NDNF低表达对博来霉素致肺纤维化小鼠炎症反应、氧化应激和EMT的影响  

Effect of silencing NDNF expression on inflammatory response,oxidative stress,and EMT in mice with bleomycin-induced pulmonary fibrosis

作　　者：冯碧莹 李平 杜玮 FENG Biying;LI Ping;DU Wei(Department of Respiratory and Critical Care Medicine,PLA Southern Theater Command General Hospital,Guangzhou 510010,Guangdong,China)

机构地区：[1]南部战区总医院呼吸与危重症医学科,广东广州510010

出　　处：《贵州医科大学学报》2024年第5期665-671,共7页Journal of Guizhou Medical University

基　　金：广东省中医药局科研项目资助项目(20211185);广州市科技计划项目(202102080518)。

摘　　要：目的探讨神经元源性神经营养因子(NDNF)低表达对博来霉素致肺纤维化小鼠肺炎症反应、氧化应激和上皮-间质转分化(EMT)的影响。方法采取气管注入博来霉素(5 mg/kg)构建肺纤维化小鼠模型(肺纤维化组)、尾静脉注射siRNA-NDNF腺病毒载体建立NDNF低表达肺纤维化组(siRNA-NDNF组)、同时设置空载体组及健康组;于造模成功后2周时,采用HE及Masson染色观察小鼠肺组织病理变化(肺泡炎症评分)和肺纤维程度(肺纤维化占比),ELISA法检测肺组织炎性因子[肿瘤坏死因子-α(TNF-α)、转化生长因子-β1(TGF-β1)、白细胞介素(IL)-4和IL-6]、氧化应激因子[丙二醛(MDA)、活性氧(ROS)和超氧化物歧化酶(SOD)]及羟脯氨酸(HYP)水平,Western blot检测各组小鼠肺组织EMT相关蛋白[E-钙黏蛋白(E-cadherin)、α平滑肌肌动蛋白(α-SMA)、Snail1和Collagen I]表达。结果肺纤维化组小鼠肺泡结构破坏程度严重、肺泡残缺严重、明显炎症浸润、大面积胶原纤维沉积,肺系数、肺纤维化占比、肺泡炎症评分和肺组织HYP水平均高于健康组(P<0.05);siRNA-NDNF组小鼠肺泡结构损伤、炎性浸润和胶原纤维蓝色区域较空载体组减少,肺系数、肺纤维化占比、肺泡炎症评分和肺组织HYP水平低于空载体组(P<0.05);肺纤维化组小鼠肺组织TNF-α、TGF-β1、IL-6、IL-4、MDA、ROS水平和α-SMA、Snail1、Collagen I蛋白表达均高于健康组(P<0.05),肺组织SOD水平和E-cadherin蛋白表达低于健康组(P<0.05);siRNA-NDNF组小鼠肺组织TNF-α、TGF-β1、IL-6、IL-4、MDA、ROS水平和α-SMA、Snail1、Collagen I蛋白表达均低于空载体组(P<0.05),肺组织SOD水平和E-cadherin蛋白表达高于空载体组(P<0.05)。结论低表达NDNF可以改善小鼠肺纤维化损伤,其机制可能与减轻炎症及氧化应激反应、调节肺EMT相关蛋白表达有关。Objective To explore the impact of silencing neuron-derived neurotrophic factor(NDNF)on pulmonary inflammation,oxidative stress and epithelial-mesenchymal transition(EMT)in mice with bleomycin-induced pulmonary fibrosis.Methods A mouse model of pulmonary fibrosis was established by tracheal injection of bleomycin(5 mg/kg,fibrosis group),and NDNF knockdown pulmonary fibrosis group was established by tail vein injection of siRNA-NDNF adenovirus vector(siRNA-NDNF group),with concurrent establishment of empty vector group and healthy group.At 2 weeks after successful modeling,HE and Masson staining were used to observe the pathological changes in mouse lung tissues(alveolar inflammation score)and the degree of pulmonary fibrosis(fibrosis ratio).ELISA was used to detect the levels of inflammatory factors[tumor necrosis factor-alpha(TNF-α),transforming growth factor-beta1(TGF-β1),interleukin(IL)-4,and IL-6],oxidative stress factors[malondialdehyde(MDA),reactive oxygen species(ROS),and superoxide dismutase(SOD)]and hydroxyproline(HYP)in lung tissues.Western blot was performed to detect the expressions of EMT-related proteins[E-cadherin,α-smooth muscle actin(α-SMA),Snail1,and CollagenⅠ]in mouse lung tissues in each group.Results Pulmonary fibrosis group exhibited severe damage in alveolar structure,severe alveolar damage,remarkable inflammatory infiltration and extensive deposition of collagen fibers.Pulmonary coefficient,fibrosis ratio,alveolar inflammation score and HYP level in pulmonary tissues were all higher in pulmonary fibrosis group than those of healthy group(P<0.05).The damage in alveolar structure,inflammatory infiltration,and blue areas of collagen fibers in mouse lung tissues were reduced in siRNA-NDNF group when compared to empty vector group(P<0.05).The pulmonary coefficient,fibrosis ratio,alveolar inflammation score and HYP level in pulmonary tissues were lower in siRNA-NDNF group than those in empty vector group(P<0.05).The levels of TNF-α,TGF-β1,IL-6,IL-4,MDA,and ROS as well as the protein expr

关 键 词：肺纤维化 博来霉素 神经元源性神经营养因子 上皮-间质转分化 炎症反应 

分 类 号：R563.1[医药卫生—呼吸系统]