Objective. Human leukocyte-associated immunoglobulin-like receptor 1 (hLAIR-1) is an immune inhibitory receptor for collagen that is expressed on most immune cells. We previously showed that the LAIR-1-collagen interaction could be antagonized by the secreted homolog hLAIR-2, which can be detected in the synovial fluid of rheumatoid arthritis (RA) patients. In addition, the extracellular part of hLAIR-1 is a putative antagonist upon shedding from the cell membrane. The purpose of this study was to determine the relative roles of hLAIR-2 and soluble hLAIR-1 (shLAIR-1) in the regulation of the LAIR-1-collagen interaction.Methods. The ability of recombinant LAIR proteins to abrogate LAIR-1-collagen binding was tested by flow cytometry and adhesion assays. Collagen binding capacity was analyzed by surface plasmon resonance. Plasma, urine, and synovial fluid were screened for the presence of sLAIR-1 and LAIR-2 by enzyme-linked immunosorbent assay.Results. Recombinant LAIR-2 proteins abrogated the binding of collagen to LAIR-1 more efficiently than did recombinant sLAIR-1. Consistent with these findings, surface plasmon resonance analysis showed that LAIR-2 had a higher affinity for collagen than did LAIR-1. Activated CD4؉ T cells were the main producers of LAIR-2, whereas the source of sLAIR-1 remains elusive. Both soluble LAIR-1 and LAIR-2 could be detected in the plasma and urine of healthy control subjects and patients with RA. Urinary levels of both proteins were significantly increased in RA patients, and LAIR-2 levels in urine were significantly correlated with markers of inflammation. Conclusion. Our data suggest that LAIR-2 is a more potent antagonist of LAIR-1 function in vivo, while both sLAIR-1 and LAIR-2 are potential biomarkers that may be used to monitor urine samples for evidence of systemic inflammation.Leukocyte-associated immunoglobulin-like receptor 1 (LAIR-1) is an inhibitory immune receptor for collagens and is expressed on most immune cells (1-3). Upon triggering, LAIR-1 inhibits various cellular functions (2,4-8). Since receptor and ligand are widely expressed, the regulation of the interaction must be tightly controlled to allow for appropriate cellular activation. We and other investigators have previously shown that upon maturation or activation, LAIR-1 expression on neutrophils, T cells, and B cells decreases, providing a regulatory mechanism at the level of receptor expression (8-11).

The collagenous C-type lectin, SP-D, is a multitrimeric glycoprotein present at mucosal surfaces and is involved in host defense against infections in mammals. SP-D has immunomodulatory properties, but the underlying mechanisms are incompletely understood. SP-D contains collagen domains. LAIR-1 is an inhibitory immune receptor at the cell surface of various immune-competent cells that binds collagen. We hypothesized that the immunomodulatory functions of SP-D can be mediated via interactions between its collagen domain and LAIR-1. Binding assays show that SP-D interacts via its collagenous domain with LAIR-1 and the related LAIR-2. This does not affect the mannan-binding capacities of SP-D, which induces cross-linking of LAIR-1 in a cellular reporter assay. Functional assays show that SP-D inhibits the production of FcαR-mediated reactive oxygen via LAIR-1. Our studies indicate that SP-D is a functional ligand of the immune inhibitory receptor LAIR-1. Thus, we have identified a novel pathway for the immunomodulatory functions of SP-D mediated via binding of its collagenous domains to LAIR-1. This may provide a mechanism for the unexplained immunomodulatory function of the collagenous domains of SP-D.

Activation of complement may cause severe tissue damage in antibody-mediated allograft rejection and other antibody-mediated clinical conditions; therefore, novel potent complement inhibitors are needed. Previously, we described binding of the inhibitory receptor LAIR-1 and its soluble family member LAIR-2 to collagen. Here, we investigated binding of LAIR-1 and LAIR-2 to the complement proteins C1q and MBL, which both have collagen-like domains, and evaluated the effect of this binding on complement function. We demonstrate specific binding of recombinant LAIR proteins to both C1q and MBL. Surface plasmon resonance experiments showed that LAIR-2-Fc protein bound C1q and MBL with the highest affinity compared to LAIR-2-HIS. We, therefore, hypothesized that LAIR-2-Fc is a potent complement inhibitor. Indeed, LAIR-2-Fc inhibited C4 fixation to IgG or mannan, reduced activation of C4 by aggregated IgG in plasma and inhibited iC3b deposition on cells. Finally, LAIR-2-Fc inhibited complement-mediated lysis of cells sensitized with anti-HLA antibodies in an ex vivo model for antibody-mediated transplant rejection. Thus, LAIR-2-Fc is an effective novel complement inhibitor for the treatment and prevention of antibody-mediated allograft rejection and antibody-mediated clinical conditions.

No abstract

The soluble form of the inhibitory immune receptor leukocyte-Associated Ig-like Receptor-1 (sLAIR-1) is present in plasma, urine and synovial fluid and correlates to inflammation. We and others previously showed inflammatory protein expression in normal amniotic fluid at term. We hypothesized that sLAIR-1 is present in amniotic fluid during term parturition and is related to fetal lung function development. sLAIR-1 was detectable in all amniotic fluid samples (n=355) collected during term spontaneous deliveries. First, potential intra-uterine origins of amniotic fluid sLAIR-1 were explored. Although LAIR-1 was expressed on the surface of amniotic fluid neutrophils, LAIR-1 was not secreted upon ex vivo neutrophil stimulation with LPS, or PMA/ionomycin. Cord blood concentrations of sLAIR-1 were fourfold lower than and not related to amniotic fluid concentrations and placentas showed no or only sporadic LAIR-1 positive cells. Similarly, in post-mortem lung tissue of term neonates that died of non-pulmonary disorders LAIR-1 positive cells were absent or only sporadically present. In fetal urine samples, however, sLAIR-1 levels were even higher than in amniotic fluid and correlated with amniotic fluid sLAIR-1 concentrations. Second, the potential relevance of amniotic fluid sLAIR-1 was studied. sLAIR-1 concentrations had low correlation to amniotic fluid cytokines. We measured neonatal lung function in a convenient subset of 152 infants, using the single occlusion technique, at a median age of 34 days (IQR 30-39). The amniotic fluid concentration of sLAIR-1 was independently correlated to airway compliance (ρ=0.29, P=.001). Taken together, we show the consistent presence of sLAIR-1 in amniotic fluid, which originates from fetal urine. Concentrations of sLAIR-1 in amniotic fluid during term deliveries are independent from levels of other soluble immune mediators. The positive association between concentrations of amniotic fluid sLAIR-1 and neonatal lung compliance suggests that amniotic fluid sLAIR-1 may be useful as a novel independent marker of neonatal lung maturation.