柚皮素对博莱霉素诱导的小鼠肺纤维化的改善作用及其作用机制

Effect of naringin on bleomycin-induced pulmonary fibrosis in mice and its mechanism

目的探讨柚皮素(NAR)对博来霉素诱导小鼠肺纤维化的影响及其可能作用机制。方法将60只雄性ICR小鼠随机分成正常对照组(Control)、模型组(Model)、阳性对照地塞米松组(DXM,3 mg/kg)、NAR低剂量组(L-NAR,25 mg/kg)和NAR高剂量组(H-NAR,100 mg/kg),每组12只。采用气管内一次性滴注博莱霉素(5 mg/kg)建立小鼠肺纤维化模型,药物干预4周后处死小鼠。取肺组织称重,计算肺指数;碱水解法测定肺组织羟脯氨酸(HYP)含量;HE染色观察肺组织病理改变并评分;Masson染色观察肺纤维化程度并评分;ELISA法检测肺组织中白细胞介素1β(IL-1β)、白细胞介素18(IL-18)和转化生长因子β1(TGF-β1)水平;RT-PCR检测肺组织中NLRP3、ASC和caspase-1 mRNA表达水平;Western blot检测肺组织中NLRP3、ASC、caspase-1、Collagen I、α-SMA、E-cadherin和Vimentin蛋白表达水平。结果与Control组比较,Model组小鼠肺组织损伤明显,纤维化程度较为严重,肺指数和肺组织中HYP、IL-1β、IL-18、TGF-β1含量增加,NLRP3、ASC、caspase-1的mRNA和蛋白表达水平以及Collagen I、α-SMA和Vimentin蛋白表达水平均上升,而E-cadherin蛋白表达水平下降,差异有统计学意义(P<0.05);与Model组比较,DXM组和H-NAR组小鼠肺损伤与纤维化程度得到明显改善,小鼠肺指数、肺组织中HYP、IL-1β、IL-18、TGF-β1含量,NLRP3、ASC、caspase-1 mRNA和蛋白表达水平以及Collagen I、α-SMA和Vimentin蛋白表达水平均下降,E-cadherin蛋白表达水平上升,差异有统计学意义(P<0.05)。而L-NAR组与Model组比较,相关指标差异无统计学意义。结论 NAR能有效抑制博莱霉素诱导的小鼠肺纤维化过程,其机制可能与抑制NLRP3炎症小体活化有关。

Objective To investigate the effect of Naringin(NAR) on bleomycin-induced pulmonary fibrosis in mice and its possible mechanism.Methods Sixty male ICR mice were randomly divided into Control group,Model group,dexamethasone group(DXM,3 mg/kg),low-dose naringin group(L-NAR,25 mg/kg) and high-dose naringin group(H-NAR,100 mg/kg),with 12 mice in each group.Mouse pulmonary fibrosis model was established by intratracheal one-time infusion of bleomycin(5 mg/kg),and mice were sacrificed after 4 weeks of drug intervention.Lung mass was taken and lung index was calculated.Levels of hydroxyproline(HYP) in lung tissue were detected by alkaline hydrolysis.The pathological changes of lung tissues were observed and scored by HE staining.The degree of pulmonary fibrosis was observed and scored by Masson staining.The levels of IL-1β,IL-18,TGF-β1 in lung tissue were detected by ELISA.The expression levels of NLRP3,ASC and caspase-1 mRNA in lung tissues were detected by RT-PCR.The protein expression levels of NLRP3,ASC,caspase-1,Collagen I,α-SMA,E-cadherin and Vimentin in lung tissues were detected by Western blot.Results Compared with the Control group,the Model group showed significant lung tissue damage and severe degree of fibrosis,the lung index and lung tissue contents of HYP,IL-1β,IL-18 and TGF-β1 increased.The mRNA and protein expression levels of NLRP3,ASC,caspase-1,as the well as the protein expression levels of Collagen I,α-SMA and Vimentin increased,while the expression levels of E-cadherin significantly decreased,and the difference was statistically significant(P<0.05).Compared with the Model group,the degree of lung injury and fibrosis were significantly improved in the DXM group and the H-NAR group.The lung index and lung tissue contents of HYP,IL-1β,IL-18 and TGF-β1 decreased.The mRNA and protein expression levels of NLRP3,ASC,caspase-1,as the well as the protein expression levels of Collagen I,α-SMA and Vimentin decreased,while the expression levels of E-cadherin increased,and the difference was statistically significant(P<0.05).However,there was no significant difference in relevant indicators between the L-NAR group and the Model group.Conclusion NAR can effectively inhibit bleomycin-induced pulmonary fibrosis in mice,and the mechanism may be related to the inhibition of NLRP3 inflammatory corpuscles activation.