FEASIBILITY STUDY--ANTIGEN PRESENTATION IN HLA B27 TRANSGENIC ANIMALS

可行性研究--HLA B27转基因动物中的抗原呈递

• 批准号: 6235858
• 负责人: ROBERT A. COLBERT
• 金额: $ 10.8万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-06-01 至 1998-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6235858
• 关键词: HY antigen; antigen presentation; autoantigens; binding proteins; biological polymorphism; cytotoxic T lymphocyte; flow cytometry; genetically modified animals; histocompatibility antigens; immunogenetics; immunoprecipitation; laboratory mouse; laboratory rat; major histocompatibility complex; minor histocompatibility loci; peptide structure; polymerase chain reaction; protein structure function

HLA-B*2703, a subtype of HLA-B27 that has not been associated with susceptibility to spondyloarthropathies, differs from all other major histocompatibility complex (MHC) class I molecules including other B27 subtypes at position 59 in the A pocket. Alloreactive cytotoxic T lymphocytes (CTL) directed against the most common B27 subtype, B*2705, show partial recognition of B*2703, while the vast majority of anti-B*2703 CTL clones recognize B*2705. Furthermore, B*2703 is deficient in the binding and presentation of a B*2705-restricted influenza A nucleoprotein peptide (SRYWAIRTR). Binding and presentation are restored by substituting Arg for the naturally-occurring Ser at position 1 (P1) of the peptide, indicating the importance of this amino acid in maintaining high affinity peptide binding to B*2703. These and other preliminary results suggest that B*2703 may bind and present only a subset of those peptides presented by B*2705. The consequences of this unique polymorphism at position 59, on both the physiologic function of B27 in protective immunity, as well aas the pathogenic role this molecule may play in susceptibility to spondyloarthropathies, are unknown. We are engaged in several lines of investigation to determine the extent of peptide binding and presentation differences between these two closely related HLA-B27 subtypes. In this Developmental and Feasibility project application we propose to use B*2703/humanbeta2-microglobulin (hbeta2m) transgenic mice that we are in the process of producing, along with currently existing B*2705/hbeta2m animals, to determine the functional significance of these differences. First, we will address the hypothesis that B*2703 presents only a subset of B*2705-restricted self peptides by performing reciprocal skin grafts and assessing tissue rejection. If our hypothesis in correct, B*2703/hbeta2m mice will reject B*2705/hbeta2m tissue, while B*2705 animals will accept B*2703 tissue. Alloreactive cytotoxic T lymphocyte responses accompanying graft rejection will be evaluated. Second, we will examine B*2705 and B*2703-restricted responses to known viral or self peptide epitopes introduced by immunization with DNA vectors encoding these peptides. We will vary the P1 amino acid to determine its role in influencing in vivo peptide presentation by these HLA-B27 subtypes, and how this affects immune responsiveness. Third, we will produce B*2703 transgenic rats to determine whether this subtype can induce the spontaneous inflammatory disease seen in rats transgenic for B*2705. These studies in transgenic animals will help to define the role of a unique A pocket polymorphism in peptide presentation by MHC class I molecules, and may provide the basis for understanding differential susceptibility to spondyloarthropathies conferred by two closely related HLA-B27 subtypes.

人类白细胞抗原B*2703，人类白细胞抗原B27的一个亚型，尚未与 对脊椎关节病的易感性，与所有其他主要疾病不同 组织相容性复合体(MHC)I类分子，包括其他B27 A口袋中第59位的亚型。同种异体反应性细胞毒性T 淋巴细胞(CTL)针对最常见的B27亚型B*2705， 显示部分认可B*2703，而绝大多数反B*2703 CTL克隆识别B*2705。此外，B*2703在 B*2705限制性甲型流感核蛋白的结合和呈递 多肽(SRYWAIRTR)。通过替换以下内容来恢复绑定和显示 Arg用于肽的第1(P1)位上自然产生的Ser， 表明这种氨基酸在维持高亲和力方面的重要性 与B*2703结合的多肽。这些和其他初步结果表明 B*2703可以仅结合和呈现所呈现的那些多肽的子集 B*2705。59位的这种独特的多态的结果是 B27在保护性免疫中的生理作用以及AAS 该分子在易感性中可能起的致病作用 脊椎关节病，是未知的。 我们正在进行几项调查，以确定 两者在多肽结合和呈递方式上的密切差异 相关的人类白细胞抗原B27亚型。在这个开发性和可行性项目中 我们建议使用B*2703/人β2-微球蛋白(Hbeta2m) 我们正在生产的转基因小鼠，以及 目前已有B*2705/hbeta2M动物，以确定其功能 这些差异的意义。首先，我们将讨论这一假设 B*2703仅呈现B*2705限制性自体多肽的子集 进行相互皮肤移植和评估组织排斥反应。如果我们的 假设正确，B*2703/hbeta2m小鼠将拒绝B*2705/hbeta2m 组织，而B*2705动物将接受B*2703组织。同种异体反应 伴随移植排斥反应的细胞毒性T淋巴细胞反应将是 已评估。其次，我们将检查B*2705和B*2703-受限响应 通过DNA免疫引入的已知病毒或自身多肽表位 编码这些多肽的载体。我们会将P1氨基酸改变为 通过这些确定其在影响体内多肽递呈中的作用 人类白细胞抗原-B27亚型，以及这如何影响免疫反应性。第三，我们 将产生B*2703转基因大鼠，以确定该亚型是否可以 诱导自发性炎症性疾病的转基因大鼠 B*2705。 这些对转基因动物的研究将有助于确定一种 MHC I类分子在多肽呈递中独特的A口袋多态性 分子，并可能为理解差异提供基础 两个密切相关的人对脊柱关节病的易感性 人类白细胞抗原B27亚型。