Specific direct bradykinin (BK) binding and competitive inhibition was detected in human neutrophil and peripheral blood mononudear celt (PBMC) detergent solubilized extracts and purified plasma membranes using in vitro radioreceptor ligand binding. Scatchard analyses of [~2sI]-BK binding revealed an equilibrium dissociation constant (K a ) of 2.9x10 ~' M for neutrophils and 5.6x10 ~ M for PBMC using [des-argg]-BK a B1 agonist; 2.6x10 ~a M for neutrophils, 6.2x10 ~' M for PBMC with BK a B2 agonist; 5.4x10 ~ M for PBMC using Lys-BK a B2 agonist. The number of binding sites (B) was calculated to be 0.113 fM/p.g protein (720 receptors per cell) for neutrophils and 0.200 fM/p.g protein (1289 receptors per ceil) for PBMC with the B1 agonist while with the B2 agonists the values were 0.128 fM/~g protein (818 receptors per cell) for neutrophils and 0.157 fM/~tg protein (1005 receptors per cell) for PBMC with BK, and 0.293 fM/~tg protein (1870 receptors per cell) with Lys-BK for PBMC.In a competitive binding inhibition assay using neutrophil and PBMC glycerol purified plasma membranes, high affinity binding in the nanomolar range was detected to Lys-BK and BK but with [des-argg]-BK a 10-100 fold lower order affinity was observed this being indicative of pharmacologically defined B2 characteristics.

Summary Our aim was to determine whether heterogeneity ofthe IgE (Ce) gene could be demonstrated in patients with chronic urticaria (CU). We performed Southern blots on DNA extracted from peripheral blood leucocytes of 20 patients with CU and 20 normal controls. Using a human Ce gene probe containing four exons ofthe constant segment ofthe IgE heavy chain, we showed the presence of a restriction fragment length polymorphism of the Ce gene segment in four of 20 patients with CU. but in none of 20 normal subjects. Family studies of two propositi revealed the presence of this Ce gene polymorphism in other family members. Our data show that a proportion of patients with CU have a polymorphism ofthe constant segment ofthe Ce gene. Further studies of this polymorphic gene fragment indicated that it was derived from duplication ofthe 3rd and 4th exons of functional Ce gene and was very likely to be located close to this gene. It raises the possibility that polymorphism ofthe functional Ce gene may affect expression of this gene. This could possibly lead to dysfunctional IgE-receptor interaction with consequent alteration in mediator release.

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The non-integrin 37/67-kDa laminin receptor (LAMR1) is a complex protein with diverse functions. LAMR1 is widely expressed in epithelial cells and recently it was reported on neutrophils and a subset of activated T cells. Ligation of LAMR1 on peripheral blood mononuclear cells (PBMC) downregulated LPS-induced TNFα production, suggesting immune functions. However, its expression on primary monocytes remain unknown. Interestingly, LAMR1 mRNA is downregulated in PBMC of patients with early rheumatoid arthritis (RA), and low gene expression is an independent predictor of poor response to anti-TNFα treatment, suggesting a role in RA pathogenesis. We found LAMR1 was constitutively expressed on all peripheral blood monocytes and a subset of B cells from healthy individuals and patients with RA and it was abundantly present in synovial tissue of patients with RA. On monocytes and synovial tissue lower levels of LAMR1 expression tended to correlate with increased disease activity scores. In vitro treatment of monocytes with IFNγ or IL-10 up-regulated surface LAMR1 in healthy individuals and patients with RA with greater effects observed in healthy individuals. Importantly, treatment with IFNγ significantly increased specific binding of monocytes to laminin-1. TNFα and IL-1β caused marginal downregulation of LAMR1 in patients but effects in controls were variable. Taken together, constitutively expressed LAMR1 on monocytes is differentially regulated by pro-inflammatory and immune-regulatory cytokines suggesting LAMR1 may regulate the threshold and amplitude of their activation and migration. Decreased levels in patients with RA may indicate loss of this potentially critical homeostatic regulation thereby contributing to the excessive inflammation.