【作者】 谢冰斌；

【导师】 刘月辉；

【作者基本信息】 南昌大学 ， 耳鼻咽喉—头颈外科， 2008， 硕士

【摘要】 目的:通过建立变应性鼻炎鼠类实验动物模型,探讨吸入性皮质类固醇激素对变应性鼻炎鼻黏膜重塑的影响。方法:选健康SD大鼠300只,随机分成正常对照组A和实验组,A组60只,实验组240只。首先对实验组大鼠予以卵清白蛋白进行基础致敏、致敏加强、局部激发建立变应性鼻炎动物模型,A组正常对照组则以等量生理盐水代替进行腹腔注射及滴鼻。然后将实验组动物随机分为B、C、D、E四组,每组60只。分组后,对A组继续予以等量生理盐水每周两次滴鼻;B组不再予以任何刺激及治疗;C组不再予以OVA刺激,每天予以丙酸氟替卡松每侧鼻腔50μl/次喷鼻; D组继续予以等浓度等量OVA双侧鼻腔滴鼻,每周两次;E组继续予以等浓度等量OVA双侧鼻腔滴鼻每周两次的同时,每天予以丙酸氟替卡松每侧鼻腔50μl/次喷鼻。分别于治疗后第1、2、4、8、12、及16周末各从A、B、C、D、E组中随机挑选10只进行处死。从各组的10只大鼠中随机挑选一只行鼻黏膜微血管铸型,其余9只取鼻黏膜分别送病理检查、PCR及透射电镜检查,以观察鼻黏膜形态学改变、微血管铸型情况及TGF-β1、FGF-2表达情况。结果:实验组大鼠动物模型建立成功,均表现出明显的抓鼻、喷嚏及流涕等症状。在接受变应原激发后鼻黏膜发生重塑改变,光镜下见鼻黏膜出现上皮剥脱,但尚未到基底层;并存在上皮杯状细胞化生,黏膜固有层出现腺体及血管增多,腺管增粗,炎症细胞浸润明显;电镜示上皮层纤毛不同程度脱落,排列欠整齐,基底膜板层网状结构增厚,可见胶原沉积及纤维增生,血管周围不同程度炎性细胞浸润,细胞明显水肿,组织间隙水肿也明显,但细胞间的结构尚未破坏;黏微血管明显增多,迂曲,分叉,粗细不一,微血管密度增加,并交织成网状;TGF-β1、FGF-2免疫阳性表达明显增高,TGF-β1mRNA表达升高。B组由于避免了变应原的接触,至第八周时,开始出现黏膜上皮剥脱减轻,至第十六周时黏膜上皮损伤情况才减轻较明显,炎症细胞浸润减少。TGF-β1、FGF-2免疫阳性表达及TGF-β1mRNA表达在第八周时开始下降。予以激素治疗后,C组在第八周上皮剥脱程度减轻,炎症细胞浸润减少,微血管相对减少,在第十六周鼻黏膜上皮层基本完整,腺样化生减轻,炎症细胞浸润不明显。而TGF-β1、FGF-2免疫阳性表达及TGF-β1mRNA表达在第四周时开始下降,至第十六周时已下降接近正常对照组水平。D组由于持续变应原接触,重塑进一步加重,TGF-β1、FGF-2免疫阳性表达及TGF-β1mRNA表达持续高水平。E组形态学改变无明显变化,至第十六周时仍可见到上皮杯状细胞化生明显,并可见大量黏液样物沉积于其中,黏膜下血管增粗。而TGF-β1、FGF-2免疫阳性表达及TGF-β1mRNA表达在第八周时开始下降,至第十六周时与C组比较无统计学差异。结论:1以卵清白蛋白为变应原,可通过大SD大鼠成功建立稳定的变应性鼻炎实验动物模型。2变应性鼻炎鼻黏膜可发生黏膜重塑现象,在避免变应原的持续接触后,其症状可缓解,鼻黏膜的重塑可自行减轻;如果变应原持续接触,其重塑将进一步加重并呈现不可逆性改变。3在能够避免变应原接触的前提下,皮质类固醇激素可较好的控制变应性鼻炎的症状,并在一定程度逆转重塑,起到一定的治疗作用;而在变应原持续接触的情况下,皮质类固醇激素虽可控制症状,但并不能在短期内完全逆转鼻黏膜的重塑,对已经发生了不可逆性改变的黏膜结构并无逆转或明显修复作用。4变应性鼻炎鼻黏膜中TGF-β1、FGF-2等生长因子可出现高表达,而皮质类固醇激素可以下调TGF-β1、FGF-2等生长因子的表达水平,这可能是皮质类固醇激素治疗变应性鼻炎机理之一。5对于变应性鼻炎的治疗,避免变应原的接触仍是首选方法,辅以皮质类固醇激素治疗可起到较好的疗效。更多还原

【Abstract】 Objective: To investigate the influence of the inhaled corticosteroids to the remodeling of the nasal mucosa in the allergic rhinitis, by establishing the allergic rhinitis model of the rats, which induced by OVA.Methods: 300 Sprague-Dawley(SD) rats are selected and randomly divided into two groups: the normal Group A used for control and the experimental group. The number of the rats in Group A is 60 and the experiment group is 240.First of all, make the rats in experimental group be allergic with ovalbumin(OVA) and enhance it and do local stimulation in order to establish the animal model of allergic rhinitis. While do intra-peritoneal injection with physiological saline in the same volume as that of ovalbumin(OVA) in the experimental group and intranasal dropping on the rats in normal Group A. Next, the rats in experimental group are randomly divided into four groups, i.e. Group B, Group C, Group D, Group E. The number of rats in each group is 60.Randomly select 10 rats to kill from each groups respectively at the end of the first, second, fourth, eighth, twelfth and sixteenth week after treatment. Randomly select one from that ten rats in each group for micro-vascular casting of nasal mucosa, and the remaining nine are to get pathological examination, PCR and transmission electron microscope in order that I can observe the morphological changes, the micro-vascular casts and the expression of TGF-β1,FGF-2 in the nasal mucosa. And then, the rats in Group A get the intranasal dropping with physiologic saline in the same volume twice a week; rats in Group B are under no stimulation or treatment; rats in the Group C are no longer stimulated with OVA, but they are going to have fluticasone propionate(FP) nasal spray, each side 50μl /per day; rats in Group D still get the intranasal dropping on each side with OVA in the same volume and concentration twice a week; rats in Group E also get the intranasal dropping on each side with OVA in the same volume and concentration twice a week, moreover, they get FP nasal spray, each side 50μl /per day.Results: The model on the rats in experimental group is established successfully: all the rats in the experimental groups show obvious symptoms: grasping noses, sneezing and running nose. After allergen stimulation, the nasal mucosa showed remodeling changes such as epithelial denudation, but it does not occur to the bottom and inflammatory cells infiltration becomes obvious with the help of optical microscope. And there are metaplasia of the goblet cells in epithelia and the number of gland and blood vessels increase in the mucosal layer and glandular vessels become thick. Cilia of epithelial shed to different extent and are uneven under the electron microscope, and the layers of reticular formation of basal membrane become thick, and collagen deposition and fabric hyperplasia are seen. There is inflammatory cells infiltration of different degrees around the blood vessels and edema in cells. Edemas in tissue spaces are also obvious and the structures between cells are not destroyed. And the micro-vascular in mucosa increase apparently. They are tortuous, branching, thick and thin. Density of blood capillary is increased and blood capillary becomes netted. The positive expression of immunohistochemistry of TGF-β1, FGF-2 rise obviously, in the meantime, TGF-β1 mRNA expression rises as well. Because of avoiding the exposure of allergen, epithelial denudations of mucosa in Group B are reducing from the eighth week. Till the sixteenth week, epithelial damage of the mucosa and inflammatory cells infiltration reduced obviously. TGF-β1、FGF-2 positive expression of immunity and TGF-β1m RNA expression drop in the eighth week. Under the treatment with corticosteroids, the degree of epithelial denudations and inflammatory cells infiltration of the rats in Group C are reduced, and micro-vascular are lessened in the eighth week. In the sixteenth week, epithelia of nasal mucosa are basically complete, glandular metaplasia alleviates, inflammatory cells infiltration become unobvious. The positive expression of immunohistochemistry of TGF-β1, FGF-2 and TGF-β1 mRNA expression decline in the fourth week, however, in the sixteenth week they draw close to the normal groups’. The positive expression of immunohistochemistry of TGF-β1, FGF-2 and TGF-β1 mRNA expression last high in Group D due to the continuously contact with allergen and enhancing remodeling. There are no obvious changes in morphology of the rats in Group E. Even in the sixteenth week, metaplasias of the goblet cells in epithelia are clear to be seen. Besides, a lot of mucus deposits in goblet cell, micro-vascular under the mucosa become thick. The positive expression of immunohistochemistry of TGF-β1, FGF-2 and TGF-β1 mRNA expression decline in the eighth week. Compared with Group C there is no differences in statistics in the sixteenth week.Conclusion: 1. The animal model of the allergic rhinitis can be established on the SD rats using the ovalbumin as the allergen. 2. The remodeling of the nasal mucosa can be found in allergic rhinitis. If allergen can be avoided, the symptoms can be alleviated and the remodeling will reduce by itself; if the allergen are continuously contacted, the remodeling will aggravate. 3. Corticosteroids can control the symptoms of the allergic rhinitis better and reverse the remodeling to some extent and perform the therapeutical effects. While if allergen can not be avoided, although the symptoms can be controlled with corticosteroids somehow, it is impossible to reverse the remodeling completely in a short time. And it cannot retro-converse or repair the nasal mucosa while the irreversibile change has occurred. 4. Growth factors such as TGF-β1, FGF-2 express obviously in the mucosa in allergic rhinitis, and the expression could be lowed down by corticosteroids, maybe that’s one of the mechanisms of the corticosteroids to cure the AR. 5. Avoid allergen is still the preferred approach for the treatment of the allergic rhinitis. If we can make sure to avoid allergen, the therapy supplemented by corticosteroid is one of good choice.更多还原