摘要
目的 观察虾青素(astaxanthin,AST)对支气管哮喘大鼠气道炎症及气道重构的影响.方法 将50只雄性Wistar大鼠按随机数字表法分为5组(n=10):生理盐水致敏+生理盐水激发组(对照组)、支气管哮喘组(哮喘组)、支气管哮喘+虾青素5 mg/kg灌胃处理组(AST 5 mg/kg组)、10 mg/kg灌胃处理组(AST 10 mg/kg组)、50 mg/kg 灌胃处理组(AST 50 mg/kg 组).酶联免疫吸附(enzyme-linked immunosorbent assay,ELISA)法测定支气管肺泡灌洗液(bronchoalveolar lavage fluid,BALF)、白介素 5(interleukin-5,IL-5)、白介素 13(interleukin-13,IL-13)、干扰素-γ(interferon-γ,IFN-γ)、转化生长因子 β(tansforming growth factor-β,TGF-β)、丙二醛(malondialdehyde,MDA)、超氧化物歧化酶(superoxide dismutase,SOD)及血清总IgE水平;肺组织切片苏木素-伊红(hematoxylin-eosin,HE)染色观察气道炎症细胞浸润和上皮细胞脱落程度;过碘酸雪夫染色(periodic acid schiff,PAS)观察气道上皮杯状细胞增生程度;Masson染色观察气道上皮下胶原纤维沉积的程度;逆转录聚合酶链式反应(reverse transcription-polymerase chain reaction,RT-PCR)测肺组织中黏蛋白 5AC(mucin 5A and C,MUC5AC)信使核糖核酸(messenger ribonucleic acid,mRNA)表达;免疫组织化学染色测定大鼠气道上皮MUC5AC蛋白表达.结果 哮喘组大鼠BALF中IL-5、IL-13、TGF-β、MDA、血清总IgE分别为[(36.73±2.29)、(53.99±2.70)、(60.89±2.54)ng/mL、(18.65±0.76)umol/L、(54.50±2.91)ng/mL]、炎症细胞(46.24±4.26)、嗜酸性粒细胞(2.09±0.13)、脱落上皮/气道上皮[(6.09±0.45)％]、杯状细胞/上皮细胞面积比值[(13.65±1.90)％]、胶原纤维/基底膜下20pm区域面积[(17.58±2.14)％]、肺组织MUC5AC mRNA、肺组织MUC5AC蛋白表达IOD值(187±12)均高于对照组(P＜0.01 或 P＜0.001),IFN-y[(26.38±1.70)ng/mL]、SOD[(16.37±1.22)U/L]低于对照组(P＜0.001);AST处理组IL-5、IL-13、总IgE、TGF-β、MDA水平、气道上皮炎症细胞浸润和上皮细胞损伤脱落程度、气道上皮杯状细胞增生程度、气道上皮下有胶原纤维沉积程度、肺组织MUC5AC mRNA、气道上皮细胞MUC5AC蛋白表达均低于哮喘组(P＜0.05或P＜0.01或P＜0.001),IFN-γ、SOD高于哮喘组(P＜0.05或P＜0.01).结论 虾青素可抑制支气管哮喘大鼠气道炎症,下调气道MUC5AC蛋白表达,抑制杯状细胞增生,减轻气道重构.
Abstract
Objective To observe the effects of astaxanthin(AST)on the airway inflammation and remodeling in the asthmatic rats.Methods Fifty male Wistar rats were randomly divided into five groups(n=10 for each group):saline-sensitized and-saline-challenged group(the control group),bronchial asthma group(the asthma group),bronchial asthma+astaxanthin 5 mg/kg gavage treatment group(the AST 5 mg/kg group),bronchial asthma+10 mg/kg gavage treatment group(the AST 10 mg/kg group),and bronchial asthma+50 mg/kg gavage treatment group(the AST 50 mg/kg group).The level of interleukin-5(IL-5),interleukin-13(IL-13),interferon-γ(IFN-γ),tansforming growth factor-β(TGF-β),malondialdehyde(MDA)and superoxide dismutase(SOD)in the bronchoalveolar lavage fluid(BALF)and the total IgE level in the serum were measured using enzyme linked immunosorbent assay(ELISA).The infiltration of airway inflammatory cells and the degree of airway epithelial cells detachment,the extent of goblet cell hyperplasia and the severity of subepithelial collagen deposition were evaluated on the hematoxylin eosin(HE),periodic acid Schiff(PAS)and Masson trichrome stained lung sections.reverse transcription-polymerase chain reaction(RT-PCR)was used to measure the expression of mucin 5A and C(MUC5AC)messenger ribonucleic acid(mRNA)in lung tissue;Immunohistochemical staining was used to determine the expression of MUC5AC protein in the rat airway epithelium.Results The level of IL-5,IL-13,TGF-β,MDA and the total IgE in the serum respectively[(36.73±2.29),(53.99±2.70),(60.89±2.54)ng/mL,(18.65±0.76)umol/L,(54.50±2.91)ng/mL],the extent of inflammatory cells infiltration(46.24±4.26),the extent of eosinophils infiltration(2.09±0.13),the extent of epithelial cells detachment[(6.09±0.45)％],the extent of goblet cell hyperplasia[(13.65±1.90)％],the extent of subepithelial collagen deposition[(17.58±2.14)％],the MUC5AC mRNA expression level,and the lung tissue MUC5AC protein expression IOD value(187±12)in the asthma group were all higher than those in the control group(P＜0.01 or P＜0.001),the level of IFN-γ and SOD in the BALF[(26.38±1.70)ng/mL],[(16.37±1.22)U/L],was lower than that in the control group(P＜0.001);The level of IL-5,IL-13,total IgE,TGF-β,MDA,the inflammatory cells infiltration in the airway epithelial,the degree of epithelial cell damage and detachment,the degree of goblet cell hyperplasia,the degree of subepithelial collagen deposition,the MUC5AC mRNA expression in lung tissue,and the MUC5AC protein expression in airway epithelial cells in the AST treated groups were all lower than those in the asthma group(P＜0.05 or P＜0.01 or P＜0.001),the level of IFN-y,SOD in the BALF was higher than that in the asthma group(P＜0.05 or P＜0.01).Conclusion Astaxanthin can inhibit airway inflammation,downregulate airway MUC5AC expression,inhibit goblet cell proliferation,and alleviate airway remodeling in rats with bronchial asthma.
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