TLR4受体在LPS诱导的ApoE^(-/-)小鼠急性肺损伤中的意义

The role of Toll-like receptor 4 on acute lung injury induced by LPS in ApoE^(-/-) mice

目的:研究表明小鼠注射脂多糖(LPS)能形成狼疮样自身抗体及狼疮样肾小球肾炎,成功诱导小鼠狼疮模型。然而,狼疮动脉粥样硬化小鼠中肺损伤的发病机制罕见报道。本研究用LPS(TLR4激动剂)及/或TAK-242(TLR4抑制剂)干预ApoE-/-和C57BL/6小鼠,旨在探讨TLR4受体在肺组织病理变化中的作用。方法:将20只雌性9~10周龄ApoE-/-小鼠随机分为4组,每组5只,分别给予腹腔注射生理盐水、脂多糖(2.5 mg/kg)、脂多糖+TAK-242(脂多糖2.5 mg/kg,并TAK-242 0.3 mg/kg)、TAK-242(0.3 mg/kg),将20只普通雌性C57BL/6(B6)小鼠进行相应处理,每周两次;连续注射4周。然后HE和Masson染色观察肺组织的形态及病理变化;ELISA检测小鼠血清中ANA、抗-dsDNA抗体水平及免疫炎症相关因子IFN-!、TNF-"、IL-1β的水平;免疫组化检测肺组织中TLR4受体、NF-κB p65及BAFF的表达量。结果:ApoE-/-和B6两种小鼠都发生肺毛细管炎、肺间质有不同程度的纤维化、血清ANA和抗-dsDNA抗体的水平升高,并伴有免疫炎症相关因子IFN-!、TNF-"、IL-1β水平升高,肺组织中TLR4、NF-κB p65及BAFF的表达量较盐水组明显升高(P<0.05),上述表现ApoE-/-小鼠皆重于B6小鼠(P<0.01);与脂多糖组相比,TAK-242可明显减轻两种小鼠肺毛细管炎、改善肺间质纤维化,降低血清自身抗体及炎症因子水平,使肺组织中TLR4、NF-κB p65及BAFF的表达量明显减少(P<0.01)。结论:脂多糖在ApoE-/-小鼠较容易诱导出肺毛细管炎,TLR4受体在肺的免疫炎症损伤中起到了重要的作用。

Objective：Studies have indicated Lipopolysaccharide（LPS） injection is a well-established lupus model which mice develop lupus-specific autoantibodies and glomerulonephritis.However,the mechanism of lung injury in lupus with atherosclerosis mice is scarely reported.The purpose of this research was to investigate the pathologic mechanisms of Toll-like receptor 4（TLR4） in lung injury after LPS（TLR4 agonists） and/or TAK-242（TLR4 antagonist） were administered to ApoE-/-and C57BL /6 mice.Methods：Twenty female ApoE-/ mice aged 9-10 weeks were randomized into 4 groups,5 in each group.Saline,LPS（2.5 mg/kg）,TAK-242（0.3 mg/kg）,LPS（2.5 mg/kg） and TAK-242（0.3 mg/kg） were administered respectively by intraperitoneal injection,twice a week.Corresponding interventions were implemented to C57BL /6 mice.After 4 weeks,pathomorphology changes were determined by Hematoxylin-Eosin（HE） and Masson staining in lung paraffin sections.In addition,the levels of antinuclear antibody（ANA）,anti-double-stranded DNA（anti-dsDNA） and inflammatory cytokines（interferon-gamma,tumor necrosis factor,interleukin-1β） were measured by enzyme-linked immunosorbnent assay（ELISA）.TLR4,NF-κB p65 and B cell activating factor belonging to the TNF family（BAFF） in lung specimens were examined by immunohistochemistry.Results：As compared with NS group,pulmonary capillaritis and interstitial fibrosis occurred after LPS intraperitoneal injection in both n ApoE-/-and B6 mice.Meanwhile,the levels of ANA,anti-dsDNA and immune inflammatory mediators（IFN-!,TNF- and IL-1β） rosed significantly.TLR4,NF-κB p65 and BAFF expressions were also increased.However,the above-mentioned manifestations were more serious in ApoE-/-than in B6 mice（P0.01）.After TAK-242 therapy,pulmonary capillaritis and interstitial fibrosis ameliorated markedly.Serum autoantibodies and inflammatory cytokines reduced.And TLR4,NF-κB p65 and BAFF expressions were obviously decreased（P0.01）.Conclusion：Pulmonary capillaritis induced by LPS is more obvious in ApoE-/-than in C57BL /6 mice.And TLR4 is crucial for its development of immune and inflammatory injury.