Development of an HLA-DP2 Transgenic Murine Model of Chronic Beryllium Disease

慢性铍病 HLA-DP2 转基因小鼠模型的建立

基本信息

批准号：
8389617
负责人：
Andrew P. Fontenot
金额：
$ 17.1万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-01-01 至 2014-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8389617
关键词：
Alleles Alveolitis Amino Acids Animal Disease Models Animal Model Animals Antigens Beryllium Binding Binding Sites Blood Breathing C3H/HeJ Mouse C57BL/6 Mouse CD4 Positive T Lymphocytes Cells Charge Chronic berylliosis Data Development Disease Disease susceptibility Exposure to FVB/N Mouse Fibrosis Generations Genetic Predisposition to Disease Glutamic Acid Granulomatous HLA-DP Antigens HLA-DP2 HLA-DP4 HLA-DPB1 gene Hilar Histocompatibility Antigens Class II Human Immune System Diseases Immune response Inbred Strains Mice Individual Inflammation Inflammatory Response Knowledge Lead Link Lung Lung diseases Lymphocyte Lysine Mediating Metals Modeling Mononuclear Mus Nature Oxides Pathology Patients Positioning Attribute Public Health Publishing Research Personnel Risk Sampling Site Spleen Structure T-Cell Activation T-Cell Receptor T-Lymphocyte Testing Threonine Time Transgenes Transgenic Mice Transgenic Organisms Workplace animal model development base cytokine human disease human subject interstitial lymph nodes lymphocyte proliferation mouse model response

项目摘要

DESCRIPTION (provided by applicant): Chronic beryllium disease (CBD) is a granulomatous lung disorder caused by beryllium exposure in the workplace and is characterized by the accumulation of beryllium-specific CD4+ T cells in the lung. Disease susceptibility has been linked to HLA-DPB1 alleles, including DPB1*0201, that possess a glutamic acid residue at position 69 of the HLA-DP2 2-chain (2Glu69). We have shown that beryllium directly binds to HLA-DP2 molecules and that the majority of beryllium-specific CD4+ T cells recognize beryllium in the context of HLA- DP. Importantly, the DP molecules that present beryllium match those implicated in disease susceptibility, confirming that the HLA contribution to disease is based on the ability of those molecules to bind and present beryllium to T cells. We have recently crystallized HLA-DP2 and identified a potential beryllium binding site involving ¿Glu69 and two other invariant glutamic acids at positions ¿26 and ¿68. Functional studies confirmed that beryllium recognition was dependent on this cluster of negative charges. To date, no animal model of beryllium-induced disease exists. Previous attempts to generate a murine model failed to document a beryllium-specific adaptive immune response. Importantly, mice lack a major histocompatibility complex class II (MHCII) molecule of similar sequence and structure to HLA-DP2. Thus, we hypothesize that the generation of a humanized transgenic mouse containing the cluster of negative charges unique to HLA-DP2 will result in a disease-specific murine model with pathology reflective of the human disease. HLA-DP2 and -DP4 transgenic mice on an FVB/N background have been generated by researchers at NIOSH. However, these mice express murine MHC and lack human CD4, an important coreceptor for optimal T cell activation. No studies characterizing the response of these mice to inhaled beryllium have been published. Preliminary data show peribronchovascular mononuclear cell infiltrates occurring only in beryllium-exposed HLA-DP2 transgenic mice after beryllium oxide (BeO) exposure, strongly suggesting that the inflammatory response is due to the presence of the HLA-DP2 transgene and supporting our hypothesis that ¿Glu69-containing HLA-DP molecules are required for the generation of a murine model of beryllium-induced disease. In the first specific aim, we will re-derive the HLA-DP2 transgene in C57BL/6 mice lacking murine MHCII molecules and expressing human CD4. In the second aim, we will directly compare the beryllium-induced adaptive immune response in these animals and DP2-expressing FVB/N mice, testing the hypothesis that stabilization of the interaction between HLA-DP2 and T cell receptor with the human CD4 coreceptor and elimination of the confounding effects of mouse MHCII will optimize the beryllium-specific immune response. Together, these studies will lead to a disease-specific murine model, targeting the exact molecule necessary for the development of a human disease.

描述（由适用提供）：慢性铍疾病（CBD）是工作场所中的铍暴露引起的肉芽肿性肺疾病，其特征是肺中芽孢特异性CD4+ T细胞积累。疾病的易感性已与HLA-DPB1等位基因（包括DPB1*0201）有关，该等位基因在HLA-DP2 2链（2GLU69）的位置上具有谷氨酸保留率（2GLU69）。我们已经表明，腹部直接与HLA-DP2分子结合，并且大多数腹部特异性CD4+ T细胞在HLA-DP的背景下识别腹膜。重要的是，呈腹部腹膜的DP分子与疾病敏感性实现的DP分子相匹配，证实HLA对疾病的贡献基于这些分子结合并将腹膜呈现给T细胞的能力。我们最近已经结晶了HLA-DP2，并确定了涉及的潜在腹膜结合位点»glu69和另外两个不变的谷氨酸在位置»26和68。功能研究证实，腹膜识别取决于这一负电荷的群集。迄今为止，尚未存在腹膜诱导疾病的动物模型。以前的生成鼠模型的尝试未能记录腹膜特异性适应性免疫响应。重要的是，小鼠缺乏与HLA-DP2相似的序列和结构的主要组织相容性复合物II类（MHCII）分子。这就是我们假设，含有HLA-DP2独有的负电荷簇的人源性转基因小鼠的产生将导致具有反映人类疾病的病理学的疾病特异性鼠模型。 NIOSH的研究人员生成了FVB/N背景上的HLA -DP2和-DP4转基因小鼠。然而，这些小鼠表达鼠MHC，缺乏人类CD4，这是最佳T细胞激活的重要共核能。尚未发表表征这些小鼠对继承腹膜反应的反应的研究。初步数据表明，在暴露于硫甘蓝的HLA-DP2转基因小鼠（BEO）暴露后仅发生的纤维隆举止单核细胞浸润，强烈表明炎症反应是由于HLA-dp2透射的存在，并支持了我们的摩尔摩尔，并支持了我们的摩尔生成型Hla69-contaken-contains-contage hla，腹膜诱导疾病的鼠模型。在第一个特定目的中，我们将重新启用缺乏鼠MHCII分子和表达人CD4的C57BL/6小鼠中的HLA-DP2变换。在第二个目的中，我们将直接比较这些动物和表达DP2表达FVB/N小鼠的腹膜诱导的自适应免疫响应，从而检验了一个假说，即HLA-DP2和T细胞受体之间的相互作用稳定在人体CD4 coceptor和CDECECTIUID-SPERTOR之间的相互作用稳定在鼠类MHCII IMMHCII IMMHCICICIFIFE的效应中。总之，这些研究将导致特异性鼠模型，以针对人类疾病发展所需的确切分子。