接到pGMVL-SC5干扰载体,瞬时转染293T细胞。RT-q PCR检验各靶点对PD-L1的干扰效率,将干扰效率最佳的shRNA载体进行慢病毒包装,转染A20细胞,获得稳定低表达PD-L1的A20淋巴瘤细胞株(CD274-sh A20),将A20细胞株和CD274-sh A20细胞株以等数体外培养48 h,MTT法比较体外增殖的差异;同时使用2株细胞分别对免疫功能正常的BALB/c小鼠进行荷瘤,对照组皮下注射等体积PBS缓冲液;通过活体荧光成像评估成瘤情况,记录各组小鼠生存期。结果:CD274-sh A20细胞体外增殖速率显著低于PD-L1高表达的A20细胞(P<0.05)。与普通A20细胞荷瘤组相比,PD-L1干扰荷瘤组小鼠外观及活体荧光成像均可见其瘤体显著缩小。小鼠解剖后称取瘤重,测量瘤体积,结果显示,瘤重及瘤体积明显缩小(P<0.05)。小鼠生存期也较PD-L1高表达组有一定延长。结论:PD-L1在淋巴瘤的发生发展中起到重要作用,干扰其表达能抑制细胞的体外增殖,对于瘤体的生长有一定的抑制作用。Objective: To investigate the potential inhibitory effect of interference with PD-L1 on B cell lymphoma in m ice. Methods: Three shRNA vectors for mouse CD274(PD-L1) were constructed and transiently transfected into 293 T c ells. RT-q PCR was used to validate the interference efficiency of CD274. The shRNA vector that interfere efficiently with CD274 expression was packaged by using lentivirus packaging system to generate shRNA lentivirus, and then transfected into A20 lymphoma cell line. The methyl thiazol terazolium(MTT) assay was used to detect proliferation after 48 h culture o f CD274-sh A20 cells. Meanwhile, BALB/c mice were hypodermically infected with CD274-sh A20 cells. Infected mice were observed daily and assessed to visualize tumor by in vivo fluorescence imaging. Results: The proliferation rate of CD274-sh A20 cells in vitro was significantly lower than that of A20 cells(P<0.05). The tumor size detected by in vivo f luorescence imaging showed a significant reduce in tumor bearing mice with CD274-sh compared with other tumor bearing mice. And the weight and size of tumor in CD274-sh group were also significantly reduced compared with other group(P<0.05). Moreover, the survival time of tumor bearing mice in CD274-sh group was longer than that of the PD-L1 h igh expression group. Conclusion: PD-L1 plays an important role in the incidence and the progression of lymphoma, a nd the shRNA-based PD-L1 knockdown can inhibit cell proliferation of A20 cells and partly suppress tumor growth.

关 键 词：PD-L1 RNA干扰 B系淋巴瘤 

分 类 号：R733[医药卫生—肿瘤]