目的 通过对豚鼠整体行为表现、病理组织切片的观察,炎性细胞(嗜酸性粒细胞、中性粒细胞、淋巴细胞)、炎性细胞因子(IL-1β、TNF-α、IL-4、IL-8、IL-13)和抗炎细胞因子(IL-10)在豚鼠支气管肺泡灌洗液(BALF)中含量的变化,初步探讨寒喘舒和安喘舒治疗过敏性支气管哮喘模型豚鼠的作用机制.方法 将188只豚鼠按随机数字表法分为6组,应用鸡卵白蛋白雾化吸入法建立稳定的豚鼠过敏性支气管哮喘模型.正常对照组(予生理盐水模拟抗原攻击,A组,n=20);过敏性支气管哮喘模型组(予生理盐水模拟治疗,B组,n=40);寒喘舒组(C组)予低剂量(C1组,n=32)、高剂量 (C2组,n=32) 寒喘舒治疗;安喘舒组(D组)予低剂量(D1组,n=32)、高剂量(D2组,n=32)安喘舒治疗.肉眼观察各组动物饮食、活动、抓鼻、咳嗽、喷嚏等过敏行为变化;各组于治疗后1、2、8、24 h处死动物,观察肺色泽、大小等;用生理盐水行支气管肺泡灌洗,收集BALF,离心后取沉淀细胞作细胞涂片,计数细胞;用Lowrys法测量BALF上清液中总蛋白含量,用双抗夹心ELISA法测定上清液中TNF-α、IL-1β、IL-4、IL-8、IL-10、IL-13的含量,计算各时间点各组细胞因子含量与总蛋白、抗炎因子与致炎因子(IL-10/TNF-α、IL-10/IL-1β)的比值.结果 光镜下可见:A组肺泡管和肺泡壁结构完整;B组肺泡管和肺泡壁结构受损,肺泡腔内充满渗出液,可见坏死脱落的肺泡上皮细胞和渗出的白细胞,肺间质水肿,炎性细胞浸润;C1、C2组肺泡管和肺泡壁结构受损程度轻于B组,肺泡腔内见极少量炎症细胞游于其中,支气管管腔及肺泡周围少见炎性细胞浸润.D1、D2组组织结构基本正常,未见明显损害.BALF中细胞学变化:与B组相比,C1、C2、D1、D2组嗜酸性粒细胞、中性粒细胞、淋巴细胞数量显著下降(P<0.05),巨噬细胞显著上升(P<0.05).BALF中细胞因子含量变化:C1、C2、D1、D2组BALF炎性因子IL-1β、TNF-α、IL-4、IL-8、IL-13含量随时间增加有所下降或趋于正常,而抗炎因子IL-10随时间增加逐渐增加,C2和D2组在治疗后24 h仍维持高水平(P＜0.05).C2、D1、D2组IL-10/TNF-α、IL-10/IL-1β的比值较B组差异有统计学意义(P＜0.05).结论 提前应用安喘舒对过敏性支气管哮喘豚鼠有良好的保护效果,提示安喘舒对哮喘的发生有一定的预防作用.%Objective To explore the action mechanisms of Hanchuashu and Anchuanshu in a guinea pig model of allergic bronchial asthma through investigating overall behavior of animals , histopathological changes ,and contents of inflammatory cells (eosinophils ,neutrophils and lymphocytes) ,inflammatory cytokines (IL-1β ,TNF-α ,IL-4 ,IL-8, IL-13 ) and anti-inflammatory cyto-kines (IL-10) in broncho alveolar lavage fluid (BALF) .Methods A total of 188 guinea pigs were randomly divided into six groups :normal saline control group (group A ,n=20) ,allergic bronchial asthma model group (group B , n= 40 ),low-dose Hanchuanshu treatment group (group C1, n-32) ,high-dose Hanchuanshu treatment group(group C2 ,n= 32 ),low-dose Anchuanshu treatment group(group D1 ,n=32) and high-dose Anchuanshu treatment group (group D1 , n=32) .Allergic bronchial asthma was induced by chicken ovalbumin inhalation . Animal diet ,daily activity ,scratching nose,coughing ,sneezing and other allergic manifestations were observed .Animals were sacrificed 1 ,2 ,8 and 24 hours after treatment and pulmonary color and size were recorded .In addition ,bronchoalveolar lavage was performed with normal saline .BALF was collected for preparation of cell smears after centrifugation and cells were counted .Total protein content in BALF was measured by Lowry 's method .The contents of TNF-α ,IL-1β ,IL-4 ,IL-8, IL-10 and IL-13 were determined by double-antibody sandwich ELISA . The ratios of cytokines to total protein ,anti-inflammatory cytokines and inflammatory cytokines were calculated at each time point .Results Optical microscopy showed complete alveolar ducts and alveolar walls in group A ,D1 and D2 ,and destruction of alveolar walls and alveolar ducts in group B ,C1 and C2 .Furthermore ,alveolar space exudate,alveolar epithelial cell necrosis , leukocyte exudation ; pulmonary interstitial edema, inflammatory cell infiltration ,pulmonary interstitial edema and inflammatory cell infiltration were found in group B ,and a very small amount of inflammatory cells in the alveolar cavity and rare in -flammatory cell infiltration in the bronchial lumen and around the alveolus in group C 1 and C2 . Compared with group B ,the number of macrophages obviously increased and the numbers of eosinophils ,neutrophils and lymphocytes in BALF significantly decreased in group C1 ,C2 ,D1 and D2( P<0 .05 ) .The contents of IL-1β,TNF-α, IL-4, IL-8 and IL-13 decreased and IL-10 increased with time in group C1 ,C2 ,D1 and D2 ,and IL-10 contents maintained at a high level 24 hours after treatment in group C2 and D2 .Moreover,the ratio of IL-10 to TNF-α and ratio of IL-10 to IL-1β in group B were significantly different from those in group C2 ,D1 and D2 (P<0 .05) .Conclustion Anchuanshu has a preventive effect on asthma in guinea pigs .
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