卡介苗对哮喘小鼠TH1/TH2平衡的影响

目的　探讨卡介苗 (BCG)对哮喘小鼠TH1 TH2平衡的影响。方法　C5 7BL 6小鼠以卵白蛋白 (OVA)致敏激发建立哮喘模型。于第一次抗原激发前 2周以BCG皮内注射干预 ,在最后 1次抗原激发后 4 8h收集支气管肺泡灌洗液 (BALF)和外周血 ,ELISA方法测定外周血清及BALF中IL 5与IFN γ水平 ,并采用三色光流式细胞分析法测定外周血TH1 TH2细胞比。结果　与阴性对照组相比 ,OVA致敏激发组外周血清及BALF中IL 5水平升高而IFN γ水平降低 ,BCG干预组外周血清及BALF中IL 5较OVA致敏激发组低而IFN γ较后者升高 ,与阴性对照组水平相近。此外 ,OVA致敏激发组外周血中TH2 TH1比值 (1.5 7± 0 .5 6 )较阴性对照组 (0 .37± 0 .0 5 )明显增高 (P <0 .0 1) ,BCG干预后TH2 TH1比值 (0 .5 9± 0 .11)较OVA致敏激发组下降 (P <0 .0 5 ) ,同时Tc2 Tc1比值 (0 .6 2± 0 .0 7)也较OVA致敏激发组 (1.15± 0 .18)降低 (P <0 .0 5 )。结论　哮喘小鼠中TH2型免疫应答占优势 ,BCG干预不仅在细胞因子水平 ,也在细胞数量上校正了TH1 TH2失平衡。     

卡介苗对哮喘小鼠气道炎症、TH细胞分化及血清OVA特异性IgE的影响

目的 探讨卡介苗（BCG）对哮喘小鼠气道炎症、TH细胞分化及血清OVA特异性IgE的影响。方法 动物随机分为阴性对照组、OVA致敏激发组和BCG干预组。检测气管血管旁嗜酸性粒细胞（EOS）数目、气管上皮杯状细胞增生指数和黏液储存指数、小鼠外周血上清液中细胞因子白细胞介素5（IL-5）及γ干扰素（IFN-γ）浓度、小鼠肺组织中T-bet和GATA-3mRNA水平及小鼠血清中OVA特异性IgE表达。结果BCG干预组和OVA致敏激发组中小鼠肺气管血管旁单位面积EOS数目分别为（65．09±32．58）个/mm^2、（941．86±592．03）个/mm^2，两者比较差异有统计学意义（P〈0．05）；BCG干预组和OVA致敏激发组杯状细胞增生指数分别为（19．25±12．27）个／mm、（54．39±24．50）个／mm，BCG干预组和OVA致敏激发组黏液储存指数分别为（6．47±3．94）％、（15．53±11．38）％，两者比较差异有统计学意义（P〈0．001）；BCG干预组和OVA致敏激发组IL-5浓度分别为（83．40±11．33）pg／ml、（192．30±44．30）pg/ml，1FN-γ浓度分别为（28．69±6．05）pg/ml、（18．12±3．78）pg/ml，T-bet／GATA．3mR—NA比值分别为1．02±0．18、0．74±0．07，两者比较均差异有统计学意义（P〈0．01）；BCG干预组和OVA致敏激发组血清中OVA特异性IgE的表达分别为0．61±0．27、1．05±0．45，两者比较差异有统计学意义（P〈0．05）。结论 BCG能抑制哮喘小鼠气道EOS浸润和黏液高分泌状态，影响T-bet／GATA-3表达水平，降低IL-5水平而增高IFN-γ水平，并能抑制哮喘小鼠血清中OVA特异性IgE的表达。     

嗜酸性粒细胞提呈抗原对肺组织CD4+细胞分化的影响

目的:探讨嗜酸性粒细胞(EOS)对肺组织中的CD4 细胞分化的影响。方法:以鸡卵清蛋白(OVA)致敏和雾化吸入刺激BALB/c小鼠以诱发气道的EOS在气道聚集后将其分离纯化,然后与来源于致敏小鼠肺组织的CD4 细胞共同培养。在部分试验中加入抗CD80和(或)抗CD86mAb。同时,将EOS注入致敏小鼠气道,3d后,取出肺组织,并将其制成细胞悬液进行培养。收集培养上清液,用ELISA测定其中IL-4、IL-5、IL-13及INF-γ的含量。结果:在和EOS体外共同培养的条件下,肺组织CD4 淋巴细胞产生IL-4、IL-5、和IL-13等Th2类因子,但不产生INF-γ等Th1类因子;抗CD80和(或)抗CD86mAb可以明显抑制Th2类细胞因子的产生。将EOS注入致敏小鼠气道后,同样在体内能激发肺组织CD4 活化,产生Th2细胞因子IL-4,IL-5,IL-13,但不产生INF-γ,与体外结果相似,抗CD80和(或)抗CD86mAb可以抑制EOS在体内的抗原提呈过程。结论:EOS细胞在体内或体外均能摄取和处理抗原,激发CD4 的Th2反应。     

地塞米松对呼吸道合胞病毒感染哮喘加重小鼠气道TSLP分泌及炎症的影响

目的:探讨地塞米松(DEX)对呼吸道合胞病毒(RSV)感染哮喘加重小鼠胸腺基质淋巴细胞生成素(TSLP)分泌及气道炎症的影响。方法:雌性BALB/c小鼠32只,随机分成4组,分别为磷酸盐缓冲液(PBS)对照组、鸡卵白蛋白(OVA)组、OVA/RSV组、OVA/RSV/DEX组;应用OVA腹腔注射致敏、OVA气道雾化结合RSV滴鼻激发哮喘,地塞米松1mg/kg肌肉注射;无创肺功能检测各组小鼠气道反应性;ELISA法检测小鼠血清IL-4、IL-5、IL-13、IFNγ-和气管灌洗液(BALF)TSLP含量;小鼠肺组织病理观察炎症反应,免疫组化观察小鼠气道上皮细胞TSLP表达水平。结果:无创肺功能检测显示地塞米松抑制RSV感染哮喘加重小鼠气道反应性的增高,OVA/RSV/DEX组小鼠Penh值明显低于OVA/RSV组(P<0.01);OVA/RSV/DEX组小鼠血清IL-4、IL-5、IL-13、IFNγ-浓度[分别为(86.78±27.04)、(227.66±40.87)、(194.65±73.27)和(17.33±3.06)pg/ml]和BALF中TSLP浓度[(1 873±10)pg/ml],均明显低于OVA/RSV组[分别为(274.2±103.7)、(293.3±46.1)、(330±93.5)、(30.1±5.7)、(2 127±46)pg/ml](P<0.01);病理观察显示地塞米松显著减轻RSV感染哮喘小鼠气道炎症细胞浸润;免疫组化染色证实地塞米松抑制RSV感染哮喘小鼠气道上皮细胞TSLP表达。结论:地塞米松可以抑制RSV感染哮喘加重小鼠气道上皮细胞表达TSLP,减轻RSV感染哮喘加重小鼠气道炎症反应。     

PPD对TH1细胞的在体调节及对卵蛋白致敏小鼠气道炎症的影响

目的：探索结核菌素纯蛋白衍生物（purified protein derivative，PPD）对卵蛋白（ovalbumin，OVA）致敏小鼠肺组织TH1细胞的在体调节和对气道炎症的。方法：取37只Balb/c小鼠，雌雄兼有，6-8周。OVA雾化连续10d，每次雾化20min，复制OVA致敏模型。观察记录小鼠耗氧量、肺组织淋巴细胞反应和支气管肺泡灌洗液（bronchoalveolar lavage fluid，BALF）中炎性细胞变化。结果：OVA致敏组小鼠肺组织CD4^ T淋巴细胞增加，CD8^ T淋巴细胞无明显变化，主要表现为γ-IFN阴性CD4^ T淋巴细胞的增加，γ-IFN^ /CD4比较降低。PPD治疗后，肺组织CD4^ 淋巴细胞降低，但γ-IFN^ 细胞明显增加。OVA致敏小鼠消耗氧增加，PPD治疗后降低。结论：使用本实验体系PPD可上调TH1细胞并减轻实验性哮喘的气道反应。     

呼吸道合胞病毒对致敏小鼠气道炎症和CD8+ T细胞功能的影响

目的观察呼吸道合胞病毒（RSV）对致敏小鼠气道炎症和CD8^＋T细胞功能的影响.方法BALB/c小鼠40只,随机分成4组,分别为磷酸盐缓冲液（PBS）对照组、鸡卵白蛋白（OVA）组、RSV组、OVA/RSV组;应用OVA腹腔注射致敏、OVA气道雾化结合RSV滴鼻激发哮喘;支气管肺泡灌洗液（BALF）作细胞分类计数;酶联免疫吸附试验（ELISA）测定BALF上清中白细胞介素（IL）-4、IL-5、干扰素（IFN）-γ含量;苏木精-依红（HE）染色观察肺病理变化;采用三色光流式细胞分析法测定气管旁淋巴结（PBLN）中CD4^＋、CD8^＋T细胞及细胞内细胞因子IFN-γ、IL-4、IL-5表达与TH2/TH1、Tc2/Tc1比值变化.结果（1）BALF中细胞总数及分类：与OVA组比较,OVA/RSV组细胞总数、嗜酸性粒细胞、淋巴细胞均明显增加（分别P＜0.01）;与RSV组比较,OVA/RSV组细胞总数、嗜酸性粒细胞明显增加（分别P＜0.01）.（2）BALF上清中细胞因子含量：与OVA组比较,OVA/RSV组IFN-γ、IL-4、IL-5含量均明显升高（分别P＜0.01）;与RSV组比较,OVA/RSV组IFN-γ无明显变化,而IL-4、IL-5显著上升（分别P＜0.01）.（3）肺组织病理：OVA/RSV组与其他各组比较气道黏膜增厚,管腔狭窄、收缩,上皮破坏,管壁周围炎症细胞浸润明显加重.（4）PBLN中CD4^＋（WN-γ^＋、IL-4^＋、IL-5^＋）、CD8^＋（IFN-γ^＋、IL-4^＋、IL-5^＋）T细胞各占CD3^＋T细胞百分比及TH2/TH1、Tc2/Tc1比值变化：与OVA组比较,OVA/RSV组CD8^＋（IFN-γ^＋、IL-4^＋、IL-05^＋）T细胞百分比、Tc2/Tc1比值增加（分别P＜0.01）,TH2/TH1比值无明显变化.与RSV组比较,OVA/RSV组CD4^＋（IL-4^＋、IL-5^＋）T细胞、TH2/TH1比值、CD8^＋（IL-4^＋、IL-5^＋）T细胞、Tc2/Tc1比值均明显上升（分别P＜0.01）.结论（1）OVA致敏小鼠RSV感染后可明显加重气道炎症,TH2、TH1型炎症均加强,且以TH2型炎症加重为主.（2）OVA致敏小鼠RSV感染后可引起CD8^＋T细胞数量及功能改变,即由产生IFN-γ^＋为特征的Tc1细胞向产生IL-4^＋、IL-5^＋为特征Tc2细胞转化,并可能与气道内IL-4、IL-5升高及嗜酸性粒细胞的大量募集有关.     

HDM通过肺泡巨噬细胞TLR4诱导哮喘小鼠气道炎症的机制研究

目的:研究HDM通过肺泡巨噬细胞(AM)Toll样受体4(TLR4)的高表达及诱导AM的活化,探讨其对哮喘小鼠气道炎症的影响.方法:BALB/c小鼠随机分为哮喘模型组(OVA)A,HDM处理组(HDM+OVA)B,对照组(生理盐水)C.用卵白蛋白(OVA)致敏与激发建立小鼠哮喘模型;HE染色观察小鼠气道及肺组织病理变化;光学显微镜下观察小鼠支气管肺泡灌洗液(BALF)中细胞分类及计数;酶联免疫吸附试验(ELISA)检测小鼠BALF上清中IL-4、IL-5、IL-13和IFN-γ的含量,实时定量PCR法测定AM的TLR4的表达,流式细胞技术(FCM)检测AM的CD80、CD86的表达.结果:与A组相比,B组BALF上清中IL-4、IL-5和IL-13的水平显著增高(P<0.05),而IFN-γ差异无统计学意义(P>0.05),与A组相比,AM的TLR4mRNA表达明显增高(P<0.05),CD80的表达差异无统计学意义,CD86的表达水平显著增高,差异有统计学意义(P<0.05).结论:HDM通过AM的TLR4的高表达诱导AM的活化,加重哮喘的气道炎症.     

Th17/Treg相关因子在哮喘小鼠中的表达变化及其意义

目的检测哮喘模型小鼠辅助性T17细胞(Th17)和调节性T细胞(Treg)相关细胞因子及受体的表达变化,探讨Th17/Treg在哮喘发病机制中的作用。方法 20只清洁级BALB/C雌鼠,鼠龄6周,体质量18~20 g。随机分为对照组和哮喘组,每组10只。哮喘组小鼠腹腔注射卵白蛋白(OVA)与氢氧化铝的氯化钠混悬液3次后,给予雾化的OVA激发,建立哮喘小鼠模型。对照组给予等量的0.9%氯化钠溶液。末次激发后24 h处死小鼠,检测支气管肺泡灌洗液(BALF)中细胞总数和细胞分类计数;酶联免疫吸附分析(ELISA)、实时定量聚合酶链反应(Q-PCR)和Western blot法测定小鼠外周血和肺组织中的Th17/Treg细胞相关因子及受体的表达水平。结果哮喘组小鼠BALF中细胞总数、嗜酸性细胞比例、中性粒细胞比例、淋巴细胞比例显著高于对照组。哮喘组外周血和肺组织中白细胞介素(IL)-17和IL-18高于对照组[IL-17(124.36±2.14)pg/mL、(21.70±1.02)pg/mL vs(25.98±5.23)pg/mL、(19.95±3.21)pg/mL;IL-18(227.06±24.14)pg/mL、(413.97±32.01)pg/mL vs(120.38±10.32)pg/mL、(238.21±15.33)pg/mL],IL-10和转化生长因子β(TGF-β)低于对照组[IL-10(60.42±24.12)pg/mL、(65.14±15.11)pg/mL vs(61.07±14.36)pg/mL、(104.36±28.16)pg/mL;TGF-β(38.72±12.12)pg/mL、(58.65±24.21)pg/mL vs(30.19±10.16)pg/mL、(121.06±39.12)pg/mL];哮喘组维甲酸相关孤儿素受体γt(RORγt)水平高于对照组[(8.77±2.35)、(4.46±1.05)vs(3.03±1.02)、(3.45±1.25)],肺叉头蛋白3(FoxP3)水平低于对照组[(5.15±1.84)vs(12.35±3.21)];并且外周血和肺组织中相关细胞因子及受体的表达差异具有显著统计学意义(P0.01)。结论 Th17/Treg及其主要相关因子在哮喘的发病中具有重要作用。     

OVA特异性免疫治疗对哮喘小鼠IL-23/Th17轴的影响及机制研究

目的腹部皮下注射大剂量卵白蛋白(OVA)建立小鼠哮喘免疫治疗模型,探讨IL-23/Th17轴在小鼠哮喘模型经皮免疫治疗过程中的作用。方法选择18只BALB/c小鼠为建模研究对象,按随机数字表法分为3组:空白对照组、哮喘对照组和哮喘免疫治疗组,每组6只小鼠。哮喘免疫治疗组予10μg OVA于0、7 d腹腔注射致敏,21~27 d持续1周予1 mg OVA腹部皮下注射诱导免疫耐受,35~41 d予1%OVA雾化激发;哮喘对照组在21~27 d OVA皮下注射等量生理盐水,其余处置同哮喘免疫治疗组;空白对照组采用等量生理盐水致敏及激发。50 d予10%OVA加强激发1次。末次激发24 h内检测气道反应性;收集支气管肺泡灌洗液(BALF)计数细胞总数及细胞分类计数;ELISA法检测血清OVA特异性Ig E、BALF中IL-5、IFN-γ、IL-23、IL-10的表达;HE染色观察肺组织病理改变;流式细胞仪检测各组脾、肺组织Treg、Th17细胞比例;q-PCR检测肺组织转录因子。结果哮喘免疫治疗组气道反应性、BALF中嗜酸性粒细胞计数、IL-23水平及血清OVA特异性Ig E水平明显低于哮喘对照组,差异有统计学意义(P0.05);而BALF中IFN-γ水平与哮喘对照组比较差异无统计学意义(P0.05);同时HE染色发现哮喘免疫治疗组肺组织炎症较哮喘对照组明显减轻;流式细胞技术检测发现外周血Treg细胞百分比明显高于哮喘对照组,差异有统计学意义(P0.05);q-PCR检测肺组织转录因子表明免疫治疗组Foxp3明显高于哮喘对照组,而RORγt明显低于哮喘对照组,差异均具有统计学意义(P0.05)。结论大剂量OVA特异性免疫治疗能够减轻哮喘小鼠气道慢性炎症反应,同时引起Th17细胞下降伴随IL-23表达降低;其机制可能与纠正肺部IL-23/Th17轴有关。     

口服卡介苗对哮喘小鼠TH1/TH2平衡的影响

卡介苗不仅被应用于结核病的预防,还应用于哮喘、慢支、肿瘤等疾病的治疗,国内外学者以皮内注射卡介苗用于实验性哮喘的治疗取得了很好的疗效。但卡介苗口服用于哮喘防治国内还未见报道。本实验通过口服卡介苗后哮喘小鼠支气管肺泡灌洗液(BALF)和外周血中IL-4、IFN-γ水平的变化,并用流式细胞仪来检测Y_H1/T_H2细胞平衡的影响,进一步探讨哮喘发病机理,为口服卡介苗治疗哮喘提供理论依据。32只BALB/c小鼠分为正常对照组、哮喘组、皮内注射卡介苗 哮喘组、口服卡介苗 哮喘组,以卵蛋白(OVA)致敏激发建     

Genetic polymorphism and TH1/TH2 orientation

BACKGROUND: It is likely that besides developmental and environmental factors, genetic factors also play an important role in Th1/Th2 orientation and susceptibility to related disorders. Thus, for each genetic factor involved one would expect an opposite pattern of susceptibility towards Th1- and Th2-associated diseases. METHODS: We report a comparative analysis of the pattern of association of four genetic polymorphisms with bronchial asthma (Th2 disease) and Crohn's disease (CD; Th1 disease). The study population included 291 Roman children with bronchial asthma and 72 adult Romans with CD, and haptoglobin, adenosine deaminase (ADA), acid phosphatase locus 1 (ACP1) and MN phenotypes were determined. RESULTS: Compared with controls from the same population, the pattern of phenotype association observed in bronchial asthma is exactly opposite to that observed in CD. The analysis of pairwise gametic type distribution for ACP1, ADA and MN polymorphisms has shown that the pattern of differences between bronchial asthma and controls is opposite to that observed between CD and controls. CONCLUSIONS: The pattern of differences between bronchial asthma versus CD is compatible with the hypothesis that some of the genetic systems considered contribute to Th1/Th2 orientation.     

Biology of human TH1 and TH2 cells

Evidence is accumulating to suggest the existence of polarized human T-cell responses, reminiscent of T_H1 and T_H2 subsets described for mouse T cells. Human T_H1 cells preferentially develop during infections by intracellular bacteria and trigger phagocyte-mediated host defense, whereas T_H2 cells, which predominate during helminthic infestations and in response to common environmental allergens, are responsible for phagocyte-independent host response. Human T_H1 and T_H2 cells exhibit not only different functional properties but probably also distinct surface markers; T_H2, but not T_H1, clones express membrane CD30 and release the soluble form of CD30, a member of the TNF receptor superfamily. The cytokine profile of natural immunity evoked by different offending agents in the context of different host genetic backgrounds appears to be the most critical factor in determining the phenotype of the subsequent specific response. IL-12 and IFN- and produced by macrophages and NK cells favor the development of T_H1 cells, whereas the early production of IL-4 by a stillunidentified cell type favors the development of T_H2 cells. Clearly, polarized human T_H1 and T_H2 responses not only play different roles in protection, they can also promote different immunopathological reactions. Strong and persistent T_H1 responses seen to be involved in organ-specific autoimmunity, contact dermatitis, and some chronic inflammatory disorders of unknown etiology. In contrast, polarized T_H2 responses favor a reduced protection against the majority of infectious agents (including HIV) and, in genetically predisposed hosts, are responsible for triggering of allergic atopic disorders.     

第9、10、11肋间神经和肋下神经的血供

本文观察了25具成人专供研究用的尸体,对50侧第9、10、11肋间神经和肋下神经的血供进行了研究,对该四对神经的营养动脉数目、外径、长度、入神经干部位和来源动脉进行观察测量。发现营养动脉外径较细,长度亦较短,不宜作为肋间神经的血管蒂进行吻合,而肋间神经营养动脉的来源动脉——肋间后动脉,其外径较粗,并有足够长度,作为血管蒂进行吻合较为理想。     

Immunologic influences on allergy and the TH1/TH2 balance

TH2 cell-mediated immune responses against "innocuous" antigens play a triggering role in atopic allergy. Several epidemiologic studies have clearly shown that the reduced microbial exposure of children caused by the westernized lifestyle is responsible for the increased prevalence of allergy that has occurred in the last decades in developed countries ("hygiene hypothesis"). The immunologic changes caused by the reduced exposure to pathogenic and nonpathogenic microbes during childhood are still controversial. The initial interpretation has been a lack of shift of allergen-specific responses from the TH2 to the TH1 phenotype. This is because of reduced production of IL-12 and IFNs by cells of the natural immunity stimulated by bacterial products through their Toll-like receptors (missing immune deviation). Another interpretation emphasizes the importance of reduced activity of T-regulatory cells (reduced immune suppression). However, although there are impressive amounts of data in favor of the missing immune deviation, experimental evidence supporting the role of reduced immune suppression in explaining the increased prevalence of allergy is currently weak or even contradictory. The solution to this question is very important not only from a theoretic point of view but also because of its therapeutic implications.     

哮喘患儿激素治疗后TH1/TH2的变化

目的观察哮喘患儿血清中白细胞介素IL12、IL13和IgE水平的变化及糖皮质激素对其的影响。方法采用ELISA法分别检测哮喘患儿急性期及口服强的松5～7天后、健康对照组血清中IL12、IL13和IgE的水平。结果哮喘组患儿IL12水平急性发作期较治疗后和健康对照组低,差异均有显著性（P〈0.01）。IL13水平急性发作期较治疗后和健康对照组高,差异均有显著性（P〈0.01）,IgE水平急性发作期较治疗后和健康对照组高,差异均有显著性（P〈0.01）。直线相关分析表明,血清中IL12与IL13负相关,与IgE呈负相关,IL13与IgE呈正相关。结论哮喘患者血清中IL13、IgE水平升高,IL12水平下降,3者的变化相关,糖皮质激素可使IL13水平下降,IL12水平升高。