塵蹣蛋白致敏小鼠短期攝取塵蹣蛋白產生口服耐受性的機制探討

Study on oral Tolerance Induced by Short-term Feeding of Dp2 Protein to Mice sensitized and Challenged with Dp2 Protein

許銘志(Ming-Chih Hsu)

指導教授 ： 林璧鳳

國立臺灣大學/生命科學院/微生物與生化學研究所/碩士(2007年)

https://doi.org/10.6342/NTU.2007.01691

摘要

塵蹣 ( Dermatophagoides pteronyssinus ) 是引起國人過敏性氣喘的主要過敏原，約80% ~ 90% 塵蹣過敏患者對塵蹣蛋白第二類蛋白質 ( Dp2 ) 產生過敏反應。當經口服食多量過敏原而引起免疫不反應性稱為口服耐受性，由本研究室先前結果已知，連續七天餵食塵蹣蛋白有效誘發口服耐受性，減緩過敏性氣喘模式小鼠的過敏免疫反應，且微陣列基因分析顯示小腸細胞 CRIP3 和 CD226 基因表現顯著增加。因此，本實驗欲探討口服耐受性的影響機制，分析短期攝取塵蹣蛋白對後續的腹腔注射致敏和支氣管刺激時與免疫調節相關基因的影響。結果顯示，連續七天口服過敏原 Dp2 後，小腸免疫組織皮耶氏體淋巴結的 CRIP3、CD226、Foxp3之 mRNA 表現量顯著增加、脾臟細胞 IFN-g 分泌量顯著上升，顯示免疫調節傾向第一型免疫反應。在後續的致敏中 CD226 雖反而下降，但是調節型 T 細胞的轉錄因數 Foxp3 mRNA 的表現量在致敏過程中仍持續增高，因而脾臟與 Peyer’s patch 表現 CD4+CD25+ 的調節型 T 細胞也顯著上升，顯示餵食大量過敏原經由腸道免疫產生較多調節型免疫細胞，可影響脾臟免疫細胞的分化與調節。因此，後續的致敏過程中脾臟細胞 IL-4 和 IL-5 皆顯著下降，調節型細胞激素 TGF-b1 也隨致敏漸增，且比氣喘組顯著較高。口服耐受組在腹腔致敏時使脾臟細胞 Dp2 特異性增生反應顯著下降，而血清總 IgE 抗體也顯著降低，後續支氣管 Dp2 刺激下仍顯著低於過敏氣喘組，顯示口服耐受性顯著抑制過敏原特異性的增生和減少總過敏抗體的生成。當進行支氣管刺激時，口服耐受組肺部呼吸道阻力 ( airway hyperresponsiveness ) 也顯著下降，肺沖洗液的 IL-4 和 IL-13 皆有顯著下降，IL-5 與 eotaxin 也有下降趨勢，顯示過敏免疫反應受到抑制。同時，血清 Dp2 特異性 IgE 抗體也有下降趨勢。因此，本研究結果顯示，短期餵食塵蹣蛋白可改變 CRIP3、CD226 和 Foxp3 的基因表現，而口服耐受組在致敏過程中仍有較高的 Foxp3 表現，並誘發調節型 T 細胞產生，抑制過敏免疫反應，使短期的餵食可達到長期免疫調節的效果。

關鍵字

塵蹣 ； 口服耐受性 ； 調節型 T 細胞 ； 過敏氣喘

並列摘要

The dust mite is a major allergen for allergic asthma, an airway chronic inflammatory disease. About 80 ~ 90% of patients who react to mite extract react specifically to dust mite protein group 2, Dp2. Oral tolerance is a state of immunological unresponsiveness to allergen induced by feeding. Our previous study demonstrated that oral tolerance was induced efficiently by continuous feeding of Dp2 for seven days. Therefore, to investigate mechanisms of oral tolerance used in asthma, the mRNA expression and immune response in the process of oral tolerance induction, three trials including a short-term oral administration of Dp2, an added three intra-peritoneal (IP) boosting and an extra intra-tracheal (IT) challenge, were conducted. In the short-term feeding trial, the tolerance group significantly enhanced mRNA expressions of CRIP3, CD226 and Foxp3 in Peyer’s patch (PP) and Dp2-stimulated IFN-g secretion from splenocytes, indicating a Th1 dominant immunomodulation by oral induction. Although CD226 became lower in oral tolerance group after IP boosting and IT challenge, Foxp3, a transcriptional factor in regulatory T cell, remained significantly higher during the sensitization process. The CD4+CD25+ level, a regulatory T cell marker, in splenocytes and PP were also significantly higher suggesting that regulatory immune cells were induced and differentiation of splenocytes might be affected by oral tolerance. Therefore, IL-4 and IL-5 secretion were suppressed, and TGF-b1 secretion was increased in oral tolerance group during the following sensitization. The Dp2-stimulated proliferation of splenocyte and serum total IgE level decreased significantly in oral tolerance group after IP boosting. Airway hypersensitivity, Th2 cytokine such as IL-4 and IL-13 in BALF, both serum total IgE and Dp2-specific IgE levels significantly decreased in the tolerance group after IT challenge. In summary, short-term oral administration of Dp2 altered long term intestinal gene expression such as Foxp3, and induced more regulatory T cells and suppressed allergic immune responses to achieve the long-term tolerance outcome.

並列關鍵字

Dp2 ； oral tolerance ； regulatory T cell ； allergic asthma

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