Role of LAIR-1 and CD300 Family Receptors in Regulating Inflammation

LAIR-1 和 CD300 家族受体在调节炎症中的作用

基本信息

批准号：
7964542
负责人：
JOHN COLIGAN
金额：
$ 62.07万
依托单位：
NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7964542
关键词：
Affect Agreement Allergic Animals Antigens Avidity B-Lymphocytes Binding CD4 Positive T Lymphocytes CD8B1 gene Cells Chemical Models Collagen Collagen Type I Collagen Type XVII Cytoplasmic Tail Defect Dendritic Cells Family Family member Gene Cluster Goals Hematopoietic stem cells Human IRp60 ITIM IgG1 Immune Immune response Immunoglobulin Class Switching Immunologic Receptors In Vitro Inflammation Ligand Binding Ligands Ligation Light Lymphocyte Lymphoid Modeling Mus Mutation Myeloid Cells Natural Killer Cells Organ Peptides Peripheral Peritoneum Peritonitis Proliferating Proteins Publishing Research Personnel Role Signal Transduction Site Surface Plasmon Resonance T-Lymphocyte cell type crosslink cytotoxicity eosinophil extracellular in vivo macrophage mast cell matrigel member monocyte neutrophil receptor research study response trafficking

项目摘要

We have generated LAIR-1 -/- mice on a B6 background to study the in vivo function of LAIR-1. Phenotypic analysis of lymphoid organs did not show large differences between LAIR-1 +/+ and LAIR-1 -/- animals. We have observed a slight increase in the percentage of splenic B cells in the LAIR-1 -/- mice, along with a decrease in T cells, mostly because of a decrease in CD8 T cells. This probably does not result from abnormal trafficking of T lymphocytes, since LAIR-1 +/+ and LAIR-1 -/- T cells traffic equally to peripheral lymphoid organs. In the gut we have observed a slight increase of T cells in the LAIR-1 -/- mice, along with an increase in the NKG2D expression. In vitro experiments showed that OT-II LAIR-1-/- CD4 T cells proliferated less than the OT-II LAIR-1+/+ CD4 T cells when they are cultured with APC loaded with OT-II OVA peptide. To study the immune response in vivo, we have immunized the animals with TNP-OVA and found that class switching is affected in LAIR-1 -/- mice. These animals produced lower levels of IgG2a and IgG2b, while switching to IgG1 is not affected. By using T cell specific LAIR-1 -/- animals (CD4 Cre LAIR-1flox/flox), we confirmed that the defect in class switching is T cell specific. Previous investigators have published that mouse B cells do not express LAIR-1; however, recently we have found that marginal zone B cells are positive for LAIR-1 expression. Other preliminary results have shown that in the LAIR-1 -/- mice there are significant alterations in the recruitment of macrophages and eosinophils into the peritoneum in a model of chemical peritonitis, suggesting a role for LAIR-1 in the trafficking of these cell types towards sites of inflammation. We investigated the effect of a conservative R65K mutation on LAIR-1 ligand binding and function. Compared with LAIR-1 wild-type (wt)-expressing cells, LAIR-1 R65K cells show markedly reduced binding to collagen, which correlates with a reduced level of LAIR-1 polarization to the site of interaction with collagens. Both LAIR-1 wt and R65K cells can generate intracellular signals when ligated by anti-LAIR-1 mAb, but only LAIR-1 wt cells respond to collagens or matrigel. In agreement, surface plasmon resonance analyses showed that LAIR-1 R65K protein has markedly reduced avidity for collagen type I compared with LAIR-1 wt. Likewise, LAIR-1 R65K protein has decreased avidity for cells expressing transmembrane collagen XVII. Thus, a single residue, Arg65, is critical for the interaction of LAIR-1 with collagens.

我们已经在B6背景上产生了LAIR-1 - / - 小鼠，以研究Lair-1的体内功能。淋巴器官的表型分析并未显示Lair-1 +/ +和Lair-1 - / - 动物之间的巨大差异。我们已经观察到Lair-1 - / - 小鼠中脾B细胞百分比的百分比略有增加，以及T细胞的降低，主要是因为CD8 T细胞的降低。这可能不是由T淋巴细胞的异常运输引起的，因为Lair-1 +/ +和Lair-1 - / - T细胞同样流动到外周淋巴机构。在肠道中，我们观察到Lair-1/ - 小鼠中T细胞的略有增加，以及NKG2D表达的增加。体外实验表明，OT-II LAIR-1 - / - CD4 T细胞在用载有OT-II OVA肽的APC培养时，比OT-II Lair-1+/+ CD4 T细胞少于OT-II LAIR-1+/+ CD4 T细胞。为了研究体内的免疫反应，我们已经用TNP-ova免疫了动物，发现类切换在lair-1 - / - 小鼠中受到影响。这些动物产生了较低的IgG2a和IgG2B，而切换到IgG1的动物不受影响。通过使用T细胞特异性Lair-1-/ - 动物（CD4 CRE LAIR-1FLOX/FLOX），我们证实了类切换中的缺陷是T细胞特异性的。先前的研究人员已经发表了小鼠B细胞不表达巢穴-1。但是，最近我们发现边缘区B细胞对LAIR-1表达呈阳性。其他初步结果表明，在Lair-1 - / - 小鼠中，在化学腹膜炎模型中，巨噬细胞和嗜酸性粒细胞募集到腹膜中存在重大变化，这表明LAIR-1在将这些细胞类型贩运到炎症部位的运输中起作用。我们研究了保守的R65K突变对LAIR-1配体结合和功能的影响。与Lair-1野生型（WT）表达细胞相比，LAIR-1 R65K细胞显示出与胶原蛋白的结合显着降低，这与Lair-1降低水平与与胶原蛋白相互作用的位点相关。当通过抗LAIR-1 MAB结扎时，LAIR-1 WT和R65K细胞都可以产生细胞内信号，但只有Lair-1 WT细胞对胶原蛋白或Matrigel响应。一致，表面等离子体共振分析表明，与LAIR-1 WT相比，LAIR-1 R65K蛋白已显着降低了I型胶原蛋白的自发性。同样，lair-1 r65k蛋白也减少了表达跨膜胶原蛋白XVII的细胞。因此，单个残基Arg65对于Lair-1与胶原蛋白的相互作用至关重要。