日本血吸虫感染对过敏性哮喘影响的实验研究

目的 建立日本血吸虫感染并发过敏性哮喘实验动物模型及探讨日本血吸虫感染对过敏性哮喘的作用. 方法 取25只BALB/c小鼠,随机分为3组,A组为日本血吸虫感染并发过敏性哮喘组,B组为单纯过敏性哮喘组,C组为健康对照组.A组小鼠经腹部皮肤感染日本血吸虫尾蚴(23±3)条.8周后,以卵白蛋白(OVA)分别对A组、B组小鼠进行致敏激发,建立过敏性哮喘模型.12周后,取小鼠脾脏制成单细胞悬液并培养72 h,收集脾细胞上清液,检测细胞因子IL-4和IFN-γ水平;收集支气管肺泡灌洗液(BALF),观察嗜酸性粒细胞(EOS)所占的百分比;取小鼠肺组织作病理切片,观察小鼠肺组织的炎症变化. 结果 与B组比较,A组小鼠BALF涂片中EOS数减少(P＜0.05),A、B两组EOS百分比分别为(7.30±5.01)%和(43.60±2.53)%,而C组小鼠BALF中仅见少数气管上皮细胞,无炎细胞;ELISA法检测A、B、C组小鼠脾细胞培养上清IL-4水平分别为(96.02±38.54) pg/ml、(414.86±175.12) pg/ml和(13.55±1.66) pg/ml,IFN-γ水平分别为(11.60±4.18) pg/ml、(7.43±3.60)pg/ml和(7.55±1.57) pg/ml.与B组比较,A组小鼠IL-4水平降低(P＜0.05),IFN-γ水平无显著性变化(P＞0.05),IL-4/IFN-γ比值降低(P＜0.05).肺部病理改变A组小鼠较轻,B组较重,C组无炎症反应. 结论 日本血吸虫感染对过敏性哮喘的发生有抑制作用.     

日本血吸虫感染鼠树突状细胞对哮喘抑制作用的实验研究

目的通过过继转移日本血吸虫感染鼠树突状细胞(DC),探讨DC在抑制过敏性哮喘中的作用及机制。方法用CD11c 磁珠分离纯化日本血吸虫感染鼠及正常鼠DC亚群CD11c 细胞。将15只BALB/c小鼠随机分为3组,A组为过继转移日本血吸虫感染鼠DC并诱发过敏性哮喘组,B组为过继转移正常鼠DC并诱发过敏性哮喘组,C组为单纯过敏性哮喘组。A、B两组小鼠经尾静脉过继转移1×106DC,2 h后A、B、C 3组均开始诱发哮喘。4周后处死小鼠,收集肺泡灌洗液,检测细胞因子IL-4I、L-5及IFN-γ水平;取小鼠肺组织作病理切片,观察其炎症变化。结果与B、C组比较,A组肺部炎症明显减轻,按Underwood标准,A、B、C 3组总评分分别为5.00±1.58、11.40±2.07和13.20±2.86,差异有统计学意义(P<0.05)。A、B、C组小鼠肺泡灌洗液IL-4水平分别为(23.25±1.57)pg/ml、(40.70±3.28)pg/ml和(65.23±4.84)pg/ml,差异有统计学意义(P<0.05);IL-5水平分别为(16.20±4.11)pg/ml、(41.41±7.14)pg/ml和(64.75±16.96)pg/ml,IFN-γ水平分别为(6.60±2.34)pg/ml(、6.94±2.19)pg/ml和(5.87±2.44)pg/ml,差异无统计学意义(P>0.05)。结论日本血吸虫感染鼠DC通过影响受体鼠Th2细胞因子的分泌明显抑制过敏性哮喘的发生。     

加味苏子降气汤对哮喘模型大鼠细胞因子功能紊乱的调节

目的观察加味苏子降气汤对支气管哮喘模型大鼠细胞因子功能紊乱的影响。方法清洁级雄性SD大鼠40只,按随机数字表法分为正常对照组、哮喘模型组、加味苏子降气汤组和地塞米松组。采用卵清白蛋白致敏法建立哮喘大鼠模型。酶联免疫吸附试验(ELISA)测定肺泡灌洗液中白细胞介素(IL)-4、干扰素(IFN)-γ的水平。结果哮喘模型组肺泡灌洗液中IL-4水平上升(P<0. 01);加味苏子降气组及地塞米松组肺泡灌洗液中IFN-γ水平上升(P<0. 05),IL-4下降(P<0. 05)。结论加味苏子降气汤可以改善哮喘患者体内细胞因子水平,调节细胞因子失衡从而达到治疗目的。     

小鼠哮喘模型中补体C5a对IL-17表达的调节作用

目的 探讨小鼠哮喘模型中补体C5a对IL-17表达的调节作用.方法 用卵清蛋白(OVA)致敏、激发的方法 建立Balb/C小鼠哮喘模型.用体内抗体中和试验,观察致敏前中和炎症介质补体C5a的功能是否影响小鼠肺泡灌洗液中IL-17细胞因子的表达水平,并探讨其机制.结果 病理结果 显示模型组小鼠的肺组织出现肺泡腔扩张、腔内大量炎性细胞的浸润.小鼠致敏前给予抗C5a抗体干预上调了肺部IL-17及IL-6 的表达,导致肺炎症明显加重.结论 小鼠哮喘模型中体内C5a分子对IL-17表达具有负调节作用.     

细胞因子在小鼠支气管哮喘发病中的作用及梓醇的治疗效应

目的 研究细胞因子IL-4、IL-5、IL-8、IFN-r及 NO与哮喘发生的关系,观察中药梓醇对支气管哮喘的治疗作用.方法　将30只Balb/c小鼠随机分为三组:生理盐水对照组、哮喘模型组、梓醇治疗组,每组各10只.除对照组外,余两组采用卵蛋白和氢氧化铝混合溶液注射致敏,并雾化吸入建立哮喘模型.治疗组小鼠从第15天起每次雾化攻击前1 h给予梓醇5 mg/kg腹腔注射,连续7 d.其余两组用生理盐水替代卵蛋白进行注射.第22天处死小鼠,收取血清,并取其肺、脾组织,制备支气管肺组织匀浆,收集支气管肺泡灌洗液(BALF),脾淋巴细胞和巨噬细胞培养上清液.用放射免疫法检测IL-8的水平,酶联免疫法(ELISA)检测IFN-γ、IL-4、IL-5的水平,用酶法检测 NO含量,计数小鼠BALF中白细胞的数量.结果 哮喘模型组肺泡灌洗液中白细胞计数、支气管肺组织中IL-4、脾淋巴细胞培养上清液中IL-5、血清与脾淋巴细胞培养上清液中IL-8含量及脾巨噬细胞培养上清液中NO含量明显高于对照组(P<0.01);而治疗组上述指标明显低于哮喘模型组(P<0.01或P<0.05);哮喘模型组脾淋巴细胞产生INF-γ含量明显低于对照组,治疗组明显高于哮喘模型组(P<0.01).结论 细胞因子IL-4、IL-5、IL-8、IFN-γ和NO水平的变化反映了Th1和Th2的失衡和巨噬细胞功能的增强,可能参与了哮喘发病的病理生理过程,中药梓醇对支气管哮喘有一定的治疗作用.     

粉螨性小鼠哮喘模型的建立与评估

目的探讨采用椭圆食粉螨提取浸液建立C57BL/6小鼠哮喘模型的方法。方法 45只C57BL/6小鼠随机分成3组:PBS阴性对照组、卵清蛋白(OVA)阳性对照组、椭圆食粉螨(Ale o)哮喘模型组。用酶联免疫吸附试验(ELISA)检测支气管肺泡灌洗液(BALF)、脾细胞培养上清液中IL-4、IL-17和IFN-γ的含量,计数BALF中细胞总数并分类,同时切取小鼠未灌洗侧肺组织进行病理学观察。结果 OVA阳性对照组、Ale o模型组小鼠脾细胞上清和BALF中IL-4、IL-17及IFN-γ含量与PBS阴性对照组相比,差异均具有统计学意义(P<0.05或P<0.01);OVA阳性对照组、Ale o模型组小鼠BALF中细胞总数及各类细胞数与PBS阴性对照组相比,差异有统计学意义(P<0.05或P<0.01);小鼠未灌洗侧肺组织病理学观察显示,OVA阳性对照组和Ale o模型组小鼠肺组织可见大量炎症细胞浸润,以嗜酸性粒细胞、淋巴细胞及巨噬细胞为主。结论用椭圆食粉螨提取浸液建立的C57BL/6小鼠哮喘模型具有过敏性哮喘的主要特征。     

重楼总皂甙对小鼠哮喘模型气道炎症的影响及机制

目的探讨重楼总皂甙对小鼠哮喘模型的保护作用及可能机制。方法将24只BALB/c小鼠随机分为空白对照组,模型组,重楼总皂甙低剂量组(LRTPS)和重楼总皂甙高剂量组(HRTPS),每组6只。除空白对照组小鼠外,其余各组采用卵蛋白致敏建立支气管哮喘小鼠模型,治疗组在造模的同时灌胃给予不同剂量的重楼总皂甙治疗。通过肺组织病理切片、酶联免疫反应(ELISA)法检测肺泡灌洗液中的白细胞介素(IL)-4和干扰素(IFN)-γ水平,Q-PCR检测肺组织IL-4和IFN-γmRNA表达水平。结果空白对照组小鼠肺组织支气管无炎性细胞浸润,气道无明显黏液分泌,模型组支气管和血管周围炎性细胞浸润明显,黏膜及黏膜下层水肿明显。治疗组支气管和血管周围炎性细胞浸润明显减少,肺泡结构清晰,肺泡腔和支气管腔无渗出物。哮喘组小鼠的肺泡灌洗液IL-4水平和肺组织IL-4 mRNA表达显著高于对照组;治疗组显著低于哮喘组(P<0.05)。哮喘组小鼠的肺泡灌洗液IFN-γ水平和肺组织IFN-γmRNA表达显著低于对照组,治疗组显著高于哮喘组(P<0.05)。结论重楼总皂甙能明显减轻哮喘气道炎性反应,其机制可能与恢复局部Th1/Th2细胞因子平衡有关。     

不同内型哮喘小鼠模型中的小气道功能研究及其机制初探

目的 探讨不同内型哮喘小鼠模型中,小气道功能是否存在异常及其相关机制.方法 卵清蛋白(OVA)致敏、激发建立T2型哮喘模型,OVA联合臭氧暴露(OVA+ozone)建立非T2型哮喘模型.模拟强迫振荡系统检测小鼠小气道功能,激发试验检测气道反应性.酶联免疫吸附试验法检测支气管肺泡灌洗液(BALF)中的细胞因子;苏木精-伊...     

卡介苗多糖核酸通过抑制支气管哮喘小鼠核因子κB影响支气管肺泡灌洗液中细胞因子水平的研究

目的 观察卡介苗多糖核酸(BCG-PSN)对支气管哮喘(简称哮喘)小鼠Th1/Th2型免疫反应的调节作用,探讨核因子κB(NF-κB)与细胞因子干扰素γ(IFN-γ)、白介素4(IL-4)、IL-5的关系及BCG-PSN的调节作用.方法 35只小鼠随机分为对照组、卵白蛋白(OVA)组、BCG-PSN 6周龄组、BCG-PSN 6周龄+OVA组、BCG-PSN 1周龄+OVA组.所有动物于第40天和第41天雾化吸入OVA进行气道激发,第42天行支气管肺泡灌洗;取支气管、肺组织标本.用ELISA方法测定支气管肺泡灌洗液(BALF)中INF-γ、IL-4、IL-5的水平,用免疫组织化学染色方法检测支气管、肺组织NF-κB的表达.结果 OVA致敏后BALF中IL-4、IL-5的水平及支气管、肺组织中NF-κB的表达明显增加,且NF-κB的表达与IL-4、IL-5的水平呈正相关;BCG-PSN免疫后IL-4、IL-5的水平及NF-κB的表达下降,BCG-PSN 1周龄+OVA组IFN-γ的水平有显著升高.结论 NF-κB对IL-4、IL-5具有上调作用,BCG-PSN免疫可提高BCG-PSN 1周龄鼠IFN-γ水平.     

卡介苗多糖核酸通过抑制支气管哮喘小鼠核因子κB影响支气管肺泡灌洗液中细胞因子水平的研究

目的 观察卡介苗多糖核酸(BCG-PSN)对支气管哮喘(简称哮喘)小鼠Thl/Th2型免疫反应的调节作用,探讨核因子kB(NF-κB)与细胞因子干扰素7(IFN-γ)、白介素4(IL-4)、IL-5的关系及BCG-PSN的调节作用.方法 35只小鼠随机分为对照组、卵白蛋白(OVA)组、BCG-PSN 6周龄组、BCG-PSN 6周龄+OvA组、BCG-PSN 1周龄+OVA组.所有动物于第40天和第41天雾化吸入OVA进行气道激发,第42天行支气管肺泡灌洗;取支气管、肺组织标本.用ELISA 方法测定支气管肺泡灌洗液(BALF)中INF-γ、IL-4、IL-5的水平,用免疫组织化学染色方法检测支气管、肺组织NF-κB的表达.结果 OVA致敏后BALF中IL-4、IL-5的水平及支气管、肺组织中NF-κB的表达明显增加,且NF-κB的表达与IL_4、IL-5的水平呈正相关;BCG-PSN免疫后IL-4、IL-5的水平及NF-κB的表达下降,BCG-PSN 1周龄+OVA组IFN-γ的水平有显著升高.结论 NF-γB对IL-4、IL-5具有上调作用,BCG-PSN免疫可提高BCGPSN 1周龄鼠IFN-γ水平.     

粉尘螨Ⅲ类重组变应原致敏的小鼠哮喘模型致敏效果分析

目的 探讨粉尘螨Ⅲ类重组变应原（Der f3）诱导的小鼠哮喘模型的致敏效果。方法 30只6~8周龄雌性SPF级BALB/c小鼠随机分为PBS组（阴性对照组）、卵清蛋白（OVA）组（阳性对照组）和Der f3组（实验组）,OVA组和Der f3组分别使用OVA和纯化Der f3蛋白于第0、7、14天进行腹腔注射致敏,于第21d开始雾化吸入,连续7d,PBS组则换成PBS进行腹腔注射和雾化吸入。最后一次雾化吸入24h后,分别观察肺组织病理变化、支气管肺泡灌洗液（BALF）中的白细胞计数,酶联免疫吸附试验（ELISA）测定BALF和脾细胞培养上清中IL-4、IL-2、IL-17和IFN-γ的含量及血清中IgG1、IgE抗体水平变化;结果OVA组和Der f3组小鼠肺部支气管、血管粘膜下及周围肺组织有明显的炎性细胞浸润、支气管上皮细胞部分断裂及脱落,血管壁明显水肿等,而PBS组未见明显病理改变;Der f3组的BALF中白细胞总数[(19.99±1.03)×106个/ml]及嗜酸性粒细胞（EOS）[(1.81±0.07)×106个/ml]计数均明显高于PBS组（P<0.01）;与PBS组相比：BALF中IL-4 [(80.99±9.06) pg/ml]、IL-17 [(209.69±31.86) pg/ml]呈高水平表达（P<0.01）,而IL-2 [(8.29±1.27) pg/ml]和IFN-γ [(55.04±18.85) pg/ml]含量则显著下降（P<0.01）;上述指标在脾细胞培养上清中出现类似的变化。Der f3组血清中IgE [(31.92±2.68) U/ml]、IgG1 [(16.46±4.32) μg/ml]的含量表现为Th2型反应增强趋势;但OVA组和Der f3组间所有指标差异均无统计学意义（P>0.05）;结论 粉尘螨重组Der f3蛋白可成功建立小鼠变态反应性气道及肺部炎症动物模型。     

外源性间充质干细胞减轻支气管哮喘小鼠气道炎症的研究

目的 观察鸡卵清蛋白诱导小鼠支气管哮喘(简称哮喘)模型中外源性间充质干细胞(mesenchymal stem cells,MSC)在哮喘小鼠肺组织气道炎症中的作用.方法 45只雌性SPF级C57BL/6小鼠,体质量18～22 g.随机分为对照组(P:P:P)、哮喘组(O:P:O)和MSC治疗组(O:M:O).哮喘组与MSC治疗组第1天和第8天致敏,第15天、第16天和第17天使用OVA气道内滴入激发哮喘.MSC治疗组于哮喘造模第14天移植外源性MSC.对照组小鼠予PBS处理.三组小鼠于末次激发结束后24 h(第18天)处死,取支气管肺泡灌洗液上清,ELISA检测IL-5、IL-9及β-氨基己糖苷酶;支气管肺泡灌洗液细胞计数总细胞数、嗜酸粒细胞数;取肺组织行病理切片苏木精-伊红染色观察肺部气道炎症情况.结果 ①MSC下调了哮喘小鼠气道局部炎症;②MSC减轻了哮喘小鼠肺组织中的炎细胞浸润;③MSC减轻了哮喘小鼠气道中的肥大细胞脱颗粒现象;④MSC抑制了哮喘小鼠过度的Th2变态反应.结论 外源性MSC通过抑制Th2变态反应,减轻哮喘肺组织的气道炎症.     

外源性间充质干细胞减轻支气管哮喘小鼠气道炎症的研究

目的观察鸡卵清蛋白诱导小鼠支气管哮喘（简称哮喘）模型中外源性间充质干细胞（mesenehymal stem cells,MSC）在哮喘小鼠肺组织气道炎症中的作用。方法45只雌性SPF级C57BL／6小鼠,体质量18-22g。随机分为对照组（P：P：P）、哮喘组（O：P：O）和MSC治疗组（O：M：O）。哮喘组与MSC治疗组第1天和第8天致敏,第15天、第16天和第17天使用OVA气道内滴入激发哮喘。MSC治疗组于哮喘造模第14天移植外源性MSC。对照组小鼠予PBS处理。三组小鼠于末次激发结束后24h（第18天）处死,取支气管肺泡灌洗液上清,ELISA检测IL-5、IL-9及β-氨基己糖苷酶;支气管肺泡灌洗液细胞计数总细胞数、嗜酸粒细胞数;取肺组织行病理切片苏木精-伊红染色观察肺部气道炎症情况。结果①MSC下调了哮喘小鼠气道局部炎症;②MSC减轻了哮喘小鼠肺组织中的炎细胞浸润;③MSC减轻了哮喘小鼠气道中的肥大细胞脱颗粒现象;④MSC抑制了哮喘小鼠过度的Th2变态反应。结论外源性MSC通过抑制Th2变态反应,减轻哮喘肺组织的气道炎症。     

High mobility group box 1: a novel mediator of Th2-type response-induced airway inflammation of acute allergic asthma

Background

High mobility group box 1 (HMGB1) is an inflammatory mediator involved into the advanced stage of systemic inflammatory response syndrome (SIRS), and is over-expressed in bacterial sepsis and hemorrhagic shock. Recently, it has been found that the HMGB1 was abnormally expressed in induced sputum and plasma of asthmatic patients. However, the precise role of HMGB1 in the acute allergic asthma is unclear. Therefore, we aim to investigate the role HMGB1 in regulating airway inflammation of acute allergic asthma and its possible mechanism in this study.

Methods

Forty-eight BALB/c female mice were randomly divided into four groups: control group (Control), asthma group (Asthma), HMGB1 group (HMGB1) and anti-HMGB1 (HMGB1 monoclonal antibody of mice) group (Anti-HMGB1). Acute allergic asthma mice models were established by ovalbumin (OVA)-challenge. Then, we measured the levels of HMGB1 in bronchoalveolar lavage fluid (BALF) and lung tissue of mice. Finally, after exogenous HMGB1 and/or anti-HMGB1 administration, pulmonary function test, histological analysis, Western blot, cytological analysis and ELISA assay were performed to explore the effect of HMGB1 in acute allergic asthma.

Results

The levels of HMGB1 in BALF and lung tissue and the expression of HMGB1 protein in the lung tissue of asthma group were significantly higher than those in control group, respectively (P<0.01). Moreover, the HMGB1 group was showed an increased mucus secretion and infiltration of eosinophils and neutrophils in the airway of asthma mice, and a decrease of pulmonary function, compared to control group (P<0.01, respectively). Meanwhile, exogenous HMGB1 could increase the levels of IL-4, IL-5, IL-6, IL-8 and IL-17, whereas could reduce the IFN-γ in the BALF and lung tissue (P<0.05, respectively). Exogenous HMGB1 could enhance GATA3 expression of Th2 cells and attenuate the T-bet expression of Th1 cells (P<0.05, respectively), which could be abrogated after inhibiting HMGB1.

Conclusions

HMGB1 could aggravate eosinophilic inflammation in the airway of acute allergic asthma through inducing a dominance of Th2-type response and promoting the neutrophilic inflammation.     

Transfer of T cells from intranasal ovalbumin-immunized mice ameliorates allergic response in ova-sensitized recipient mice.

Mucosal immunotherapy is suggested as a treatment strategy for tolerance induction in allergic diseases. The purpose of this study was to determine the effect of transferred splenic T cells from intranasal ovalbumin (OVA)-immunized mice to naive mice before sensitization on its impact of cytokine production and airway histopathology. BALB/c mice in group I received intranasal immunotherapy (days1-6), carboxylfluorescein succinyl ester (CFSE)-labeled splenocytes or splenic T cells were i.v. transferred to naive recipients (group II) before OVA sensitization. Acute murine asthma model was established by two i.p. OVA injections (days 21 and 28) and seven OVA nebulizations (days 42-48) in groups I, II and III. Groups III and IV served as asthma model and control, respectively. CFSE-labeled cells in splenocytes and lymph node lymphocytes, lung histopathology, IL-4, IL-10, and interferon (IFN) gamma cytokines of recipients were analyzed 24 hours after OVA nebulization challenge. CFSE-labeled T cells from group I were detected in spleen and regional lymph nodes of the OVA-sensitized recipients (group II). Smooth muscle and thickness of airways were less in intranasal OVA immunotherapy and OVA-sensitized recipients when compared with the asthma model (p < 0.05). Area of inflammation was significantly suppressed in OVA-sensitized recipients compared with the asthma model (p < 0.01). IL-10 and IFN-gamma levels in splenocyte supernatants were significantly increased in intranasal immunotherapy and OVA-sensitized recipients compared with asthma model and controls (p < 0.01). IL-4 levels were significantly less in intranasal immunotherapy group and the OVA-sensitized recipient group when compared with asthma the model group (p < 0.05). This study suggests that intranasal immunotherapy with allergens regulates T-cell responses and ameliorates airway histopathology in sensitized mice, hence, encouraging mucosal tolerance induction as a suitable treatment of allergic diseases.     

Protective effect of Bifidobacterium infantis CGMCC313-2 on ovalbumin-induced airway asthma and b-lactoglobulininduced intestinal food allergy mouse models

AIM To determine whether oral administration of Bifidobacterium infantis CGMCC313-2(B. infantis CGMCC313-2) inhibits allergen-induced airway inflammation and food allergies in a mouse model.METHODS Ovalbumin(OVA)-induced allergic asthma and b-lactoglobulin-induced food allergy mouse models were used in this study. Following oral administration of B. infantis CGMCC313-2 during or after allergen sensitization, histopathologic changes in the lung and intestine were evaluated by hematoxylin and eosin(HE) staining. In the allergic asthma mouse model, we evaluated the proportion of lung-infiltrating inflammatory cells. OVAspecific IgE and IgG1 levels in serum and cytokine levels in bronchoalveolar lavage fluid(BALF) were also assessed. In the food allergy mouse model, the levels of total Ig E and cytokines in serum were measured.RESULTS Oral administration of B. infantis CGMCC313-2 during or after allergen sensitization suppressed allergic inflammation in lung and intestinal tissues, while the proportion of infiltrating inflammatory cells was significantly decreased in the BALF of allergic asthma mice. Moreover, B. infantis CGMCC313-2 decreased the serum levels of total Ig E in food allergy mice, and reductions in IgE and IgG1 were also observed in OVA-induced allergic asthma mice. The expression of interleukin-4(IL-4) and IL-13 in both serum and BALF was suppressed following the administration of B. infantis CGMCC313-2, while an effect on serum IL-10 levels was not observed.CONCLUSION B. infantis CGMCC313-2 inhibits the secretion of allergen-induced IgE, IL-4 and IL-13, and attenuates allergic inflammation.     

反义内皮素转换酶核酸纳米粒对变应性气道炎症的作用

目的 观察气管给予12-烷基化壳聚糖纳米粒(12-ACSs)包裹的反义内皮素转换酶(ECE)核酸表达质粒对小鼠变应性气道炎症的影响.方法 将40只健康雌性Balb/c小鼠按完全随机设计原则分为4组,每组10只,分为健康对照(N)组、OVA致敏激发并给予包裹反义ECE核酸的12-ACSs(NM)组、OVA致敏激发气道炎症模型(As)组及OVA致敏激发并给予裸反义ECE核酸(DNA)组.NM组、As组、DNA组在致敏后于首次激发前24 h分别通过气管灌注包裹反义ECE核酸的12-ACSs 100μl生理盐水100μl和反义ECE核酸5μg加生理盐水(总量100μl),N组用等体积生理盐水处理.末次激发后24 h收集BALF,计算BALF中细胞总数、分类计数.HE染色观察各组小鼠肺组织的病理变化.ELISA法检测肺组织匀浆、脾细胞培养卜清液白细胞介素(IL)-4、5、10、13、干扰素-γ(IFN-γ)和内皮素-1(ET-1)水平.流式细胞仪分别检测分泌IL-4、IFN-γ和IL-10脾细胞比例.对各组数据进行正态检验和单因素方差齐性检验,采用LSD法进行两两比较.结果 NM组小鼠BALF中细胞总数为(5.4±0.6)×105/ml、嗜酸性粒细胞为(1.8±0.4)×105/ml,均明显低于As组[分别为(8.5±0.9)×105/ml、(3.8±0.6)×105/ml]和DNA组[分别为(8.3±1.1)×105/ml、(3.8±0.5)×105/m1],均P<0.01;NM组小鼠肺组织的病理损害明显轻于As组和DNA组.肺匀浆N组ET-1水平为(11.7±1.7)ng/L,明显低于NM组的(18.5±1.4)ng/L、As组的(28.9±2.3)ng/L和DNA组的(27.1±1.6)ng/L,均P<0.01,而NM组ET-1水平则明显低于As组和DNA组(均P<0.01),As组和DNA组差异无统计学意义.N组IL-5水平均明显低于NM组、As组和DNA组(均P<0.01),而NM组IL-5水平则明显低于As组和DNA组(均P<0.05),Ag组和DNA组差异无统计学意义.IL-4、IL-13结果与IL-5、ET-1相似.各组IFN-γ的水平差异无统计学意义.NM组IL-10水平高于As组和DNA组(均P<0.01),与N组比较差异无统计学意义,脾细胞培养上清液细胞因子检测结果与肺匀浆结果相似.4组中CD4、IL-4表达均为阳性的细胞表达率分别为N组3.1%、NM组3.3%、As组5.9%、DNA组6.6%;表达IFN-γ、CD4阳性的细胞表达率分别为N组1.2%、NM组1.6%、As组1.7%、DNA组1.6%;表达IL-10、CD4、CD25阳性的细胞分别为N组1.9%、NM组4.3%、As组3.4%、DNA组3.1%.结论 气管内给予包裹反义ECE核酸表达质粒的12-ACSs纳米粒,可减少ET-1、IL-4、IL-5、IL-13的合成,增加IL-10的分泌,减轻变应性气道炎症.     

Timing of infection and prior immunization with respiratory syncytial virus (RSV) in RSV-enhanced allergic inflammation

Respiratory syncytial virus (RSV) infection has been shown to be a risk factor for the development of allergy in humans and mice. The allergy-enhancing properties of RSV may be dependent on atopic background and an individual's history of RSV infection. We examined the influence of the timing of infection and prior inoculation with RSV in a mouse model of allergic asthma. Mice were sensitized to and challenged with ovalbumin (OVA) and were inoculated with RSV either before or during the sensitization or challenge period. One group of mice was inoculated with RSV both before sensitization to OVA and during challenge with OVA. Increased pulmonary expression of interleukin (IL)-4, IL-5, and IL-13 mRNA and aggravated alveolitis and hypertrophy of mucus-producing cells were observed only when OVA-sensitized mice were inoculated with RSV shortly before or during challenge with OVA. Despite protection against viral replication, prior inoculation with RSV did not abrogate RSV-enhanced, OVA-induced expression of T helper 2 (Th2) cytokines in the lung. In conclusion, inoculation with RSV enhances allergic disease only when the immune system has already been Th2-primed by the allergen (i.e., OVA). This RSV-enhanced allergy is not completely abrogated by prior inoculation with RSV.     

呼吸道合胞病毒对致敏小鼠T辅助细胞相关细胞因子表达的作用

目的　探讨经卵白蛋白 (OVA)雾化致敏的小鼠受呼吸道合胞病毒 (RSV)感染后肺内炎症发展的特点及与T辅助细胞有关的细胞因子的反应。方法　 40只小鼠随机分成 4组 ,每组 1 0只。对照组 :磷酸盐缓冲液 (PBS)雾化 1 0天 ,HEP 2细胞液滴鼻 ;RSV组 :PBS液雾化 ,RSV滴鼻 ;OVA组 :OVA致敏 ,HEP 2细胞液滴鼻 ;OVA +RSV组 :OVA致敏 ,RSV滴鼻。第 1 6天 ,每组各 6只小鼠支气管肺泡灌洗作细胞计数及分类 ,酶链免疫吸附试验 (ELISA)测定白细胞介素 4(IL 4)、γ干扰素 (IFN γ)的含量 ;余 4只取肺组织用于病理分析和提取总RNA ,经逆转录 聚合酶链反应 (RT PCR)测IL 4、IL 5、IFN γ的mRNA表达量。结果　 (1 )支气管肺泡灌洗液 (BALF)中细胞总数 :单纯RSV组为 (1 4 5±5 4)× 1 0 6 /ml、OVA +RSV组为 (1 6 8± 4 9)× 1 0 6 /ml,与对照组 [(7 7± 2 4)× 1 0 6 /ml]比较差异有显著性 (P <0 0 5) ;淋巴细胞RSV组为 0 63± 0 0 5、OVA +RSV组为 0 77± 0 0 9,与OVA组 (0 36± 0 0 3)、对照组 (0 2 8± 0 0 5)比较差异有显著性 (P <0 0 5) ;嗜酸细胞 :OVA +RSV组 (0 0 690± 0 0 1 0 0 )与其它三组 (0 0 0 30± 0 0 0 1 0、0 0 0 90± 0 0 0 50、0 0 1 0 0± 0 0 0 4 0 )比较差异也有显著性 (P均 <0     

中性粒细胞性支气管哮喘小鼠模型建立的探索

目的利用不同剂量脂多糖（lipopolysaccharides,LPS）干预卵清蛋白（ovalbumin,OVA）致敏和激发的小鼠,探索中性粒细胞性哮喘（neutrophilic asthma,NA）小鼠模型的建立。方法 80只BALB/c小鼠随机分为正常对照组（A组）、肺损伤对照组（B组：B1～B4）、嗜酸细胞性哮喘模型对照组（C组）、NA模型探索组（D组：D1～D4）。分致敏和激发两阶段建模,致敏阶段：A组PBS腹腔注射及PBS滴鼻,B组PBS腹腔注射及LPS滴鼻,C组OVA腹腔注射,D组OVA腹腔注射及LPS滴鼻;激发阶段：A、B组生理盐水雾化,C、D组5%OVA雾化。通过观察小鼠雾化时的症状、肺组织病理改变,检测支气管肺泡灌洗液（bronchoalveolar lavage fluid,BALF）细胞总数及分类计数和血清OVA-sIgE等评价NA小鼠模型建立。结果①D3组小鼠出现类似C组小鼠呼吸急促、大小便失禁等表现。②D3组BALF炎症细胞总数较A组显著升高（P〈0.01）;D3组中性粒细胞（neutrophil,NEU）百分比（NEU%）较A、C、D1、D2组显著升高（P值均〈0.01）,嗜酸粒细胞（eosinophil,EOS）百分比（EOS%）较A组显著升高而较C组显著低下（P值均〈0.01）。③D3组支气管管壁及肺泡间隔增厚变形,部分断裂,气道黏膜下除了EOS浸润外还存在明显NEU浸润。④D3组血清OVA-sIgE水平显著高于A组而低于C组（P值均〈0.05）。⑤D3组IL-4水平显著高于A组（P〈0.05）,与C组无显著差异（P〉0.05）;D3组IFN-γ水平显著低于A组（P〈0.05）,与C组无显著差异（P〉0.05）。结论 10μg LPS滴鼻吸入＋50μg OVA腹腔注射三次与5%OVA连续激发两周即D3组可成功建立NA小鼠模型。