Toll样受体2和Toll样受体4在小鼠变应性鼻炎中的表达

发布时间：2018-01-16 10:42

本文关键词：Toll样受体2和Toll样受体4在小鼠变应性鼻炎中的表达　出处：《中国医科大学》2010年硕士论文　论文类型：学位论文

【摘要】： 目的变应性鼻炎是一种吸入变应原而导致的以鼻黏膜嗜酸性粒细胞、肥大细胞、T淋巴细胞等多种炎性细胞浸润,IgE介导的免疫细胞释放化学介质引起的Ⅰ型变态反应炎症性疾病,以鼻痒、打喷嚏、流清涕等为主要症状,可并发哮喘、鼻-鼻窦炎和分泌性中耳炎等疾病,严重影响患者生活质量。近年来对变应性鼻炎的研究虽然有了很大进展,但对其发病机制仍需要进一步的研究。Toll样受体(TLRs)是近年发现的介导天然免疫反应的关键受体,它们识别特定的病原相关分子模式并启动了一系列防御反应。由于TLR2和TLR4的配体已经涵盖了自然界绝大多数对人体致病的微生物类别,因此,二者在人Toll样受体家族中最为重要。以前的研究已经证实TLR2和炎症因子之间的协同促进表达在炎症中的重要意义。目前,对于TLRs在变应性鼻炎中的表达及作用机制的研究报道少见,且存在争议。国内报道TLRs在鼻息肉中有高表达,也有国外报道没有发现TLRs在鼻息肉组较健康对照组有高表达,但在变应性鼻炎的急性期发现高表达。由此可见,对TLRs在变应性鼻炎中表达的研究有重要意义。方法 1、变应性鼻炎模型制备 Bal b／c小鼠以卵清白蛋白(OVA)100μg加氢氧化铝(Al(OH)3)2mg溶于0.1ml的生理盐水中腹腔注射,分别在1、3、5、7、9、11、13、15天各一次,共8次,然后于第16天起每侧鼻腔用10μg卵清白蛋白滴鼻激发,连续11天,建立小鼠变应性鼻炎模型。 2、变应性鼻炎模型评价 (1)末次滴鼻激发后观察30分钟,观察小鼠打喷嚏,挠鼻,抓脸,鼻溢等症状并进行计分；(2)抽取致敏动物血,血清中加入标记荧光生物素的变应原,然后在450nm处测定光密度值,来判定血清中IgE的效价；(3)各组小鼠模型行常规HE染色,并行高倍镜下嗜酸性粒细胞计数。 3、应用免疫组织化学染色技术在蛋白水平对变应性鼻炎的Toll样受体2、4的表达进行研究。 4、统计结果均以均数±标准差表示,多组均数间比较用OneWay-ANOVA分析。均用SPSS 13.0软件进行统计分析。结果 1、成功的制备出小鼠变应性鼻炎的模型。 2、变应性鼻炎组小鼠的症状评分与正常对照组比较差异有统计学意义(P0.01)；变应性鼻炎组小鼠IgE效价与正常对照组比较差异有统计学意义(P0.01)；变应性鼻炎组小鼠的嗜酸性粒细胞计数与正常对照组比较差异有统计学意义(P0.01)。 3、免疫组化法发现TLR2和TLR4在正常小鼠鼻黏膜细胞的胞浆和胞核无阳性或极弱阳性染色；在变应性鼻炎组小鼠鼻黏膜细胞胞核和／或胞浆呈不同程度的褐色或黄棕色阳性染色。 4、变应性鼻炎组小鼠在30min时鼻黏膜细胞上TLR2表达开始增多,但与正常对照组差异无统计学意义(P0.05),2h时鼻黏膜细胞上TLR2表达明显增加(P0.01),12h时鼻黏膜细胞上TLR2表达开始下降(P0.05)。24h时鼻黏膜细胞上TLR2表达与正常对照组差异无统计学意义(P0.05)。 5、变应性鼻炎组小鼠在30min时鼻黏膜细胞上TLR4表达开始增多,与正常对照组差异有统计学意义(P0.05),2h时鼻黏膜细胞上TLR4表达明显增加(P0.01),12h时鼻黏膜上TLR4表达开始下降(P0.05)。24h时鼻黏膜细胞上TLR4表达正常对照组差异无统计学意义(P0.05)。结论 1、TLR2和TLR4在变应性鼻炎组中均有表达。 2、TLR2和TLR4在变应性鼻炎的急性期出现高表达,在变应性鼻炎的整个病程中呈现动态变化。
[Abstract]:Purpose Toll - like receptor ( TLRs ) is the key receptor in human Toll - like receptor family . Toll - like receptor ( TLRs ) is one of the most important receptors in human Toll - like receptor family . Toll - like receptor ( TLRs ) is a key receptor in human Toll - like receptor family . Toll - like receptor ( TLRs ) is a key receptor in human Toll - like receptor . method 1 . Preparation of allergic rhinitis model Balb / c mice were injected intraperitoneally with 100 渭g of aluminum hydroxide ( Al ( OH ) 3 ) 2 mg dissolved in 0.1 ml of physiological saline , once every 1 , 3 , 5 , 7 , 9 , 11 , 13 , 15 days , and then stimulated with 10 渭g of egg albumin nasal drops on each side of the nasal cavity on day 16 , and the allergic rhinitis model was established for 11 days . 2 . Evaluation of allergic rhinitis model ( 1 ) observing 30 minutes after the last nose - nose excitation , observing the symptoms and scoring of the mice , scratching , nose , face , rhinorrhea and the like ; ( 2 ) extracting the sensitized animal blood , adding the allergen of the labeled fluorescent biotin in the serum , and then measuring the optical density value at 450 nm to determine the titer of the IgE in the serum ; and ( 3 ) performing routine HE staining in each group of mouse models , and counting the eosinophils in the parallel high - magnification mirror . 3 . The expression of Toll - like receptor 2 , 4 in allergic rhinitis was studied by immunohistochemical staining . 4 . The statistical results were expressed by mean 卤 standard deviation , and one Way - ANOVA was used to analyze the multiple groups . The statistical analysis was carried out with SPSS 13.0 software . Results 1 . The model of mouse allergic rhinitis was successfully prepared . 2 . There was significant difference between the symptom scores of allergic rhinitis group and the normal control group ( P0.01 ) , the IgE titer of allergic rhinitis group was significantly different from that of the normal control group ( P0.01 ) , and the count of eosinophils in allergic rhinitis group was significantly different from that of the normal control group ( P0.01 ) . 3 . Immunohistochemical method showed that TLR2 and TLR2 were not positive or weakly positive staining in the cytoplasm and nuclei of normal mouse nasal mucosa cells , and the nuclei and / or cytoplasm of nasal mucosa cells of allergic rhinitis group were stained with brown or yellowish brown positive . 4 . The expression of TLR2 in nasal mucosa increased significantly after 30 min in allergic rhinitis group ( P0.05 ) . The expression of TLR2 in nasal mucosa increased significantly ( P0.05 ) at 2h . The expression of TLR2 in nasal mucosa was not statistically significant at 24h ( P0.05 ) . 5 . In allergic rhinitis group , the expression began to increase in nasal mucosa cells at 30 min . There was a significant difference between nasal mucosa and normal control group ( P0.05 ) , and the expression began to decrease in nasal mucosa at 12 h ( P0.05 ) . Conclusion 1 . TLR2 and TLR2 were expressed in allergic rhinitis group . 2 . TLR2 and TLR2 were highly expressed in the acute stage of allergic rhinitis and showed dynamic changes in the whole course of allergic rhinitis .

【学位授予单位】：中国医科大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R765.21

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