摘要
目的探讨丹参新醌乙对氧化低密度脂蛋白(ox-LDL)诱导的人脐静脉内皮细胞损伤的影响及机制。方法取对数生长期的人脐静脉内皮细胞,分为正常组(加入10%胎牛血清)、ox-LDL组(在正常组基础上加入100ug/mL的ox-LDL)、二甲基亚砜(DMSO)组(在ox-LDL组基础上加入0.1%浓度DMSO)、丹参新醌乙组(在ox-LDL组基础上加入由DMSO溶解的100ng/ml的丹参新醌乙),培养24 h后进行实验。微板法检测乳酸脱氢酶(LDH)表达;qRT-PCR法检测NF-κB1、核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、消皮素D(GSDMD)、白介素(IL)-1β的mRNA表达;WesternBlot法检测NF-κB1、NLRP3、caspase-1、GSDMD-N的蛋白表达。免疫荧光法检测GSDMD表达情况。结果与正常组比较,ox-LDL组、DMSO组LDH表达明显升高(P<0.01),NF-κB1、NLRP3、GSDMD、IL-1β的mRNA水平均显著上调(P<0.01),NF-κB1、NLRP3、caspase-1、GSDMD-N、IL-1β的蛋白表达水平明显升高(P<0.01);与ox-LDL组、DMSO组比较,丹参新醌乙组LDH表达有所降低(P<0.05),NF-κB1、NLRP3、GSDMD、IL-1β的mRNA水平显著下调(P<0.01),NF-κB1、NLRP3、caspase-1、GSDMD-N、IL-1β的蛋白水平显著降低(P<0.01),GSDMD在质膜表达抑制,入核受限。结论丹参新醌乙可能通过NF-κB/NLRP3信号通路,抑制NLRP3炎症小体介导的细胞焦亡,改善ox-LDL诱导的内皮细胞损伤。
Objective To investigate the protective effect of Danshenxinkun B against oxidized low-density lipoprotein(ox-LDL)-induced human umbilical vein endothelial cell(HUVEC)injury and explore the underlying mechanism.Methods HUVECs cultured in the presence of 10%fetal bovine serum were treated with ox-LDL(100μg/mL),ox-LDL+0.1%dimethyl sulfoxide(DMSO),or ox-LDL+Danshenxinkun B(100 ng/mL,dissolved in DMSO)for 24 h.The changes in lactate dehydrogenase(LDH)release was detected,and qRT-PCR was used to detect the mRNA expressions of nuclear factor-κB1(NF-κB1),nucleotide binding oligomerization domain-like receptor family pyrin domain protein 3(NLRP3),gasdermin D(GSDMD)and interleukin 1β(IL-1β).The protein expressions of NF-κB1,NLRP3,caspase-1,IL-1βand GSDMD-N were detected with Western blotting.Immunofluorescence assay was performed to examine the changes in GSDMD expression in the cells.Results Compared with the normal control cells,the cells treated with ox-LDL alone or in combination with DMSO exhibited significantly increased LDH release,mRNA expressions of NF-κB1,NLRP3,GSDMD,and IL 1βand the protein levels of NFκB1,NLRP3,IL 1β,GSDMD-N and caspase-1(P<0.01),which were all significantly lowered by treatment with Danshenxinkun B(P<0.05 or 0.01).Danshenxinkun B treatment significantly inhibited GSDMD expression on the cell membrane and restricted its entry into the cell nucleus.Conclusion Danshenxinkun B alleviates ox-LDL-induced HUVEC injury possibly by suppressing pyroptosis mediated by NLRP3 inflammatory bodies and inhibiting the NF-κB/NLRP3 signaling pathway.
作者
李洪涛
邓宇
王添乐
黄克勇
于传沛
陈朝俊
LI Hongtao;DENG Yu;WANG Tianle;HUANG Keyong;YU Chuanpei;CHEN Chaojun(Guangzhou University of Chinese Medicine,Guangzhou 510006,China;Township Center Hospital of Xiaosanjiang Town,Qingyuan 513224,China;Department of Neurology,Guangzhou Hospital of Integrated Traditional and West Medicine Affiliated to Guangzhou University of Chinese Medicine,Guangzhou 510800,China)
出处
《南方医科大学学报》
CAS
CSCD
北大核心
2023年第8期1425-1431,共7页
Journal of Southern Medical University
基金
广东省中医药局科研项目(20213014)。
作者简介
李洪涛,硕士,E-mail:leihungtou@outlook.com;通信作者:陈朝俊,主任中医师,教授,博士生导师,E-mail:ccjbs@126.com。
