右美托咪定对高迁移率族蛋白1诱导人脐静脉内皮细胞炎性反应的影响  

Effects of Dexmedetomidine on HMGB1-induced Inflammatory Responses in HUVECs

作　　者：张志发[1] 朱梦娇 杨少兵[1] 陶怡芝 王学仁 Zhang Zhifa;Zhu Mengjiao;Yang Shaobing(Department of Anesthesiology,Tongji Hospital,Tongji Medical College,Huazhong University of Scienceand Technology,Wuhan 430030,China)

机构地区：[1]华中科技大学同济医学院附属同济医院麻醉科,武汉430030

出　　处：《华中科技大学学报（医学版）》2022年第1期38-43,共6页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金资助项目(No.81371251);湖北省陈孝平科技发展基金资助项目;华中科技大学自主创新研究基金资助项目(No.2016YXMS122)。

摘　　要：目的研究右美托咪定对脂多糖(LPS)刺激人脐静脉内皮细胞(HUVECs)分泌高迁移率族蛋白1(HMGB1)的影响,及其调控HMGB1诱导HUVECs炎性反应的分子机制。方法 100 ng/mL的LPS刺激HUVECs 16 h后,用不同浓度的右美托咪定(0.01、0.1、1、10 nmol/L)分别处理细胞4 h,酶联免疫吸附实验(ELISA)检测细胞上清液中HMGB1的含量。MTT检测不同浓度右美托咪定对HUVECs细胞活力的影响,细胞免疫荧光检测HMGB1的表达及分布。将HUVECs随机分为对照组、右美托咪定组、HMGB1组、右美托咪定+HMGB1组,右美托咪定和HMGB1的浓度分别是1 nmol/L、1μg/mL。ELISA检测细胞上清液中TNF-α和IL-1β含量,Western blotting检测MAPK信号通路的磷酸化水平,再采用ERK1/2、p38 MAPK通路抑制剂进行阻断实验。结果不同浓度的右美托咪定对HUVECs活力无影响。LPS作用后,HUVECs上清液中HMGB1的含量增加(P<0.01),而右美托咪定可呈剂量依赖性地抑制HMGB1的表达(P<0.05);LPS刺激HMGB1蛋白从细胞核转移至胞质,而右美托咪定可显著抑制LPS引起的HMGB1移位(P<0.05)。相对于对照组,HMGB1组上清液中炎症因子TNF-α和IL-1β含量增加(均P<0.01),外源性HMGB1促进ERK1/2、p38 MAPK和JNK磷酸化(均P<0.05),而右美托咪定+HMGB1组可抑制上述炎症因子的分泌并部分逆转ERK1/2、p38 MAPK的磷酸化(均P<0.05)。p38 MAPK抑制剂(SB203580)可下调HMGB1组IL-1β的表达,ERK1/2阻滞剂(PD98059)可下调TNF-α和IL-1β的表达,且与右美托咪定具有协同抑制TNF-α和IL-1β表达的作用(均P<0.05)。结论右美托咪定可抑制LPS引起的HMGB1分泌,并通过抑制HMGB1相关的MAPK信号通路发挥抗炎效应。Objective To investigate effect of dexmedetomidine on the secretion of high-mobility group box-1 protein(HMGB1)in lipopolysaccharide(LPS)-stimulated human umbilical vein endothelial cells(HUVECs),and the possible molecular mechanism of dexmedetomidine-mediated inflammatory response in HUVECs with HMDB1 stimulation.Methods After incubation with 100 ng/mL LPS for 16 h, HUVECs were treated with dexmedetomidine of different concentrations(0.01,0.1,1,10 nmol/L)for 4 h.Levels of HMGB1 in supernatant of HUVECs were evaluated by enzyme linked immunosorbent assay(ELISA).The effect of dexmedetomidine on cell viability was examined by MTT,and the expression levels and distributions of HMGB1 in HUVECs were detected by immunofluorescent staining.HUVECs were randomly divided into control group, dexmedetomidine group, HMGB1 group, and dexmedetomidine+ HMGB1 group, and the final concentrations of dexmedetomidine and HMGB1 were 1 nmol/L and 1 μg/mL,respectively.TNF-α and IL-1β in supernatant of HUVECs were examined by ELISA,and the phosphorylated status of MAPK signaling pathway was determined by Western blotting.Inhibitors against ERK1/2 and p38 MAPK were used for blocking experiments.Results Dexmedetomidine showed nontoxic effect on viability of HUVECs.LPS stimulation increased the expression level of HMGB1 protein in supernatant of HUVECs(P<0.01),while dexmedetomidine had a dose-dependent suppressive effect on HMGB1 expression(P<0.05).The translocation of HMGB1 from cell nucleus to cytoplasm was verified in LPS-treated HUVECs, which was markedly inhibited by dexmedetomidine(P<0.05).Compared to the control group, the content of TNF-α and IL-1β in cell supernatant(both P<0.01),and the phosphorylation of ERK1/2,p38 MAPK and JNK(all P<0.05)were significantly increased in HMGB1 group, and those changes were partly reversed in dexmedetomidine+HMGB1 group(all P<0.05).Moreover, p38-MAPK inhibitor(SB203580)suppressed IL-1β expression, and ERK1/2 inhibitor(PD98059)attenuated the expression of both TNF-α and IL-1β,showing synergi

关 键 词：右美托咪定 脂多糖 人脐静脉内皮细胞 高迁移率族蛋白1 

分 类 号：R971.2[医药卫生—药品]