目的：探讨甘草查尔酮A( LA)对吸烟诱导的急性肺损伤小鼠的保护作用及相关机制。方法整体实验：香烟烟雾暴露构建小鼠急性肺损伤模型,取肺泡灌洗液( BALF)进行白细胞计数,测定肺内角质形成细胞衍生趋化因子( KC)、肿瘤坏死因子α( TNF-α)、白介素-1β( IL-1β)和基质金属蛋白酶9(MMP-9)的mRNA和蛋白表达水平,用试剂盒测定肺组织中过氧化物髓化酶( MPO)、超氧化物歧化酶( SOD)活力以及谷胱甘肽( GSH)水平。肺组织病理切片进行HE染色。离体实验：香烟烟雾提取物( CSE)诱导上皮细胞损伤,测定细胞内白介素-8(IL-8)、MMP-9 mRNA的表达。 Western blot分析细胞外信号调节激酶l/2( ERK1/2)、p38分裂原活化蛋白激酶( p38 MAPK)、c-Jun氨基末端激酶( JNK)的磷酸化水平和转录因子NF-κB p65的活性。结果整体实验显示LA组肺内炎症细胞数量及浸润程度均明显低于模型组。模型组肺内KC、TNF-α、IL-1β、MMP-9 mRNA和蛋白表达较正常对照组均明显升高,经LA处理后,上述指标相比模型组明显降低。与正常对照组相比,模型组肺内MPO活性明显上升,而SOD活性和GSH水平明显下降。 LA能逆转这种变化,明显降低MPO活性,升高SOD活性和GSH水平。离体实验显示,LA(2．5、5μmol·L-1)能明显降低CSE刺激的细胞内IL-8和MMP-9 mRNA的高表达。 CSE可以诱导细胞ERK1/2磷酸化和胞核内NF-κB p65的表达,经LA预处理后,上述指标可明显抑制。结论 LA可能通过阻断ERK1/2/NF-κB途径来抑制炎症介质的高表达、调节氧化/抗氧化失衡、下调金属蛋白酶,从而发挥对急性肺损伤小鼠的保护作用。%Aim To explore the protective roles of lic-ochalcone A ( LA) on mice with cigarette smoke-medi-ated acute lung injury and the related mechanisms. Methods In vivo: Mice were exposed to cigarette smoke ( CS) to establish acute lung injury model. The bronchoalveolar lavage fluid ( BALF ) was conducted for cell counting. The mRNA and protein expression of keratinocyte-derived chemokine ( KC ) , tumor necrosis factor alpha ( TNF-α) , interleukin 1β ( IL-1β) and matrix metalloproteinases ( MMP)-9 in lungs were de-termined. The myeloperoxidase ( MPO ) , superoxide dismutase ( SOD ) activities and glutathione ( GSH ) levels in lungs were quantified. The paraffin sections of lungs were prepared and stained with HE. In vitro:Human lung epithelial cells (BEAS-2B) were exposed to cigarette smoke extract ( CSE ) , which induced cell injury. The releases of interleukin 8 (IL-8) and MMP-9 were assessed. The phosphorylation of mitogen-acti-vated protein kinases ( MAPKs, including ERK1/2, p38 and JNK ) and nuclear factor-κB ( NF-κB ) p65 protein were analyzed by Western blot. Results In vi-vo: The accumulation of inflammatory cells was lower in LA groups than that in model group. In comparison with normal control group, the mRNA and protein lev-els of KC, TNF-α, IL-1βand MMP-9 were significant-ly increased in model group. Following treatment with LA, the above indicators were significantly decreased as compared to model group. In the CS-exposed mice, the MPO activity in lungs was significantly increased, meanwhile the SOD activity and GSH level were signif-icantly decreased compared with the air-exposed ani-mals. CS-induced activity of MPO was significantly in-hibited, which were accompanied by increases in SOD and GSH levels by LA. In vitro: CSE-induced mRNA levels of IL-8 and MMP-9 were significantly inhibited by LA at 2 . 5 and 5 μmol · L-1 . The CSE-induced phosphorylation of ERK1/2 and nucleus NF-κB p65 protein expression were prevented by pretreatment with LA. Conclusions LA has protective effects on CS-ex-posed acute lung injury in mice by preventing CS-in-duced pulmonary inflammation, oxidative stress and protease rise. The exploration of the mechanisms sug-gests that LA exerts protective effects via suppressing ERK1/2/NF-κB pathways.
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