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Induction of factor VIII specific unresponsiveness by intrathymic factor VIII injection in murine hemophilia A

胸腺内注射因子 VIII 诱导小鼠血友病 A 诱导因子 VIII 特异性无反应

基本信息

批准号：
17591006
负责人：
MADOIWA Seiji
金额：
$ 2.24万
依托单位：
Jichi Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-17591006/
关键词：
hemophilia A factor VIII inhibitor immune tolerance thymus regulatory T cells high resolution image system 胸腺 SIVベクター抗第VIII因子インヒビター樹状細胞

项目摘要

Hemophilia A is a congenital bleeding disorder caused by a deficiency of coagulation factor VIII. Approximately 30% of hemophilia A patients develop inhibitors against factor VIII following replacement therapy. We have previously reported that neonatal exposure of factor VIII antigen might induce antigen specific immune tolerance by IFN-γ dependent T cell anergy in hemophilic mice (Madoiwa S, et al. J Thromb Haemost 2:754-762,2004). Thymus plays crucial roles of self-tolerance with negative selection of self-reactive effector T cells and positive selection of self-reactive regulatory T cells. In this study, we focused on the induction of immune tolerance by direct thymic injection of factor VIII. Hemophilia mice were injected recombinant human factor VIII into thymus under the real time high resolution image guidance (Vevo 770, Visual Sonic Inc.). We measured the factor VIII inhibitors after repeated intravenous injection of factor VIII. The CD4+ cells, antigen presenting cells (APCs) and CD4+CD25+ cells were isolated for the proliferation and cytokine assays under in vitro stimulation of factor VIII. Anti-factor VIII inhibitory antibody titers were significantly lower in mice (n=17) with thymic injection of factor VIII than in mice (n=18) without injection (14.6 ± 3.8 vs 184.5 ± 48.6 Bethesda units/mL, respectively, p=0.0019). The CD4+ cells from thymic injected mice could not proliferate or produce IL-2, IL-12, and IFN-γ in response to factor VIII, when they were co-cultured with APCs from immunogenic mice. The CD4+CD25+ cells from thymic treated mice but not from naive mice efficiently suppressed the proliferative response of immunogenic mice derived CD4+ cells in the presence of APCs. Intrathymic administration of factor VIII could result in immune tolerance by factor VIII specific regulatory T cells induction. This immune tolerance model may provide a basis of new manipulation for the prevention of factor VIII inhibitors.

A型血友病是一种由凝血因子VIII缺乏引起的先天性出血性疾病。大约30%的A型血友病患者在替代治疗后出现抗因子VIII抑制剂。我们之前报道过，新生儿暴露于因子VIII抗原可能通过血友病小鼠IFN-γ依赖性T细胞能量诱导抗原特异性免疫耐受（Madoiwa S等）。[J] .血栓病杂志，2004(2):754-762。胸腺通过阴性选择自身反应效应T细胞和阳性选择自身反应调节性T细胞，在自身耐受中起着至关重要的作用。在本研究中，我们的重点是通过胸腺直接注射因子VIII诱导免疫耐受。在实时高分辨率图像引导（Vevo 770, Visual Sonic Inc.）下，将重组人因子VIII注射到血友病小鼠胸腺。反复静脉注射因子VIII后测定因子VIII抑制因子。分离CD4+细胞、抗原呈递细胞（APCs）和CD4+CD25+细胞，在体外因子VIII刺激下进行增殖和细胞因子测定。胸腺注射VIII因子小鼠（n=17）的抗VIII因子抑制抗体滴度显著低于未注射小鼠（n=18）（分别为14.6±3.8比184.5±48.6 Bethesda单位/mL， p=0.0019）。当与免疫原性小鼠的apc共培养时，胸腺注射小鼠的CD4+细胞不能增殖或产生IL-2、IL-12和IFN-γ，以响应因子VIII。来自胸腺治疗小鼠的CD4+CD25+细胞，而不是来自幼稚小鼠的CD4+细胞，在apc存在的情况下，有效地抑制了免疫原性小鼠来源的CD4+细胞的增殖反应。胸腺内给药因子VIII可通过诱导因子VIII特异性调节性T细胞产生免疫耐受。这种免疫耐受模型可能为预防因子VIII抑制剂的新操作提供基础。