CD34~ 细胞在变应性鼻炎外周血和鼻黏膜中的表达

目的:探讨CD34 细胞在变应性鼻炎患者外周血和鼻黏膜中的表达及髓外CD34 细胞可能参与变应性鼻炎鼻黏膜中的炎性细胞特别是嗜酸粒细胞(eosinophils,EOS)局部生成的可能性。方法:取变应性鼻炎发作期和非发作期患者各12例,两组患者分别取外周血,用流式细胞仪做CD34 细胞记数;分别取鼻黏膜,用免疫组化方法标记CD34 细胞,在尽可能排除血管内皮细胞和纤维细胞CD34 阳性表达后,计数CD34 细胞。结果:变应性鼻炎患者发作期外周血中CD34 细胞表达明显高于非发作期患者外周血中CD34 细胞的表达(P<0．05), 鼻黏膜中CD34 细胞主要表达于黏膜下层EOS聚集区,非发作期患者鼻黏膜中CD34 细胞表达明显高于发作期的鼻黏膜(P<0．01),并与活化的EOS数呈负相关。结论:变应性鼻炎外周血、鼻黏膜中 CD34 细胞的表达,提示了髓外CD34 细胞可能参与变应性鼻炎鼻黏膜中的炎性细胞特别是EOS的局部生成。图8表2参10     

变应性鼻炎患者外周血CD34+和白介素5及嗜酸粒细胞的检测

变应性鼻炎鼻黏膜组织中嗜酸粒细胞(eosinophil，EOS)的浸润与症状的严重程度密切相关，但有关EOS在鼻黏膜中持续增多的机制尚未完全阐明，但有学者认为变应性鼻炎和骨髓问可能存在一种反馈机制。已知有多种细胞因子参与了EOS的生成和趋化，其中以白细胞介素5(interleukin5，IL-5)最为重要。本研究通过流式细胞术、酶联免疫法和外周血EOS常规计数等方法，观察28例健康人和32例变应性鼻炎患者外周血CD34^ 细胞、IL-5、EOS计数的变化，以探讨它们之间的关系。     

血红素氧合酶1在变应性鼻炎豚鼠模型的鼻黏膜中的表达

目的:通过制备豚鼠变应性鼻炎动物模型,观察并分析内源性一氧化碳(CO)的限速酶血红素氧合酶1(HO1)在豚鼠鼻黏膜组织中的表达。方法:分别以卵清蛋白致敏制备豚鼠变应性鼻炎动物模型为致敏组及以地塞米松处理作为治疗组,以生理盐水处理作为正常对照组,取豚鼠鼻黏膜,苏木精伊红染色观察炎性细胞浸润,免疫组织化学染色方法观察HO1在各组豚鼠的鼻黏膜组织中的表达情况。结果:在黏膜组织中,炎性细胞的浸润与炎症程度有关,炎性细胞以嗜酸粒细胞(EOS)浸润明显,HO1主要表达在黏膜腺上皮细胞的胞质中,3组中,致敏组鼻黏膜HO1表达明显增强(P<0.01),黏膜下EOS浸润明显(P<0.01),激素治疗组HO1的表达弱于致敏组(P<0.01),黏膜下EOS浸润程度减轻,但强于对照组(P<0.01),组间差异有统计学意义(均P<0.01)。结论:HO1主要表达于鼻黏膜的腺上皮细胞质内,豚鼠变应性鼻炎模型中HO1的表达与炎症的严重程度相关,提示内源性CO可能参与了变应性鼻炎的炎症过程。     

变应性鼻炎患者外周血嗜酸粒细胞-骨髓干细胞通路指标检测在激素治疗评价中的意义

目的 检测健康人及变应性鼻炎(AR)患者治疗前后外周血嗜酸粒细胞(eosinophils,EOS)-骨髓干细胞通路相关指标CD34+、白细胞介素5(IL-5)、EOS,探讨外周血-骨髓通路在变应性鼻炎发病机制中的作用以及糖皮质激素对此通路的影响.方法 实验分2组:①试验组:常年性持续性变应性鼻炎患者44例,男24例,女20例,年龄7～68岁;给予糖皮质激素治疗4周;②健康对照组:健康体检者30例.分别检测试验组治疗前后和对照组外周血EOS计数,血清IL-5水平及CD34+细胞数,并分析各指标间的相关性.结果 试验组治疗前血清IL-5含量、CD34+数分别为(88.25±33.47)ng/L、(9.24±2.15)个/105,显著高于治疗后[(44.34±16.32)ng/L、(6.31±1.83)个/103]及健康对照组[(31.24±8.43)ng/L、(3.47±1.32)个/105].试验组治疗前后血清IL-5水平与其CD34+数呈显著正相关(r值分别为0.64、0.61,P值均＜0.01).患者血清IL-5水平与其EOS数呈显著正相关(r=0.64,P＜0.01).结论 外周血EOS、IL-5及CD34+细胞参与AR发病过程,提示AR患者病变局部组织和骨髓造血之间有相关通路存在.通过检测外周血IL-5及CD34+可评价治疗效果.     

CD30在变应性鼻炎患者鼻黏膜组织中的表达

目的 :探讨CD30在变应性鼻炎患者鼻黏膜组织中的表达 ,分析其在变应性鼻炎发病中的作用。方法 :应用免疫组织化学SP法和苏木精 伊红染色对 31例变应性鼻炎患者的下鼻甲黏膜 (变应性鼻炎组 )和 2 6例肥厚性鼻炎患者的下鼻甲黏膜 (对照组 )标本中CD30的表达进行检测 ,并进行统计学分析。结果 :变应性鼻炎组CD30细胞数显著高于对照组 ,其差异有统计学意义 (P <0 .0 5 )。结论 :在变应性鼻炎的鼻黏膜组织中存在着Th2细胞主导的黏膜反应 ,通过研究CD30的表达水平可了解变应性鼻炎中Th2反应的状态。     

变应性鼻炎患者鼻黏膜中组胺H4受体的差异性表达

目的:观察新型组胺受体H4在正常人及变应性鼻炎患者鼻腔黏膜中的表达及分布, 初步了解组胺H4受体在变应性鼻炎发病中的作用.方法:选取正常健康者及变应性鼻炎患者各10例,取鼻黏膜分别通过免疫组织化学及RT-PCR方法检测组胺H4受体在蛋白质及转录水平的表达及分布情况并进行对比.结果:变应性鼻炎患者鼻黏膜中组胺H4受体的表达(49676±8541,0.69±0.11)较正常人(25509±6441,0.42±0.08) 显著增加(P<0.05),鼻黏膜结构细胞及免疫细胞均见H4受体表达.结论:H4受体存在于正常人鼻黏膜中,在变应性鼻炎患者鼻黏膜中表达显著增强,提示组胺H4受体可能是介导组胺参与变应性鼻炎发病的重要配体之一.     

转录因子T-bet在变应性鼻炎患者鼻咽相关淋巴组织中的表达特征和意义

目的:探讨T-bet在变应性鼻炎(AR)和非变应性鼻炎(NAR)2组人群的鼻咽相关淋巴组s织(NALT)中的表达特征和意义。方法:分别用免疫组织化学单染色法检测AR患者和NAR人群扁桃体、腺样体和鼻黏膜中的T-bet阳性表达,用免疫组织化学双染色法检测CD4/T-bet表达。结果:在AR组扁桃体、腺样体和鼻黏膜组织中T-bet表达阳性率高于NAR组,差异有统计学意义(均P<0.05);AR组不同年龄之间扁桃体T-bet表达存在差异(P<0.05),其中主要是成年人T-bet表达低于儿童和青少年;在NAR组扁桃体组织T-bet表达没有年龄之间的差异。2组中扁桃体、腺样体和鼻黏膜中均有CD4和T-bet表达,大多数T-bet阳性表达位于CD4 T细胞中。结论:在AR患者NALT中T-bet的下调表达,与AR患者呼吸道局部黏膜免疫转录因子水平的Th1反应低下有关。NALT中的T-bet表达既与黏膜的变应性炎症有关,又与NALT的功能状态有关。     

CD4+CD25+调节性T细胞在变应性鼻炎患者外周血淋巴细胞中的含量研究

目的:研究CD4+CD25+调节性T细胞(CD4+CD25+Treg)在变应性鼻炎外周血淋巴细胞中的比例,并探讨其临床意义.方法:采用流式细胞术分别检测20例季节性变应性鼻炎患者发作期外周血CD4+CD25+Treg在淋巴细胞中的比例,并与8例健康对照者进行比较.结果:变应性鼻炎患者外周血CD4+CD25+Treg和CD4+CD25highTreg所占比例与对照组比较差异均有统计学意义(均P<0.05).结论:变应性鼻炎患者外周血CD4+CD25+Treg在外周血中的比例明显减少,可能与该病的发病机制有关.     

免疫治疗调节变应性鼻炎外周血调节性T细胞表达

变应性鼻炎是机体接触变应原后主要由免疫球蛋白IgE介导的鼻黏膜变态反应性炎症,大量实验研究表明变应性鼻炎是由于Thl/Th2失衡引起的。近年来研究表明Thl7与调节性T细胞在变应性鼻炎的免疫反应的发生和发展中有着重要的作用。调节性T细胞功能降低时Th2反应占优势。研究还表明变应性鼻炎患者的外周血中调节性T细胞的表达是降低的,且在进行免疫治疗后,其外周血的CD4+CD25+调节性T细胞细胞表达较之前增多。     

白介素12基因治疗对变应性鼻炎小鼠嗜酸粒细胞及白介素5的影响

目的 探讨经鼻局部给予脂质体包裹的白细胞介素(interleukin,IL)12基因治疗对变应性鼻炎(allergic rhinitis,AR)小鼠模型鼻黏膜、外周血和骨髓中嗜酸粒细胞(eosinophils,EOS)的调节作用及相关因子IL-5的影响.方法 采用6～8周雄性BALB/C小鼠,随机分成AR组、基因治疗组和健康对照组,每组12只.卵清蛋白(ovalbumin,OVA)致敏激发建立AR模型,治疗组激发前经鼻给予脂质体包裹的pGEG.m IL-12,对照组用生理盐水代替.三组分别用HE染色计数鼻黏膜中EOS的数量,用瑞氏染色计数骨髓涂片中EOS数,以流式细胞仪检测外周血中的EOS数;免疫组化染色检测鼻黏膜和骨髓中IL-5的表达,以ELISA方法检测血清中的IL-5含量.采用单因素方差分析进行统计学处理.结果 三组小鼠中,各检测指标在各组之间的差异均有统计学意义(P值均<0.01).两两比较发现,基因治疗组鼻黏膜EOS数为(4.6±2.6)个/高倍镜视野,低于AR组的(26.5±9.8)个/高倍镜视野,差异有统计学意义(P<0.05);IL-5阳性细胞数[(3.0±1.3)个/高倍镜视野]也低于AR组[(17.6±6.4)个/高倍镜视野],差异有统计学意义(P<0.05);骨髓涂片中EOS(0.040±0.029)低于AR组(0.086±0.014),差异有统计学意义(P<0.05),IL-5阳性细胞数(0.035±0.012)也低于AR组(0.083±0.025),差异有统计学意义(P<0.05).基因治疗组外周血中EOS(0.124±0.031)低于AR组(0.184±0.079),差异有统计学意义(P<0.05),IL-5[(29.51±6.68)pg/ml]也低于AR组[(56.58±16.80)pg/ml],差异有统计学意义(P<0.05).结论 经鼻局部给予脂质体包裹的pGEG.m IL-12能够通过降低骨髓、外周血和鼻黏膜中IL-5的表达,进而减少骨髓、外周血和鼻黏膜中EOS的数量,IL-12基因治疗可能为呼吸道变应性炎症开辟一种新的治疗途径.     

Isolation and phenotypic characterization of mucosal nasal lymphocytes by direct ex vivo analysis

Cellular inflammation of the nasal mucosa demonstrates a local immune response which plays an important role in allergic rhinitis. The aim of the present study was to characterize nasal mucosal lymphocytes regarding their activation and differentiation state by direct ex vivo flowcytometric analysis. Lymphocytes from the inferior turbinates were isolated by a mechanical method of preparation and, for comparison, from peripheral blood by Ficoll gradient centrifugation. Patients suffering from rhinitis or difficulty in nasal breathing were divided into an allergic (pollen-allergy, n = 13) and non-allergic group (n = 24). Expression of different T- and B-cell markers was determined by flowcytometric analysis. CD4+ T-cells from the nasal mucosa exhibited a memory phenotype (CD45RO+, 97%), were highly activated (CD69+, 43–73%), and showed low expression of the cutaneous lymphocyte antigen (CLA+, 5%). Nasal CD20+ B-lymphocytes expressed significantly higher levels of mIgE and lower levels of CD23 and CD80 than peripheral B-cells. Subsets of CD80+ (4%) and CD86+ (6%) CD20+ B-lymphocytes were identified in the nasal mucosa. No significant differences between allergic and non-allergic individuals were determined. As expected, the data show profound phenotypical differences between circulating peripheral blood and nasal mucosal lymphocytes. Activated memory lymphocytes are present in the nasal mucosa from allergic, but also non-allergic patients and may indicate to a significant role of a local inflammatory state without systemic criteria for allergy. In our study, we show that direct ex vivo isolation of lymphocytes is practicable method and offers a new technique to examine the local nasal allergic immune response using a multiparametric phenotypical analysis. Christin Wolfram and Claudia Rasche contributed equally to this work.     

白细胞介素12基因治疗小鼠变应性鼻炎的实验研究

目的探讨鼻腔局部应用EB病毒(Epstein-Barrvirus,EBV)质粒载体介导的白细胞介素12(interleukin-12,IL-12)基因治疗对变应性鼻炎炎症反应的调节作用。方法将36只6～8周雄BALB/C实验小鼠随机分为变应性鼻炎组、IL-12基因治疗组和正常对照组,每组12只。以BALB/c小鼠经卵清蛋白(ovalbumin,OVA)免疫建立变应性鼻炎模型,用阳离子脂质体包裹EBV质粒载体介导的IL-12表达质粒(pGEG.mI-L12)形成混合物EBV/lipoplex,于激发前鼻腔局部滴入后,观察小鼠变应性症状的改善情况,并检测该基因在3组实验鼠鼻黏膜局部的表达情况以及对鼻黏膜炎性细胞和Th2细胞因子的影响。结果基因治疗组小鼠鼻黏膜中IL-12mRNA阳性细胞数量明显高于变应性鼻炎组,差异有统计学意义(P<0.01);基因治疗组小鼠鼻黏膜中IL12阳性细胞数量明显高于变应性鼻炎组(P<0.05);变应性鼻炎组鼻黏膜中嗜酸粒细胞、肥大细胞和IL5阳性细胞比例显著高于基因治疗组和正常对照组(P<0.01);变应性鼻炎组外周血中总IgE含量显著高于正常对照组和基因治疗组(F=1216.21,P<0.01)。结论鼻腔局部应用EBV/lipoplex后,pGEG.mIL-12能够在鼻黏膜中高效地表达,能明显抑制鼻腔的变应性反应。EBV/lipoplex有望成为变应性鼻炎免疫治疗一种新的方法。     

The effect of the H2 antagonist cimetidine on the numbers of CD4+ and CD8+ cells in the nasal mucosa of patients with allergic rhinitis

This paper reports the effects of the H₂ antagonist cimetidine on the number of CD4⁺ and CD8⁺ cells in nasal mucosa and the IgE level of nasal secretions in patients with allergic rhinitis. The results showed the numbers of CD4⁺ cells were greater than the numbers of CD8⁺ cells in nasal mucosa, both in the patients with allergic rhinitis and normal subjects, but the ratio of CD4⁺ : CD8⁺ cells was much higher in the patients with allergic rhinitis. After treatment with cimetidine locally for 4 weeks, the numbers of CD4⁺ cells fell and the numbers of CD8⁺ cells increased in the patients with allergic rhinitis. The high IgE level of nasal secretion of the patients with allergic rhinitis was much reduced after treatment with cimetidine. The results suggest that there are high numbers of CD4⁺ cells and lower numbers of CD8⁺ cells in the nasal mucosa and a high level of IgE in the nasal secretions of the patients with allergic rhinitis. Treatment with cimetidine locally may be of some value to relieve the clinical symptoms of allergic rhinitis.     

复发性鼻息肉中T淋巴细胞亚群的表达

目的 :检测复发性鼻息肉组织中T淋巴细胞亚群的表达 ,探讨T淋巴细胞介导的免疫反应在鼻息肉复发中所起的作用。方法 :应用荧光免疫流式细胞术检测 17例复发性鼻息肉患者鼻息肉组织、外周血T淋巴细胞亚群CD4 、CD8 、CD4 5RO 的表达 ,并与正常人下鼻甲黏膜及外周血的相应指标进行比较。 结果 :复发性鼻息肉组织中有大量T淋巴细胞浸润 [(39.6 5±2 .0 8) % ],而在健康下鼻甲黏膜中几乎未见CD3 细胞。在复发性鼻息肉中 ,CD3 CD4 细胞[(6 4.4 46± 5 .2 97) % ]多于CD3 CD8 细胞 [(35 .5 5 4± 5 .2 97) % ](P <0 .0 5 ) ,CD3 CD4 5RO 细胞 [(2 2 .6 49± 2 .789) % ]也显著多于正常人外周血CD3 CD4 5RO 细胞 [(3.896± 0 .384 ) % ](P<0 .0 5 ) ,CD3 CD4 /CD3 CD8 比值为 1.95 6± 0 .0 93,复发性鼻息肉患者外周血比值为 2 .36 7±0 .12 8,正常人外周血比值为 1.6 0 6± 0 .0 96 ,其差异均有统计学意义 (均P <0 .0 5 )。结论 :复发性鼻息肉组织中有大量T淋巴细胞表达 ,且T细胞亚群比例失调 ,显示细胞免疫功能的紊乱在鼻息肉的形成与复发中起重要作用     

Distribution of rantes and interleukin-5 in allergic nasal mucosa and nasal polyps.

Eosinophil-chemoattracting cytokines are thought to be important in the pathogenesis of allergic inflammation. However, little is known about the presence and significance of RANTES in nasal allergy and nasal polyps, two well-known rhinologic disorders characterized by eosinophil infiltration in the tissue. In order to evaluate the role of RANTES in eosinophil infiltration in vivo, the tissue distributions of RANTES and interleukin-5 (IL-5) and their correlation with eosinophil infiltration were investigated. Nasal mucosa specimens were obtained from 9 allergic and 12 control subjects, and nasal polyps from 6 allergic and 9 nonallergic subjects. All the subjects were divided into 4 groups: normal mucosa, allergic mucosa, nonallergic polyps, and allergic polyps. To identify the cellular localizations of RANTES and IL-5, we used specific immunohistochemical staining. We also investigated the differences in cytokine expression among the 4 groups, and the correlation between cytokine expression and eosinophil infiltration in the tissue. RANTES was expressed in the epithelium, endothelium, and some submucosal cells, while IL-5 was confined to the cells in the submucosa. Expression of both RANTES and IL-5 significantly increased in allergic mucosa and nasal polyps compared to normal mucosa; however, there was no significant difference in their expression between allergic and nonallergic polyps. Both cytokines had a significant correlation between their expression and either total or activated eosinophil numbers. The results of this study suggest that RANTES, as well as IL-5, plays a role in eosinophil recruitment in allergic nasal mucosa and nasal polyps in vivo.     

骨髓反应与变应性鼻炎和变应性哮喘的相关性研究

目的检测变应性鼻炎和变应性哮喘动物模型骨髓中CD34^ 祖细胞,探讨IL-5对大鼠骨髓单核细胞增殖的刺激作用。方法采用6～8周雄性SD大鼠,随机分成变应性鼻炎组10只和对照组10只,变应性哮喘组10只和对照组10只,以卵清蛋白致敏激发制成变应性鼻炎和变应性哮喘模型,免疫组化检测大鼠骨髓涂片中CD34^ 祖细胞;取对照组大鼠骨髓单核细胞,培养7天,分为对照组、IL-5组计数集落数。结果两组动物模型骨髓涂CD34^ 祖细胞比例显著高于其对照组;大鼠骨髓单核细胞培养7天后,各IL-5组单核细胞集落数显著多于对照组。结论变应性鼻炎和变应性哮喘模型中骨髓CD34^ 祖细胞增殖,IL-5能刺激骨髓单核细胞增殖形成集落。提示骨髓反应是变应性疾病重要的全身反应机制。     

Identification of CD11c+ myeloid dentritic cells in adenoids and in nasal mucosa of patients with and without allergies

BACKGROUND: Dendritic cells form a link between innate and acquired immunity. They are capable to detect pathogens based on the recognition of pathogen-associated microbial molecules and trigger the appropriate type of immune responses. In humans, three major subsets of dendritic cells can be distinguished, Langerhans cells of the skin, myeloid DC (MDC) and plasmacytoid DC (PDC). It was reported that PDC infiltrate nasal mucosa in allergen-induced rhinitis. Information about the role of MDC in nasal mucosa and the corresponding mucosa-associated lymphoid tissue, the nasopharyngeal adenoids, is limited. PATIENTS AND METHODS:: Here we examined the presence of MDC in adenoids and in nasal mucosa of healthy individuals (n = 9) and in patients with allergic rhinitis. MDC were detected by flow cytometry by positive staining for MHC II and CD11c and the lack of lineage markers. Dead cells were excluded from analysis. RESULTS: In adenoids, 0.4 % of all cells were MDC. Considerable numbers of MDC could also be detected in nasal mucosa. No difference was found between healthy individuals and patients with allergies (0.3 % vs. 0.45 % MDC; p = 0.12). Interestingly, MDC were absent in patients who received treatment with glucocorticoids, while very high numbers of MDC were found in patients who recently had upper respiratory tract infections. CONCLUSION: Our results demonstrate for the first time the presence of MDC in nasal mucosa. MDC numbers were similar in healthy individuals and in patients with allergy. This study forms the basis for examining the role of MDC in the pathogenesis of allergic rhinitis, and for the modulation of MDC functional activity with microbial molecules such as CpG oligonucleotides.     

蛋白激酶C和激活蛋白-1信号级联在变应性鼻炎患者T淋巴细胞IL-5表达中的调控作用

目的：探讨蛋白激酶C（PKC）和激活蛋白-1（AP-1）信号转导级联在变应性鼻炎（AR）患者外周血T淋巴细胞IL-5表达中的作用。方法：25例AR患者和23例鼻中隔偏曲（DNS）患者为研究对象，分别从每位受试者外周血中分离T淋巴细胞，并随机分为空白组、PKC激动剂12-肉豆蔻酰-13-乙酸佛波酯（PMA）组、PMA和AP-1抑制剂姜黄素组进行培养。将培养的T淋巴细胞涂片，用免疫细胞化学染色方法检测AP-1的表达，用酶联免疫吸附（ELISA）法检测上清液中的IL-5含量。结果：①加PMAAR组T淋巴细胞的AP-1活化细胞百分比和培养上清液中的IL-5与DNS空白组与AR空白组比较，均差异有统计学意义（均P〈0．01）；与加PMADNS组及加PMA和姜黄素DNS组T淋巴细胞比较，均差异有统计学意义（均P〈0．01）；与加PMA和姜黄素AR组T淋巴细胞比较，差异亦有统计学意义（P〈0．01）；②加PMA和姜黄素AR组T淋巴细胞AP-1活化细胞百分比、培养上清液中的IL-5含量与AR空白组、加PMAAR组、DNs空白组、加PMADNS组、加PMA和姜黄素DNS组比较，均差异有统计学意义（均P〈0．01）；③T淋巴细胞的AP-1活化与IL-5表达呈显著正相关（r=0．92，P〈0．01）。结论：AR患者T淋巴细胞PKC活化后促进IL-5表达增加的生物信号可能是通过AP-1进行转导，提示T淋巴细胞PKC—AP-1信号转导级联的激活可能是AR发病机制之一。