山东大学学报 (医学版) ›› 2024, Vol. 62 ›› Issue (4): 1-8.doi: 10.6040/j.issn.1671-7554.0.2023.1016
• 基础医学 •
沈海涛,乔亚琴,董萍,路燕
SHEN Haitao, QIAO Yaqin, DONG Ping, LU Yan
摘要: 目的 探究Toll样受体4(toll-like receptor 4, TLR4)是否通过调控程序性坏死及铁死亡进一步影响对乙酰氨基酚(acetaminophen, APAP)肝损伤过程及其发生机制。 方法 体外培养人正常肝细胞L-02,采用CCK-8法检测细胞活力,筛选出最佳APAP和TAK-242浓度。将实验分为对照组、APAP组(1、4、12 h)和APAP+TAK-242组(1、4、12 h),比较各组细胞TLR4 mRNA表达;检测各组细胞匀浆液的丙氨酸氨基转移酶(alanine aminotransferase, ALT)和天门冬氨酸氨基转移酶(aspartate aminotransferase, AST)水平;检测各组细胞核因子-κB(nuclear factor-κB, NF-κB)、白细胞介素-6(interleukin-6, IL-6)、肿瘤坏死因子-α(tumor necrosis factor-α, TNF-α)水平;检测各组细胞高迁移率族蛋白1(high mobility group box 1, HMGB1)、受体相互作用蛋白激酶1(receptor interacting protein kinase 1, RIP1)、受体相互作用蛋白激酶3(receptor interacting protein kinase 3, RIP3);检测各组细胞内Fe2+含量以及NF-κB、 P53、重组溶质载体家族7成员11(solute carrier family 7 member 11, SLC7A11)、谷胱甘肽过氧化物酶4(glutathione peroxidase 4, GPX4)水平。 结果 通过CCK-8法,确定5 mmol/L APAP和100 nmol/L TAK-242作为后续实验浓度。与对照组比较,各时间点APAP组TLR4 mRNA水平上调(P<0.05);与APAP组比较,同一时间点APAP+TAK-242组TLR4 mRNA水平下调(P<0.05/3=0.016 7)。与对照组比较,各时间点APAP组ALT、AST水平升高(P<0.05);与APAP组比较,同一时间点APAP+TAK-242组ALT、AST水平下降(P<0.05/3=0.016 7)。与对照组比较,各时间点APAP组NF-κB、IL-6、TNF-α mRNA表达均上调(P<0.05);与APAP组比较,同一时间点APAP+TAK-242组NF-κB、 IL-6、TNF-α mRNA表达均下调(P<0.05/3=0.016 7)。与对照组比较,各时间点APAP组HMGB1、RIP1、RIP3蛋白水平均升高(P<0.05);与APAP组比较,同一时间点APAP+TAK-242组HMGB1、RIP1、RIP3蛋白水平均降低(P<0.05/3=0.0167)。与对照组比较,各时间点APAP组Fe2+含量、NF-κB和P53蛋白水平均上升(P<0.05),而SLC7A11和GPX4蛋白水平和mRNA表达均降低(P<0.05);与APAP组比较,同一时间点APAP+TAK-242组Fe2+含量、NF-κB和P53蛋白水平均降低(P<0.05/3=0.016 7),而SLC7A11和GPX4蛋白水平和mRNA表达均升高(P<0.05/3=0.016 7)。 结论 抑制TLR4可通过调节TLR4/HMGB1信号通路下调程序性坏死,以及可能通过调节TLR4/NF-κB信号通路下调炎症反应和铁死亡来减轻APAP肝损伤。
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