We show that LPS-stimulated circulating CD14-positive monocytes from patients with common variable immunodeficiency (CVID) express a higher proportion of intracellular IL-12-positive cells than monocytes from patients with X-linked agammaglobulinemia or normal subjects. We used four-color flow cytometry and measured IL-12 with an Ab to the p40 subunit following stimulation with LPS. The raised IL-12 is associated with an increased frequency of IFN-γ-positive T cells, but not of IFN-γ-positive CD56+ NK cells. These increases in frequency of cytokine-positive cells are due to a decrease in the absolute numbers of circulating monocytes and T cells that are negative for IL-12 and IFN-γ, respectively. The increased frequency of IL-12-positive monocytes appears to be selective because TNF-α was not increased, and is thus unlikely to reflect a general activation. Chronic infection is also unlikely to explain our data since cells from X-linked agammaglobulinemia patients with a similar Ig deficiency do not show these changes. Our data suggest a fundamental abnormality in the IL-12/IFN-γ circuit in CVID, with up-regulation of IL-12 being the “primary” factor. This imbalance is likely to skew the immune response away from Ab production and also explains the failure of CVID T cells to make Ag-specific memory cells and the chronic inflammatory and granulomatous complications that are a feature of CVID. This disease appears to be a rare example of a polarized Th1-type response and may in part be due to a genetic defect in the control of IL-12 production.

Factor J (FJ) is a complement inhibitor that acts on the classical and the alternative pathways. We demonstrated FJ-cell interactions in fluid phase by flow cytometry experiments using the cell lines Jurkat, K562, JY, and peripheral blood lymphocytes. FJ bound to plastic plates was able to induce in vitro adhesion of these cells with potency equivalent to fibronectin. As evidence for the specificity of this reaction, the adhesion was blocked by MAJ2, an anti-FJ monoclonal antibody, and by soluble FJ. Attachment of the cells required active metabolism and cytoskeletal integrity. The glycosaminoglycans heparin, heparan sulfate, or chondroitin sulfates A, B, and C inhibited to varying degrees the binding of FJ to cells, as did treatment with chondroitinase ABC. In the search for a putative receptor, a protein of 110 kDa was isolated by affinity chromatography, and microsequence analysis identified this protein as nucleolin. Confocal microscopy evidenced the presence of nucleolin in cell membrane by immunofluorescence with monoclonal (D3) and polyclonal anti-nucleolin antibodies in Jurkat cells. The interaction FJ-nucleolin was evidenced by Western blot and enzyme-linked immunosorbent assay. Furthermore, purified nucleolin and D3 inhibited adhesion of Jurkat cells to immobilized FJ, suggesting that the interaction was specific and that nucleolin mediated the binding. Factor J (FJ)1 is a complement inhibitor that is able to regulate both the classical and the alternative pathways of complement (1, 2). FJ was initially found as a soluble molecule in urine and serum (1, 3). FJ is a peculiar protein having a high sugar content and a pI Ն 9.6 (4). In addition, the existence of FJ-related molecules has been described on the surface of human circulating cells and lymphoid cell lines (5). Flow cytometry analysis with an anti-FJ monoclonal antibody (mAb) has revealed staining in a small but consistent population of lymphocytes (mean: 11%) but not in monocytes, granulocytes, erythrocytes, or platelets. Moreover, we have also found that anti-FJ stained several cell lines (Jurkat, U937, K562, and Ramos) at varying percentages.Relationships between complement and adhesion processes have been described previously; moreover, complement regulators such as vitronectin (VN) (6), clusterin (7), and more recently, factor H-like protein 1 (8) and factor H (9) have been implicated in adhesion processes. Affinity purification identified Mac1 (CD11b/CD18) as a factor H binding receptor (9). Moreover, CD11b/CD18 (Mac1) binds several soluble ligands including the complement fragment iC3b (10). Some characteristics of FJ resemble VN. Among them are the ability to regulate complement, the tendency to form aggregates, and the potential to bind heparin (11). VN is a ligand for ␤ 1 and ␤ 3 integrins. Clusterin, a different complement regulator, is capable of promoting the aggregation and adhesion of renal epithelial cells.FJ also strongly binds heparin (12). Heparin-binding proteins constitute a diverse group that includes extracellular matrix mo...

We examined the effects of intravenous immunoglobulin (IVIG) on cytokine regulation in vivo using samples taken before and after replacement-dose (200-400 mg/kg) IVIG in a group of patients with common variable immunodeficiency (CVID) and X-linked agammaglobulinaemia (XLA). The intracellular cytokine content of CD4+ and CD8+ lymphocytes, and their CD28+/- subsets, were measured following in vitro activation with phorbol myristate acetate (PMA) and ionomycin. The cytokines IL-2, interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha), and the early activation marker CD69, were assessed by four-colour flow cytometry of whole blood cultures taken before and after IVIG infusion. There was a significant increase in IL-2 expression in CD4+ (and CD4+28-) cells and an increase in TNF-alpha expression in CD8+28- cells following IVIG in CVID, but not in XLA patients. IFN-gamma and CD69 expression were not affected by IVIG infusion. This increase in TNF-alpha and IL-2, combined with unchanged IFN-gamma expression, is evidence against the putative 'anti-inflammatory' role of IVIG, and may explain the failure of resolution of granulomata in CVID patients treated with IVIG alone.

No abstract