基于NLRP3/Caspase-1/GSDMD信号通路研究生骨再造丸对BMSCs焦亡水平的干预效应
Based on the NLRP3/Caspase-1/GSDMD signaling pathway intervene effect of Shenggu Zaizao Pills on pyroptosis of BMSCs
张勇杰 1曹林忠2
作者信息
- 1. 甘肃中医药大学,甘肃兰州 730000
- 2. 甘肃中医药大学,甘肃兰州 730000;甘肃中医药大学附属医院,甘肃兰州 730000
- 折叠
摘要
目的 研究生骨再造丸通过NLRP3/Caspase-1/GSDMD信号通路对BMSCs焦亡水平的影响.方法 购买原代兔BMSCs,制备焦亡BMSCs模型,采用慢病毒技术及siRNA技术转染焦亡的BMSCs;制备生骨再造丸含药血清,对焦亡的BMSCs进行干预.将上述细胞分为空白组、模型组、慢病毒组、siRNA组、含药血清组.通过PCR检测各组BMSCs中GS-DMD基因的表达水平;WB检测各组BMSCs中NLRP3、Caspase-1、GSDMD的表达水平;ELISA检测各组BMSCs中IL-1β、IL-18水平;通过CCK-8检测BMSCs活力.结果 与空白组相比,模型组GSDMD基因表达增加(P＜0.01),NL-RP3、Caspase-1、GSDMD 蛋白水平显著升高(P＜0.05)(P＜0.01)(P＜0.01),IL-18、IL-1β水平上升(P＜0.01)(P＜0.01).与模型组相比,慢病毒组BMSCs中GSDMD基因表达升高(P＜0.01),NLRP3、Caspase-1、GSDMD蛋白水平增加(P＜0.05)(P＜0.05)(P＜0.05),IL-18、IL-1β 水平明显增加(P＜0.01)(P＜0.05);siRNA 组 BMSCs 中 GSDMD 基因表达显著降低(P＜0.01),NLRP3、Caspase-1、GSDMD 蛋白水平降低(P＜0.01)(P＜0.05)(P＜0.05),IL-18、IL-1β水平明显降低(P＜0.01)(P＜0.01);含药血清组 BMSCs 中 GSDMD 基因表达降低(P＜0.05),NLRP3、Caspase-1、GSD-MD 蛋白水平降低(P＜0.05)(P＜0.05)(P＜0.01),IL-1β、IL-18 水平降低(P＜0.05)(P＜0.05).结论 NLRP3 是调控BMSCs焦亡的关键靶点,生骨再造丸能通过降低NLRP3表达,抑制NLRP3/Caspase-1/GSDMD信号通路,调控BMSCs焦亡水平.
Abstract
Objective To study the effect of bone reconstruction pills on the level of pyroptosis in BMSCs through NLRP3/Caspase-1/GSDMD signaling pathway.Methods Pyroptosis BMSCs models were prepared,and lentivirus and siRNA technolo-gies were used to transfect pyroptosis BMSCs.The medicated serum of raw bone reconstruction pills was prepared to intervene in the BMSCs of pyroptosis.The above cells were divided into blank group,model group,lentivirus group,siRNA group and drug-containing serum group.CCK-8 was used to detect the activity of BMSCs in each group,and the expression level of GSDMD gene in BMSCs was detected by RT-qPCR.WB was used to detect the expression levels of NLRP3,Caspase-1 and GSDMD in BMSCs.ELISA was used to detect the levels of IL-1β and IL-18 in BMSCs.Results Compared with the blank group,the ex-pression of GSDMD gene in the model group increased(P＜0.01),the protein levels of NLRP3,Caspase-1 and GSDMD were significantly increased(P＜0.05)(P＜0.01)(P＜0.01),and the levels of IL-18 and IL-1 β increased(P＜0.01)(P＜0.01).Compared with the model group,the expression of GSDMD gene in BMSCs in the lentivirus group increased(P＜0.01),the protein levels of NLRP3,Caspase-1 and GSDMD increased(P＜0.05)(P＜0.05)(P＜0.05),the levels of IL-18 and IL-1β increased significantly(P＜0.01)(P＜0.05),and the expression of GSDMD gene in BMSCs in the siRNA group de-creased significantly(P＜0.01),NLRP3,The protein levels of Caspase-1 and GSDMD decreased(P＜0.01)(P＜0.05)(P＜0.05),the levels of IL-18 and IL-1 β decreased significantly(P＜0.01)(P＜0.01),the expression of GSDMD gene in BMSCs in the drug-containing serum group decreased(P＜0.05),NLRP3,Caspase-1,The levels of GSDMD protein de-creased(P＜0.05)(P＜0.05)(P＜0.01),and the levels of IL-1β and IL-18 decreased(P＜0.05)(P＜0.05).Conclu-sion NLRP3 is a key target for regulating pyroptosis of BMSCs,and bone reconstruction pills can regulate the level of pyroptosis of BMSCs by reducing the expression of NLRP3 and inhibiting the NLRP3/Caspase-1/GSDMD signaling pathway.
关键词
生骨再造丸/NLRP3/Caspase-1/GSDMD信号通路/骨髓间充质干细胞/焦亡Key words
Shenggu Zaizao Pills/NLRP3/Caspase-1/GSDMD signaling pathway/bone marrow mesenchymal stem cells/py-roptosis引用本文复制引用
基金项目
国家自然科学基金(82160915)
甘肃省兰州市科技局项目(2022-3-23)
甘肃中医药大学研究生创新基金(2023CX14)
出版年
2024
国家科技期刊平台
NSTL
万方数据