重组新城疫病毒rL-RVG通过调控NLRP3/Caspase1/GSDMD通路诱导肺腺癌PC9细胞焦亡  

Recombinant Newcastle disease virus rL-RVG induces lung adenocarcinoma PC9 cell pyroptosis by modulating the NLRP3/Caspase1/GSDMD pathway

作　　者：李洋 何英珏 贡克文 田义督 王晨 严玉兰[1,2] LI Yang;HE Yingjue;GONG Kewen;TIAN Yidu;WANG Chen;YAN Yulan(Department of Respiratory Medicine,People's Hospital Affiliated to Jiangsu University,Jiangsu Zhenjiang 212000,China;Department of Respiratory Medicine,the University of Hong Kong-Shenzhen Hospital,Guangdong Shenzhen 518000,China.)

机构地区：[1]江苏大学附属人民医院呼吸内科,江苏镇江212000 [2]香港大学深圳医院呼吸内科,广东深圳518000

出　　处：《现代肿瘤医学》2024年第14期2518-2523,共6页Journal of Modern Oncology

基　　金：广东省自然科学基金(编号:2024A1515012336);江苏省镇江市社会发展项目(编号:SH2021032)。

摘　　要：目的:探究基因工程病毒rL-RVG对肺腺癌细胞(PC9)的影响及其潜在机制。方法:通过CCK8实验评估细胞增殖能力,Transwell实验检测细胞迁移和侵袭能力。通过蛋白质印迹分析NLRP3、Caspase1、GSDMD、Asc和IL-1β蛋白表达,免疫荧光染色检测NLRP3的表达。应用Hoechst 33342/PI双染法鉴定细胞焦亡。使用NLRP3抑制剂MCC950预处理验证阻断NLRP3/Caspase1/GSDMD通路对细胞焦亡的影响。结果:rL-RVG处理后,PC9细胞表现出细胞焦亡的形态变化,细胞膨胀、气泡突出、膜破裂和胞质流出胞外。rL-RVG处理可明显抑制PC9细胞增殖,而且抑制效果超过了常规的NDV处理。蛋白质印迹分析显示,在NDV、rL-RVG处理组中,NLRP3、Caspase1-p20、GSDMD-NT、Asc和IL-1β蛋白表达显著上调,而在rL-RVG组中,这些上调效应更为显著。免疫荧光染色结果揭示,在rL-RVG组中,NLRP3的表达高于NDV组和Mock组。另外,rL-RVG处理显著减弱了PC9细胞的迁移和侵袭能力。Hoechst 33342/PI双染显示rL-RVG处理导致PI阳性细胞(红色荧光)显著增加,提示细胞焦亡,其中rL-RVG组效应更为显著。MCC950预处理实验进一步证实,阻断NLRP3/Caspase1/GSDMD信号通路可以减轻rL-RVG诱导的PC9细胞焦亡。结论:rL-RVG通过调控NLRP3/Caspase1/GSDMD通路抑制肺腺癌细胞增殖和诱导焦亡。Objective:To investigate the effect of genetically engineered virus rL-RVG on lung adenocarcinoma cells(PC9)and its underlying mechanisms.Methods:Cell proliferation ability was assessed by CCK8 assay,and cell migration and invasion ability was detected by Transwell assay.NLRP3,Caspase1,GSDMD,Asc and IL-1βprotein expression was analysed by Western blotting,and NLRP3 expression was detected by immunofluorescence staining.Hoechst 33342/PI double staining was applied to identify the pyroptosis.The effect of blocking the NLRP3/Caspase1/GSDMD pathway on the pyroptosis was verified using the NLRP3 inhibitor MCC950 pretreatment.Results:After rL-RVG treatment,PC9 cells exhibited morphological changes of cellular pyroptosis,including cell swelling,bubble protrusion,membrane rupture,and cytoplasmic atrophy.rL-RVG treatment significantly inhibited the proliferation of PC9 cells,and the inhibition effect exceeded that of conventional NDV treatment.Western blotting analysis showed that NLRP3,Caspase1-p20,GSDMD-NT,Asc and IL-1βprotein expressions were significantly up-regulated in the NDV,rL-RVG-treated groups,and these up-regulatory effects were more significant in the rL-RVG group.Immunofluorescence staining revealed that the expression of NLRP3 was higher in the rL-RVG group than in the NDV and Mock groups.In addition,rL-RVG treatment significantly attenuated the migration and invasion ability of PC9 cells.Hoechst 33342/PI double staining showed that rL-RVG treatment resulted in a significant increase in PI-positive cells(red fluorescence),suggesting cellular pyroptosis,with the effect being more pronounced in the rL-RVG group.The MCC950 pretreatment assay further confirmed that blocking NLRP3/Caspase1/GSDMD signalling pathway could attenuate rL-RVG-induced PC9 cell pyroptosis. Conclusion: rL-RVG inhibited lung adenocarcinoma cell proliferation and induced the pyroptosis by regulating the NLRP3/Caspase1/GSDMD pathway.

关 键 词：基因工程病毒 肺腺癌细胞 细胞增殖 细胞迁移 细胞焦亡 

分 类 号：R734.2[医药卫生—肿瘤]