Complement receptor 2-negative (CR2/CD21

Protein tyrosine phosphatase nonreceptor type 22 (PTPN22) gene polymorphisms are associated with many autoimmune diseases. The major risk allele encodes an R620W amino acid change that alters B cell receptor (BCR) signaling involved in the regulation of central B cell tolerance. To assess whether this PTPN22 risk allele affects the removal of developing autoreactive B cells, we tested by ELISA the reactivity of recombinant antibodies isolated from single B cells from asymptomatic healthy individuals carrying one or two PTPN22 risk allele(s) encoding the PTPN22 R620W variant. We found that new emigrant/transitional and mature naive B cells from carriers of this PTPN22 risk allele contained high frequencies of autoreactive clones compared with those from non-carriers, revealing defective central and peripheral B cell tolerance checkpoints. Hence, a single PTPN22 risk allele has a dominant effect on altering autoreactive B cell counterselection before any onset of autoimmunity. In addition, gene array experiments analyzing mature naive B cells displaying PTPN22 risk allele(s) revealed that the association strength of PTPN22 for autoimmunity may be due not only to the impaired removal of autoreactive B cells but also to the upregulation of genes such as CD40, TRAF1, and IRF5, which encode proteins that promote B cell activation and have been identified as susceptibility genes associated with autoimmune diseases. These data demonstrate that early B cell tolerance defects in autoimmunity can result from specific polymorphisms and precede the onset of disease.

Impaired immune functions leading to primary immunodeficiencies often correlate with paradoxical autoimmune complications; patients with hyper-IgM syndromes who are deficient in activationinduced cytidine deaminase (AID), which is required for classswitch recombination and somatic hypermutation, are prone to develop autoimmune diseases. To investigate the impact of AIDdeficiency on early B-cell tolerance checkpoints in humans, we tested by ELISA the reactivity of recombinant antibodies isolated from single B cells from AID-deficient patients. New emigrant/ transitional and mature naive B cells from AID-deficient patients express an abnormal Ig repertoire and high frequencies of autoreactive antibodies, demonstrating that AID is required for the establishment of both central and peripheral B-cell tolerance. In addition, B-cell tolerance was further breached in AID-deficient patients as illustrated by the detection of anti-nuclear IgM antibodies in the serum of all patients. Thus, we identified a major and previously unsuspected role for AID in the removal of developing autoreactive B cells in humans.

We studied the effect of inhaled specific antigen on airflow resistance, histamine concentration, and mast cell numbers in airway tissues in allergic dogs. In each of six allergic dogs with open chests, inhalation of specific antigen aerosol (Ascaris suum) increased airflow resistance from 0.30+/-0.40 (mean+/-SE) to 2.8+/-0.41 cmH2O/1 per s (P less than 0.05); decreased mast cell number from 38.6+/-3.2 to 24.3+/-4.5 mast cells/mm2 (P less than 0.05); decreased histamine (per mg airway tissue) from 5.3+/-0.4 to 3.5+/-0.2 ng (P less than 0.05) in lobar bronchi 5-7 mm in diam; and released histamine into the blood perfusing the lung, control arterial plasma histamine; 2.8+/-0.64 ng/ml; antigen: 154+/-1.7 ng/ml (P less than 0.005). Specific antigen caused no significant changes in mast cells or histamine in bronchi 1-2 mm in diam. Control aerosols of nonspecific antigen or methacholine did not change levels of mast cells or histamine in airway tissues. These results suggest that experimental canine asthma involves local airway reactions with degranulation of mast cells and histamine release, as well as vagally mediated reflex bronchoconstriction.

B cells play a central role in the pathogenesis of many autoimmune diseases. Therefore, understanding the mechanisms that regulate B-cell tolerance in humans is important for the development of new therapeutic strategies. Patients with monogenic diseases provide rare opportunities to study the impact of specific gene mutations on the regulation of human B cell tolerance. By this, we could show that alterations in B-cell receptor and Toll-like receptor (TLR) signaling pathways result in defective central B-cell tolerance.To further dissect the signaling pathways involved in the establishment of central B-cell tolerance in humans, we tested by ELISA and immunofluorescence the reactivity of recombinant antibodies cloned from single transitional B cells from individuals carrying CD19 mutations. CD19 is a co-receptor expressed on B cells and is involved in the amplification of B-cell responses.We found that individuals carrying CD19 mutations displayed defective central B-cell tolerance checkpoints. In addition, CD19-deficient transitional B cells were enriched in anti-nuclear clones, a feature previously observed in IRAK4-and MYD88-deficient patients in which TLR7/9 sensing nucleic acids cannot signal. Therefore, we investigated the functions of these TLRs in B cells in the absence of CD19 expression. CD19-deficient human B cells displayed defective up-regulation of activation markers after TLR7/9 triggering and failed to induce BTK, AKT but not p38 MAPK or I-Bα phosphorylation after TLR7/9 stimulation. Additionally, inhibitors blocking BTK, AKT and PI3K function impaired CD19-dependent TLR7/9 responses in healthy donor's B cells. Finally, we demonstrated that individuals carrying BTK mutations display similar defects in TLR7/9-induced B-cell activation and central B-cell tolerance.Hence, we identified a previously unsuspected role for CD19 molecules in regulating TLR7/9 functions in human B cells and central B-cell tolerance to nuclear antigens.