银杏素调节NF-κB/NLRP3/Caspase-1信号通路对高糖诱导肾小球内皮细胞焦亡的影响  

Ginkgo biloba extract regulates high-glucose-induced pyroptosis in glomerulaRendothelial cells by mediating the NF-κB/NLRP3/Casase-1 signaling pathway

作　　者：温洁[1] 高颖颖[1] WEN Jie;GAO Yingying(Baoji People’s Hospital,Shaanxi,Baoji 721000,China)

机构地区：[1]陕西省宝鸡市人民医院,721000

出　　处：《河北医药》2023年第15期2251-2255,2260,共6页Hebei Medical Journal

基　　金：陕西省自然科学基础研究计划项目(编号:S2019-JC-YB-1405)。

摘　　要：目的探讨银杏素调节核转录因子кB/Nod样受体蛋白3/半胱天冬酶-1(NF-κB/NLRP3/Caspase-1)信号通路对高糖诱导肾小球内皮细胞焦亡的影响。方法将肾小球内皮细胞(HRGECs)作为研究对象,将HRGECs细胞分为正常组(正常5 mmol/L葡萄糖)、高糖组(30 mmol/L葡萄糖)、银杏素低剂量组(30 mmol/L葡萄糖+5μmol/L银杏素)、银杏素中剂量组(30 mmol/L葡萄糖+10μmol/L银杏素)、银杏素高剂量组(30 mmol/L葡萄糖+20μmol/L银杏素)、BAY组[(30 mmol/L葡萄糖+20μmol/L银杏素+2.5μmol/L NF-κB的特异性抑制剂BAY 11-7085(BAY))]、MCC950组(30 mmol/L葡萄糖+20μmol/L银杏素+10μmol/L NLRP3抑制剂MCC950)、VX-765组(30 mmol/L葡萄糖+20μmol/L银杏素+20μmol/L Caspase-1抑制剂VX-765),各组细胞加入相应试剂后共培养24 h;MTT法检测细胞存活率;流式细胞术检测细胞焦亡情况;试剂盒法测定细胞上清IL-1β、IL-18、LDH水平;qRT-PCR法和Western blot法检测细胞NF-κB、NLRP3、Caspase-1、GSDMD的表达。结果与正常组比较,高糖组细胞存活率明显下降(P<0.05),PI+Caspase-1阳性细胞比例、细胞上清IL-1β、IL-18、LDH水平以及细胞NF-κB p65、NLRP3、Caspase-1、GSDMD表达明显升高(P<0.05);与高糖组比较,银杏素低、中、高剂量组细胞存活率明显升高(P<0.05),PI+Caspase-1阳性细胞比例、细胞上清IL-1β、IL-18、LDH水平以及细胞NF-κB p65、NLRP3、Caspase-1、GSDMD表达明显降低(P<0.05);分别使用NF-κB、NLRP3、Caspase-1特异性抑制剂BAY、MCC950、VX-765进一步升高细胞存活率,降低细胞PI+Caspase-1阳性细胞比例、细胞上清IL-1β、IL-18、LDH水平以及细胞NF-κB p65、NLRP3、Caspase-1、GSDMD表达。结论银杏素可通过抑制NF-κB/NLRP3/Caspase-1信号通路抑制高糖诱导的肾小球内皮细胞焦亡。Objective To explore the effect of ginkgo biloba extract on high-glucose-induced pyroptosis in glomerulaRendothelial cells by regulating the nucleaRtranscription factor-κB/Nod like receptoRprotein 3/caspase-1(NF-κB/NLRP3/caspase-1)signaling pathway.Methods Human glomerulaRendothelial cells(HRGECs)were treated with 5mmol/L glucose(control group),30mmol/L glucose(high glucose group),30mmol/L glucose+5μmol/L ginkgo biloba extract(low-dose ginkgo biloba group),30mmol/L glucose+10μmol/L ginkgo biloba extract(medium-dose ginkgo biloba group),30mmol/L glucose+20μmol/L ginkgo biloba extract(high-dose ginkgo biloba group),30mmol/L glucose+20μmol/L ginkgo biloba extract+2.5μmol/L NF-κB specific inhibitoRBAY 11-7085(BAY group),30mmol/L glucose+20μmol/L ginkgo biloba extract+10μmol/L NOD-like receptoRprotein 3(NLRP3)inhibitoRMCC950(MCC950 group)and 30mmol/L glucose+20μmol/L ginkgo biloba extract+20μmol/L Caspase-1 inhibitoRVX-765(VX-765 group)foR24h.Cell survival and pyroptosis were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide(MTT)assay and flow cytometry,respectively.Relative levels of IL-1,IL-18 and lactate dehydrogenase(LDH)in cell supernatant were measured by commercial enzyme-linked immunosorbent assay(ELISA)kits.The mRNA and protein expressions of NF-κB,NLRP3,Caspase-1 and gasdermin D(GSDMD)were detected by quantitative real-time PCR(qRT-PCR)and Western blot,respectively.Results Compared with those of control group,significantly decreased survival,and increased proportion of PI+Caspase-1 positive cells,relative levels of IL-1β,IL-18 and LDH in cell supernatant,and expression levels of NF-κB p65,NLRP3,Caspase-1 and GSDMD were detected in high glucose group(P<0.05).Compared with those of high glucose group,significantly increased survival,and decreased proportion of PI+Caspase-1 positive cells,relative levels of IL-1β,IL-18 and LDH in cell supernatant,and expression levels of NF-κB p65,NLRP3,Caspase-1 and GSDMD were detected in low-dose,medium-dose and high-dose ginkgo bil

关 键 词：银杏素 NF-κB/NLRP3/Caspase-1 高糖 肾小球内皮细胞 细胞焦亡 

分 类 号：R587.2[医药卫生—内分泌]