不同浓度鸦胆子苦醇对小鼠矽肺纤维化的影响  

Effects of different concentrations of brusatol on silicosis fibrosis in mice

作　　者：康慧敏 李柔 王汉钦 郑云帆 陈适 KANG Huimin;LI Rou;WANG Hanqin;ZHENG Yunfan;CHEN Shi(School of Medicine,Hunan Normal University,Changsha,Hunan 410000,China)

机构地区：[1]湖南师范大学医学院,湖南长沙410000

出　　处：《环境与职业医学》2024年第5期539-545,共7页Journal of Environmental and Occupational Medicine

基　　金：国家自然科学基金项目(82173493);湖南省自然科学基金项目(2023JJ30423)。

摘　　要：[背景]矽肺是一种肺部弥漫性纤维化疾病,由长期暴露于游离二氧化硅(SiO_(2))粉尘引起,发病机制复杂,缺乏有效的治疗。鸦胆子苦醇(Bru)具有多种生物活性,其在矽肺纤维化中的作用尚不明确。[目的]探究不同浓度Bru对SiO_(2)诱导小鼠矽肺纤维化的影响。[方法]将30只雄性C57BL/6J小鼠随机分为对照组、矽尘组、Bru低剂量(1 mg·kg^(-1))组、Bru中剂量(2 mg·kg^(-1))组、Bru高剂量(4 mg·kg^(-1))组,每组6只;除对照组外,其余各组均采用一次性非气管暴露法滴注50μL、60 mg·mL^(-1)SiO_(2)悬浊液建立矽肺小鼠模型,对照组滴注等量生理盐水;Bru组于染尘的同时腹腔注射Bru,连续注射5d,随后隔天注射,染尘28 d后处死小鼠,收集肺组织。测定小鼠肺系数;采用苏木精-伊红(HE)染色和Masson染色观察小鼠肺组织的病理变化;Western blot法检测小鼠肺组织中凋亡蛋白Cleaved-caspase 3、纤维化相关蛋白α-平滑肌肌动蛋白(α-SMA)、Ⅰ型胶原蛋白(Col-I)、自噬相关蛋白Beclin1、微管相关蛋白1轻链3(LC3)、Sequestosome 1(p62/SQSTM1)蛋白、Kelch样环氧氯丙烷相关蛋白1(Keap1),核因子E2相关因子2(Nrf2)的表达水平;实时荧光定量逆转录PCR(RT-qPCR)检测小鼠肺组织中Caspase 3、α-SMA和Col-lmRNA水平。[结果]与对照组相比,矽尘组小鼠的肺系数明显升高(P<0.01);肺组织中肺泡壁受损,出现炎性细胞浸润、纤维结节和胶原纤维沉积;Cleaved-caspase 3、α-SMA和Col-I的蛋白表达及转录水平均明显上调(P<0.01);Beclin1、LC3-Ⅱ/Ⅰ、p62、Nrf2表达水平增加(P<0.05,P<0.01),而Keap1表达水平下降(P<0.05)。与矽尘组相比,Bru低、中剂量组小鼠肺系数降低(P<0.05);肺组织的病理损伤及胶原沉积明显改善;Cleaved-caspase 3、α-SMA和Col-I的蛋白表达及转录水平下降(P<0.05,P<0.01),Beclin1、LC3-Ⅱ/Ⅰ、p62、Nrf2表达水平也均明显下降(P<0.05,P<0.01),中剂量组Keap1水平上升(P<0.05)。与矽尘组相比,Bru[Background]Silicosis is a diffuse fibrosis of the lungs caused by long-term inhalation of free silicon dioxide(SiO_(2)).It has a complex pathogenesis and lacks effective treatment.Brusatol(Bru)has a variety of biological activities,and its role in silicosis fibrosis is unclear yet.[Objective]To investigate the effects of different concentrations of Bru on SiO_(2)-induced silicosis fibrosis in mice.[Methods]Thirty male C57BL/6J mice were randomly divided into five groups:a control group,a silica group,and three Bru intervention groups with low,medium,and high doses(1,2,and 4 mg·kg^(-1)),with 6 mice in each group.Except the control group,the remaining groups were establis-hed as SiO_(2)-induced silicosis mouse models by using a single tracheal infusion of 50μL 60 mg·mL^(-1) SiO_(2) suspension.The control group was dosed with equal amount of saline.The Bru intervention groups were injected intraperitoneally with Bru for 5 consecutive days and then injected every other day.After 28 d of exposure,the mice were executed and lung tissues were collected.The lung coefficient of the mice was measured,and the pathological changes of the lung tissues were observed after hematoxylin-eosin(HE)and Masson staining.The levels of apoptotic protein Cleaved-caspase 3,fibrosis-related proteinα-smooth muscle actin(α-SMA),type I collagen(Col-I),autophagy-associated protein Beclin1,microtubule-associated protein 1 light chain 3(LC3),Sequestosome 1(p62/SQSTM1),Kelch like ECH-associated protein-1(Keap1),and nuclear factor erythroid 2 related factor 2(Nrf2)were detected by Western blot.The mRNA levels of Caspase 3,α-SMA,and Col-I were measured by realtime fluo-rescence-based quantitative PCR.[Results]Compared with the control group,the lung coefficient of mice in the silica group was significantly increased(P<0.01);the lung tissues of the silicosis mice showed damaged alveolar walls,along with infiltration of inflammatory cells,fibrous nodules,and collagen deposition;furthermore,the protein and mRNA levels of Cleaved-caspase 3,α-SM

关 键 词：矽肺 鸦胆子苦醇 肺纤维化 自噬 凋亡 

分 类 号：R114[医药卫生—卫生毒理学]