A novel heterophilic antibody interaction involves IgG4.

IgG4 has been implicated in a diverse set of complex pathologies - e.g. autoimmune pancreatitis (AIP), idiopathic membranous nephropathy - and carries unique features including lack of activation of the classical complement pathway and a dynamic Fab-arm exchange. We recently showed that the rheumatoid factor (RF)-like activity of IgG4 is achieved through a hitherto unknown, Fc-Fc (and not Fab-Fc as is the case in classical RF; CRF) interaction; hence the name, novel RF (NRF). Here, we further explore the resemblance/difference between CRF and NRF. As heterophilic interactions of human IgM RF (CRF) are well known, we checked whether this is the case for IgG4. Human IgG4 showed variable reactivity to animal IgGs: reacting intensely with rabbit and mouse IgGs, but weakly with others. The binding to rabbit IgG was not through the Fab (as in CRF) but via the Fc piece, as was recently shown for human IgG (NRF). This binding correlates with the IgG4 concentration per se and could therefore be of diagnostic usage and incidentally explain some observed interferences in biological assays. In conclusion, here is defined a novel heterophilic antibody interaction and is established the universality of the unique Fc-Fc binding, both involving IgG4.

Possible role of peripheral CD14low monocytes in the development of collagen-induced arthritis in cynomolgus monkeys (Macaca fascicularis).

The changes in levels of peripheral major lymphocyte subsets were monitored with 10 adult cynomolgus monkeys (5 females and 5 males) during the 9 weeks after immunization with chick type-II collagen in Freund's complete adjuvant. Three females and 3 males developed overt arthritis determined by swelling of small joints and increase of plasma alkaline phosphatase as well as C-reactive protein. An increase of CD16+ NK cells was observed in four non-arthritis-developed monkeys (two females and two males). There was no significant difference in the fluctuation pattern of CD4+ T cell, CD8+ T cell and CD20+ B cell levels between arthritis-developed monkeys and non-developed ones. In addition, the percentages of CD45RA+ CD4+ T cells to total CD4+ T cells, CD28- CD8+ T cells to total CD8+ T cells, and IgD- B cells to total B cells did not significantly differ between them. On the other hand, a significant increase was demonstrated in CD14-positive cells at 3 weeks after immunization in only arthritis-developed monkeys regardless of sex. The expression of CD14 antigen on the surface of increased cells was low in comparison with those appearing in blood obtained before immunization. In addition, increased CD14low cells showed no response to LPS stimulation. However, there was no significant difference in antibody titer to both chick type-II and monkey type-II collagen between arthritis-developed monkeys and non-developed ones. These results suggest that an increase in number of CD14low monocytes with immature function might be a part of the autoimmune response, and that the appearance of these cells is of pathogenic importance in the arthritic process in cynomolgus monkeys regardless of the production of autoantibody.

Collagen-induced arthritis in nonhuman primates: multiple epitopes of type II collagen can induce autoimmune-mediated arthritis in outbred cynomolgus monkeys.

OBJECTIVE: To define which regions of the type II collagen (CII) molecule result in anticollagen antibody production and the subsequent development of autoantibodies in a collagen-induced arthritis (CIA) nonhuman primate model. METHODS: Male and female cynomolgus monkeys (2-6 of each sex per group) were immunized with either chicken (Ch), human, or monkey (Mk) CII, or with cyanogen bromide (CB)-generated peptide fragments of ChCII emulsified in Freund's complete adjuvant. Monkeys were observed for the development of arthritis, and sera were collected and analyzed for anticollagen antibody specificity by enzyme-linked immunosorbent assay. RESULTS: Overt arthritis developed in all groups of monkeys immunized with intact CII and with all major CB peptide fragments of ChCII except CB8. Onset and severity of arthritis correlated best with serum anti-MkCII antibody levels. The levels of IgG autoantibody to MkCII were a result of the cross-reactivity rate of anti-heterologous CII antibodies with MkCII, which was based on the genetic background of individual monkeys rather than on sex differences. CONCLUSION: CII from several species and disparate regions of the CII molecule were able to induce autoantibody-mediated arthritis in outbred cynomolgus monkeys. The strong anti-MkCII response suggests that epitope spreading or induction of broad-based CII cross-reactivity occurred in these animals. Autoantibody levels to MkCII were higher in CIA-susceptible monkeys than in resistant monkeys, despite comparable antibody levels in response to the various immunizations of CII. These results closely parallel the type of anticollagen responses found in sera from rheumatoid arthritis patients. Perhaps this can be accounted for by similar major histocompatibility complex heterogenicity associated with an outbred population, or maybe this is a primate-specific pattern of reactivity to CII.

Specificity of antibodies to type II collagen in rheumatoid arthritis.

To reassess the role of autoantibodies to type II collagen in the pathogenesis of diseases, we studied antibodies from patients with rheumatoid arthritis (RA) and from patients with relapsing polychondritis for species specificity and collagen type specificity, using an improved enzyme-linked immunosorbent assay. Antibodies were found in the sera of 15% of the RA patients and 50% of the relapsing polychondritis patients, as well as in the cartilage of 69% of the RA patients examined. Reaction with both homologous and heterologous type II collagens was common. Analysis of 19 selected RA sera revealed that autoantibodies were generally associated with specific antibodies to some species of heterologous type II collagen. In contrast, antibodies found in 4% of the non-RA controls were specific for either bovine or chick type II collagen. These findings indicate that autoantibody formation in RA and relapsing polychondritis may occur as a result of an immune response to heterologous type II collagen. However, since RA and relapsing polychondritis patient sera differed in their reactivity with the cyanogen bromide-digested peptides, it is possible that the clinical manifestation of collagen autoimmunity might be influenced by the epitope specificity of the antibodies.

An improved enzyme-linked immunosorbent assay of anti-collagen antibodies in human serum.

An improved enzyme-linked immunosorbent assay (ELISA) for the determination of anti-collagen antibodies in human serum has been developed. The method is based on the use of serum samples diluted to 1/50 with heat-inactivated normal rabbit serum adjusted to pH 8.0 with solid Tris (0.05 M), NaCl (0.15 M) and 2 M HCl. The use of normal rabbit serum minimizes non-specific adsorption of immunoglobulin G onto the plastic surface of microtiter plate. The applicability of the method for the quantitation of anti-collagen antibodies in human serum is demonstrated with 290 specimens of sera from normal controls (194) and patients with rheumatoid arthritis (96).

Sex-linked differences in susceptibility of cynomolgus monkeys to type II collagen-induced arthritis. Evidence that epitope-specific immune suppression is involved in the regulation of type II collagen autoantibody formation.

To determine whether sex-linked factors are important in the susceptibility and resistance of cynomolgus monkeys to type II collagen-induced arthritis, we immunized 5 males and 10 females with chick type II collagen (CII) and studied their clinical and immune responses. All 10 females developed overt arthritis and produced antibodies to CII and cross-reactive antibodies to monkey type II collagen (MkII). In contrast, only 1 male monkey developed arthritis, which was transient and mild in severity. Examination of antisera obtained from the male monkeys disclosed high titers of antibody to CII, but little antibody to MkII. The cyanogen bromide peptide recognition patterns varied markedly from animal to animal, implying that monkeys are capable of recognizing multiple arthritogenic determinants on CII.