Human Keratin (K14)-MBP/MBP-TCR Transgenic Animal Model

描述(由申请人提供)：胸腺在建立自我耐受性中起着核心作用。尽管在胸腺上皮细胞中观察到广泛的异质性，但从E15.5胚胎中纯化的MTS24+MHCII+CD45-TECs的异位移植表明，这种表型的细胞足以形成表型和功能正常的胸腺微环境，启动和维持正常的胸腺生成。这表明MTS24+胸腺上皮细胞包括一组胸腺内干细胞，能够产生功能齐全的皮质和髓质胸腺上皮。提示MTS24+胸腺上皮干细胞移植具有诱导自身反应性T细胞和同种异体T细胞免疫耐受的治疗潜力。CD4+CD25+Foxp3+调节性T细胞(Tregs)在多发性硬化症、类风湿性关节炎和1型糖尿病等自身免疫性疾病的抑制和预防中发挥重要作用。CD4+CD25+Foxp3+树突状细胞是在胸腺上皮细胞表达其同源抗原时产生的。重要的是，活化的蛋白脂蛋白特异性的CD4+CD25+调节性T细胞抑制其他髓鞘抗原如髓鞘少突胶质细胞糖蛋白和髓鞘碱性蛋白诱导的EAE的发生。糖尿病相关抗原特异性调节性T细胞也比多克隆的CD4+CD25+调节性T细胞更有效地抑制糖尿病的发展。这一证据表明，CD4+CD25+Foxp3+Tregs与自身免疫性疾病相关的自身抗原反应，可能通过旁观者抑制而在自身免疫性疾病中显示出有效的抑制作用。我们将开发一种新的策略，通过使用胸腺上皮干细胞来开发与自身免疫病相关的自身抗原特异性CD4+CD25+Foxp3+Tregs。我们推测，表达MBP转基因基因的胸腺上皮干细胞移植后，移植同基因骨髓来源的造血干细胞，可以促进重组免疫系统中具有较高抑制活性的MBP特异性CD4+CD25+Foxp3+Tregs的发育，并使实验性自身免疫性脑脊髓炎(EAE)易感小鼠对EAE产生抵抗。为了解决这一假设，我们提出了在胸腺上皮细胞中表达MBP基因的TG小鼠。我们将研究MBP特异性Foxp3+Tregs在MBP-TG小鼠中的发育。 髓鞘碱性蛋白(MBP)是一种自身抗原，参与实验性自身免疫性脑脊髓炎(EAE)的发生发展。我们将建立表达MBP的小鼠，并检测MBP基因在胸腺上皮细胞中的表达是否可以抑制EAE的发展。

DESCRIPTION (provided by applicant): The thymus plays a central role in the establishment of self-tolerance. Although broad heterogeneity is observed in thymic epithelial cells, heterotopic transplantation of purified MTS24+MHCII+CD45-TECs derived from E15.5 embryo showed that cells of this phenotype are sufficient to form a phenotypically and functionally normal thymic microenvironment that initiate and sustain regular thymopoiesis. This indicates that MTS24+ thymic epithelia cells encompass a population of intrathymic stem cells capable of giving rise to fully functional cortical and medullary thymic epithelium. This information suggests that grafting of MTS24+ thymic epithelial stem cells possesses therapeutic potentials to induce immune tolerance in autoreactive T cells and allogenic T cells. CD4+CD25+ Foxp3+regulatory T cells (Tregs) have been suggested to play a critical role in the suppression and prevention of autoimmune diseases including multiple sclerosis, rheumatoid arthritis and type 1 diabetes. CD4+CD25+Foxp3+ Tregs develop when their cognate antigens are expressed in the thymic epithelial cells. Importantly, activated proteolipid protein-specific CD4+CD25+ regulatory T cells suppress the development of EAE induced by other myelin antigens such as myelin oligodendrocyte glycoprotein and myelin basic protein. Diabetes-associated antigen-specific regulatory T cells also suppress the development of diabetes more efficiently than polyclonal CD4+CD25+ regulatory T cells. This evidence suggests that CD4+CD25+ Foxp3+Tregs reacting with autoimmune disease-associated autoantigens could exhibit efficient suppression in autoimmune diseases via bystander suppression. We will develop a novel strategy to develop autoimmune disease-associated autoantigen-specific CD4+CD25+ Foxp3+Tregs by using thymic epithelial stem cells. We hypothesize that grafting of thymic epithelial stem cells expressing the MBP transgene followed by transplantation of syngeneic bone marrow-derived hematopoieticstem cells could facilitate the development of MBP-specific CD4+CD25+ Foxp3+Tregs with higher suppressive activity in the reconstituted immune system and render Experimental Autoimmune Encephalomyelitis (EAE) -susceptible mice resistant to EAE. To address this hypothesis, we propose to generate Tg mice that express MBP gene in the thymic epithelial cells. We will investigate development of MBP-specific Foxp3+ Tregs in the MBP-Tg mice. Myelin basic protein (MBP) is an autoantigen involved in development of experimental autoimmune encephalomyelitis (EAE). We will generate the mice expressing the MBP and examine whether expression of MBP gene in the thymic epithelial cells can suppress the development of EAE.