BET抑制剂(+)-JQ1通过调节NF-KB信号通路保护小鼠急性肝衰竭实验研究

Protective effect of bromo-and extra-terminal domain inhibitor(+),JQ1 on LPS/D-Gal-induced acute liver failure in mice

目的探讨溴结构域和额外末端结构域(BET)抑制剂(+)-JQ1对脂多糖/D-氨基半乳糖(LPS/D-Gal)诱导的小鼠急性肝衰竭(ALF)的保护作用及其作用机制。方法将47只C57BL/6小鼠随机分为对照组(n=15)、模型组(n=17)和(+)-JQ1干预组(n=15),采用腹腔注射LPS/D-Gal联合诱导ALF模型,其中在干预组造模前2 h腹腔注射(+)-JQ1。在造模4 h后每组处死5只小鼠,其余小鼠继续喂养至72 h,观察存活率。采用ELISA法检测血清TNF-α,采用RT-qPCR法和Western bloting法分别检测mRNA和蛋白表达。结果模型组72 h小鼠存活率为16.7%(2/12),(+)-JQ1干预组为60.0%(6/10,P>0.05);ALF组血清ALT水平为(2779.0±200.6)U/L,显著高于对照组[(44.9±2.5)U/L,P<0.05],血清AST水平为(2885.0±143.6)U/L,显著高于对照组[(76.0±5.7)U/L,P<0.05],而(+)-JQ1干预组血清ALT和AST水平较模型组显著降低[分别为(948.7±46.3)U/L和(1704.0±42.1)U/L,P<0.05];ALF组肝组织TNF-αmRNA水平为(103.7±23.0),显著高于对照组[(1.2±0.2),P<0.05],IL-6 mRNA水平为(73.4±15.8),显著高于对照组[(1.1±0.1),P<0.05],IL-1B mRNA水平为(13.5±2.7),显著高于对照组[(1.0±0.1),P<0.05],而经(+)-JQ1干预后,三种细胞因子mRNA水平均较模型组显著降低[分别为(12.8±1.2)、(11.7±0.7)和(1.5±0.3),P<0.05];ALF组血清TNF-α水平(631.6±57.5)U/L,显著高于对照组[(2.5±0.5)U/L,P<0.05],而(+)-JQ1干预组血清TNF-α水平为(139.8±8.1)U/L,显著低于ALF组(P<0.05);ALF组肝组织P65和IKB蛋白表达显著增强,而(+)-JQ1干预后显著减弱。结论(+)-JQ1对ALI小鼠肝脏具有保护作用,可能通过调节NF-KB信号通路下调促炎细胞因子的表达,从而减轻肝组织炎症反应。

Objective The aim of this experiment was to investigate the protective effect of bromo-and extra-terminal domain(BET)inhibitor,(+)-JQ1 on lipopolysaccharide/D-galactose(LPS/D-Gal)-induced acute liver failure(ALF)in mice and its mechanism.Methods 47 C57BL/6 mice were randomly divided into control(n=17),ALF model(n=15)and(+)-JQ1 intervention group(n=15).The ALF model were established by intraperitoneal injection of LPS/D-Gal,and mice in(+)-JQ1 intervention group were injected intraperitoneally with(+)-JQ12 hours before the induction of ALF.Four hours after LPS/D-Gal injection,5 mice were sacrificed in each group and the remaining mice were fed for 72 hours to observe 72-hour survival rate.Serum TNF-αwas detected by ELISA,and hepatic protein and gene mRNA were assayed by RT-qPCR and Western bloting,respectively.Results The 72-hour survival rate in model group was 16.7%(2/12),not significantly different compared to 60.0%(6/10,P>0.05)in(+)-JQ1-intervened group;serum ALT level in mice with ALF was(2779.0±200.6)U/L and serum AST level was(2885.0±143.6)U/L,both significantly higher than[(44.9±2.5)U/L and(76.0±5.7)U/L,P<0.05]in control,while they decreased significantly to[(948.7±46.3)U/L and(1704.0±42.1)U/L,respectively,P<0.05]in(+)-JQ1-intervened group;the hepatic tissue TNF-αmRNA was(103.7±23.0),significantly higher than[(1.2±0.2),P<0.05],the IL-6 mRNA was(73.4±15.8),much higher than[(1.1±0.1),P<0.05],and the IL-1B mRNA was(13.5±2.7),significantly higher than[(1.0±0.1),P<0.05]in the control,while they decreased greatly to[(12.8±1.2),(11.7±0.7)and(1.5±0.3),respectively,P<0.05]in(+)-JQ1-intervened group;serum TNF-αlevel was(631.6±57.5)U/L,much higher than[(2.5±0.5)U/L,P<0.05]in the control,while it decreased to[(139.8±8.1)U/L,P<0.05]in(+)-JQ1-intervened group;the expression of P65 and IKB in hepatic tissues in model intensified obviously,while they weaken greatly in(+)-JQ1-intervened group.Conclusion The bromo-and extra-terminal domain(BET)inhibitor,(+)-JQ1 has a protective effect on the liver in mice with LPS/D-Gal-induced ALF,which might be related to the regulation of NF-KB signaling pathway to reduce the expression of proinflammatory cytokines.