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

为了研究LAIR-1在体内的功能，我们在B6背景下培育了LAIR-1 -/-小鼠。淋巴器官表型分析显示LAIR-1 +/+和LAIR-1 -/-动物间差异不大。我们观察到LAIR-1 -/-小鼠脾脏B细胞百分比略有增加，同时T细胞减少，主要是因为CD8 T细胞减少。这可能不是由于T淋巴细胞的异常运输，因为LAIR-1 +/+和LAIR-1 -/- T细胞同样运输到外周淋巴器官。在肠道中，我们观察到LAIR-1 -/-小鼠的T细胞略有增加，同时NKG2D表达增加。体外实验表明，载有OT-II卵肽的APC培养后，OT-II LAIR-1-/- CD4 T细胞增殖能力低于OT-II LAIR-1+/+ CD4 T细胞。为了研究体内的免疫反应，我们用TNP-OVA免疫动物，发现LAIR-1 -/-小鼠的类转换受到影响。这些动物产生较低水平的IgG2a和IgG2b，而转向IgG1不受影响。通过使用T细胞特异性LAIR-1 -/-动物（CD4 Cre LAIR-1flox/flox），我们证实了类转换缺陷是T细胞特异性的。先前的研究人员已经发表，小鼠B细胞不表达LAIR-1；然而，最近我们发现边缘区B细胞LAIR-1表达阳性。其他初步结果表明，在化学性腹膜炎模型中，LAIR-1 -/-小鼠腹膜中巨噬细胞和嗜酸性粒细胞的募集发生了显著变化，这表明LAIR-1在这些细胞类型向炎症部位的运输中发挥了作用。

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.