异丁司特对脂多糖诱导小鼠急性呼吸窘迫综合征的保护作用及机制

Protective effect and mechanism of Ibudilast on LPS-induced acute respiratory distress syndrome in mice

目的:探讨异丁司特(AV-411)对脂多糖(LPS)诱导的急性呼吸窘迫综合征(ARDS)小鼠的保护作用及作用机制。方法:将24只BALB/c小鼠随机分为正常组、模型组及治疗组,每组8只。模型组和治疗组小鼠尾静脉注射LPS诱导建立ARDS小鼠模型,造模成功后治疗组小鼠给予15 mg/kg AV-411灌胃,正常组及模型组小鼠用等量生理盐水灌胃,3组均3次/d,治疗15 d后处死。经苏木素-伊红(HE)染色观察3组小鼠肺组织病理变化;采用酶联免疫吸附试验(ELISA)检测3组小鼠肺泡灌洗液中白介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)、白介素-10(IL-10)等炎症因子水平;采用实时定量聚合酶链反应(qRT-PCR)及免疫蛋白印迹(WB)检测AV-411对组织Toll样受体4(TLR4)表达影响及3组小鼠肺组织中IL-1β、TNF-α、IL-10、核因子κB的抑制蛋白α(IκB-α)的mRNA及蛋白表达水平。结果:治疗组小鼠肺泡、肺泡隔结缔组织炎性细胞浸润较模型组显著减少;与模型组比较,治疗组小鼠肺组织中TLR4蛋白及TLR4 mRNA表达水平显著降低,肺泡灌洗液中IL-1β、TNF-α水平显著降低,IL-10、IκB-α水平升高;肺组织IL-1β、TNF-α的mRNA及蛋白表达量显著降低(均P<0.05)。结论:AV-411可能通过诱导ARDS小鼠肺组织TLR4表达降低,抑制IκB-α蛋白降解及相关炎症因子表达,发挥对ARDS小鼠的治疗作用。

Objective:To investigate the protective effect and mechanism of Ibudilast(AV-411)on lipopolysaccharide(LPS)-induced acute respiratory distress syndrome(ARDS)in mice.Methods:Twenty-four BALB/c mice were randomly divided into normal group,model group and treatment group,8 rats each.Mice in the model group and the treatment group were induced by injecting LPS to establish the ARDS model.After modeling,mice in the treatment group were given 15 mg/kg AV-411 for 15 days,and the rest two groups were given the same amount of normal saline,mice in 3 groups were given three times per day,and were killed after 15 days of treatment.The morphological changes of lung tissues were observed by HE staining.Enzyme-linked immunosorbent assay(ELISA)was used to test the content of interleukin-1β(IL-1β),tumor necrosis factor-α(TNF-α),IL-10 in bronchoalveolar lavage fluid(BALF).Quantitative real-time PCR(qRT-PCR)and Western blotting were employed to detect mRNA and protein expression levels of IL-1β,TNF-α,IL-10 and inhibitor of NFκB-α(IκB-α).Results:HE staining showed that AV-411 effectively relieved the pathological changes in the treatment group and the inflammatory cells infiltration was significantly reduced as compared with the model group.The mRNA and protein levels of TLR4,IL-1β,TNF-αin lung tissues of ARDS mice in the treatment group were significantly decreased as compared with those in the model group.Besides,AV-411 promoted the expression of IL-10 and IκB-αprotein in lung tissues of mice in the treatment group,which was significantly different from the model group,but had no effect on the expression level of IκB-αmRNA.The tendency of IL-1β,TNF-αand IL-10 levels in BALF was consistent with mRNA levels in lung tissues.Conclusion:AV-411 may inhibit the degradation of IκB-αand the expression of inflammatory factors by inducing the expression of TLR4 in lung tissues in ARDS mice.