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研究生： 蔡明翰
Ming-Han Tsai
論文名稱： 探討自體免疫傾向之介白素15受器alpha鍵基因剔除小鼠之中心及周邊免疫耐受性
Study of Central and Peripheral Tolerance in the Autoimmune Prone Interleukin-15 Receptor α Chain Gene Knockout mice
指導教授： 廖南詩
Nan-Shih Liao
張子文
Tse-Wen Chang
潘榮隆
Rong-Long Pan
口試委員:
學位類別： 碩士
Master
系所名稱： 生命科學暨醫學院 - 生物科技研究所
Biotechnology
論文出版年： 2006
畢業學年度： 94
語文別： 英文
論文頁數： 47
中文關鍵詞： 介白素15 、介白素2 、介白素15受器 、自體免疫 、胸線負選擇 、紅斑狼瘡 、自體抗體 、胸線細胞 、CD4輔助T細胞
外文關鍵詞： Interlerkin-15, Interleukin-2, Interleukin-15R, Autoimmunity, Thymocyte negative selection, Lupus, Autoantibodies, Thymocytes, CD4+ T cells
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    • 介白素15號受器(interleukin-15 receptor; IL-15R)是由α、β和γ鍵所組成。其中IL-15受器α鍵本身對IL-15具有高度的親和力，然而β和γ鍵受器則是對IL-15和介白素2號(Interleukin-2; IL-2)皆具有中度的親和力。之前的研究指出在IL-2受器α鍵，IL-2/IL-15受器β鍵，或IL-2之基因剔除之小鼠會產生早期且嚴重性的自體免疫反應，然而，這些症狀卻未曾在IL-15受器α鍵基因缺損之小鼠上發現。我們之前已在IL-15受器α鍵基因剔除，且年齡大於六個月之成年母鼠發現類紅斑性狼瘡症狀，卻不曾發現於野生型的對照組。在這篇論文裡，我證實了IL-15受器α鍵基因剔除的小鼠在胸線及周邊組織表現較差的免疫耐受性。這些結果可能提供了構成成年IL-15受器α鍵基因缺損小鼠產生自體免疫疾病的一些條件，因為其較差的免疫耐受性會造成自體反應的T細胞累積，進而造成自體免疫疾病。

    Interleukin-15 receptor (IL-15R) contains α, β and γ chains. The α chain by itself binds to IL-15 with high affinity, while β and γ chains are intermediate affinity receptors for both Interleukin-2 (IL-2) and IL-15. Previous studies showed that mice deficient of IL-2 receptor α (IL-2Rα), IL-2/15 receptor β (IL-2/15Rβ) or IL-2 developed early progressing severe autoimmune diseases, a phenotype not observed in IL-15Rα knockout (IL-15Rα-/-) mice. We found that the Systemic Lupus Erythematosus (SLE)-like symptoms in female IL-15Rα-/- aged female mice older than 6 months but not in wild type (WT) counterpart. In this thesis, I demonstrated that IL-15Rα-/- mice exhibited inefficient tolerance in the thymus and in the periphery. These results may provide the mechanisms underlying the progressing autoimmune diseases in aged IL-15Rα-/- mice, which is the accumulation of autoimmune T cells resulted from inefficient immune tolerance.

    封面 中文摘要 Abstract 誌謝 Introduction Interleukin-15 (IL-15) IL-15Rα and in trans presentation Role of IL-2 and IL-15 systems in autoimmune diseases Central tolerance and autoimmune diseases Peripheral tolerance and autoimmune diseases Specific aims Materials and Methods Mice Euthanasia of mice Antibodies and Immunostaining procedures Preparation of signal-cell suspensions from lymphoid organs Negative selection models in vivo Dexamethasone sensitivity test in vitro and Fetus thymus organ culture (FTOC) Splenic Antigen presentation cells (APC) purification Negative selection models in vitro FTOC for T cells development and selection In vivo IL-15 treatment in anti-TCRβ induced negative selection Oral tolerance assay Bone marrow transplantation Statistical Analysis and Graphs Medium Results Inefficient negative selection of thymocytes in IL-15Rα-/- mice in vivo 1. Anti-TCR mAb-induced thymocyte deletion 2. Thymocytes or thymus organs of WT and IL-15Rα-/- mice had similar sensitivity to DEX 3. Endogenous superantigen-induced negative selection 4. HY TCR+ transgenic (HY) mice model Negative selection of thymocytes in vitro 1. Negative selection of thymocytes by anti-TCR mAb immobilized to plate or APC 2. Negative selection of thymocytes by anti-TCR mAb in FTOC IL-15 augmented anti-TCR induced thymocytes deletion in IL-15Rα bearing environment 1. Exogenous IL-15 can not increase the thymocytes deletion by anti-TCR mAb immobilized to plate 2. Exogenous IL-15 upgraded anti-TCR mAb-induced thymocyte deletion when cultured together with IL-15Rα-expressing splenic APC and FTOC Reduction of DX5-TCR-CD122+ thymocytes in IL-15Rα-/- mice IL-15 enhanced anti-TCRβ mAb-induced negative selection in vivo Insufficient peripheral tolerance mechanism in IL-15Rα-/- mice Discussions Discussion of experimental models for studying negative selection in vivo The inefficient negative selection in IL-15Rα-/- mice in unmanipulated mice Why can not inefficient negative selection in IL-15Rα-/- thymocytes be demonstrated in vitro? How IL-15 system parcitipated in negative selection in vivo How IL-15 modulated negative selection in thymocytes? Inefficient peripheral tolerance in IL-15Rα-/- mice provides more chance to autoimmunity Why the lupus-like autoimmunity only developed in IL-15Rα-/- female mice? Summary Figures FIGURE 1. Anti-TCRβ-mediated thymocyte deletion in vivo FIGURE 2. Anti-CD3ε mAb-mediated thymocyte deletion in vivo FIGURE 3. DEX mediated thymocyte deletion in isolated thymocytes and thymus organs FIGURE 4. IL-15Rα-/- mice had more Vβ5+ thymocytes than WT mice in the intermediate affinity I-Ab background FIGURE 5. Negative selection occurred normally in the IL-15Rα-/- HY mice FIGURE 6. Studies of the anti-TCR inducing thymocyte deletion between WT and IL-15Rα-/- thymocytes in vitro FIGURE 7. WT and IL-15Rα-/- FTOC showed similar in TCR mediated thymocyte deletion FIGURE 8. Exogenous IL-15 or IL-2 can not upgrade the TCR mediated-thymocytes deletion of isolated thymocytes FIGURE 9. Exogenous IL-15 augmented thymocytes deletion cytokine in the presence of splenic APC culture and in FTOC FIGURE 10. IL-15Rα-/- mice had less DX5-gdTCR-CD122+ thymocytes FIGURE 11. Exogenous IL-15 injection enhanced anti-TCR□ induced thymocyte deletion in vivo FIGURE 12. Inefficient oral tolerance in IL-15Rα-/- mice References Supplementary Data Supplementary Data A. Using bone marrow chimera mice to study the cell roles in anti-TCRβ mAb-induced negative selection

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