The autoantibodies that react with dopamine and serotonin are of interest in the study of bulimia nervosa. These neurotransmitters play an important role in appetite control, sexual and social behavior, and stress responses, all of which form a part of the clinical picture of bulimia nervosa. Are these autoantibodies involved in the serotoninergic hypofunctioning present in bulimia nervosa? Are they a part of an immunity regulation system essential for the cerebral system's homeostasis? To address these questions, 31 bulimic females (diagnosed according to DSM-III-R criteria) were compared with 10 control subjects (matched to the patients for sex, age, and demographic/psychosocial features). Measurement of the activity of natural autoantibodies reacting with dopamine, dopamine-beta-hydroxylase and serotonin was performed by an enzyme-linked immunosorbent assay (ELISA) for typical immunoglobulins (IgG, IgM, IgA). All of the autoantibodies of the IgG type were lower in the bulimic group than in the control group, a difference that was statistically significant for IgG anti-serotonin and IgG anti-dopamine. There was a trend for the amount of IgM anti-dopamine to be lower in patients than in controls. Dopamine and serotonin are specific components of brain cells. It can therefore be hypothesized that these antigens acting with autoantibodies could be the antigenic cerebral targets reacting with 'anti-brain' antibodies. The study of these specific autoantibodies provides information about the immunological characteristics that may be related to brain disturbances.

In the first part of this study, we investigated the rate of natural autoantibodies, in a sample of 31 female inpatients with bulimia nervosa according to DSM III-R criteria. The control (age and sex matched) group consisted in high school students including 10 females without eating disorders, depressive disorder or immunological disease. We investigated especially natural autoantibodies reacting with compounds of the central nervous system (Dopamine, Dopamine beta Hydroxylase, Serotonin). Our first conclusion is that there is a lower level of these natural auto-antibodies among female patients with bulimia nervosa. In the second part of the study, we have especially investigated the correlation between impulsivity in bulimia nervosa and the rate of natural autoantibodies against serotonin.

In the first part of this study, we investigated the rate of natural autoantibodies, in a sample of 31 female inpatients with bulimia nervosa according to DSM III-R criteria. The control (age and sex matched) group consisted in high school students including 10 females without eating disorders, depressive disorder or immunological disease. We investigated especially natural autoantibodies reacting with compounds of the central nervous system (Dopamine, Dopamine beta Hydroxylase, Serotonine). Our first conclusion is that there is a lower level of these natural auto-antibodies among female patients with bulimia nervosa. In the second part of the study, we have especially investigated the correlation between impulsivity in bulimia nervosa and the rate of natural autoantibodies against serotonine.

Brown-Norway (BN) rats injected with HgCl2 produce anti-laminin antibodies responsible for an autoimmune glomerulonephritis. The properties of three IgG1 monoclonal antibodies (mAb) previously obtained in this model, and of immunoglobulins eluted from kidneys of diseased rats, were compared in the present study. Two mAb (Hg15 and Hg16) recognized laminin only, while the third one (Hg17) was polyreactive, as were some of the kidney-eluted immunoglobulins; they reacted with laminin and with several other antigens including 2,4,6-trinitrophenyl (TNP). The Hg17 mAb and kidney-eluted polyreactive antibodies were affinity purified using a TNP-bovine serum albumin (BSA) column; their affinity for TNP was high (2 x 10(-8)M, and 1 x 10(-8)M, respectively) but less than that of a TNP-specific (LO-DNP-2) mAb (2 x 10(-11) M). The Hg17 mAb and kidney-eluted antibodies reacted more effectively with TNP28-BSA than with TNP8.5-BSA, while the TNP-specific mAb reacted equally well with both conjugates. The Hg17 mAb was the most cationic (pI: 7) of the anti-laminin mAb and this was even more evident when F(ab')2 fragments were studied (pI: 8.2). The polyreactive kidney-eluted immunoglobulins that bound TNP were also more cationic (pI: 7.4-9.3) than the fraction that did not recognize TNP (pI: 5.8-8.6). The anti-laminin mAb bound in vivo to the glomerular basement membrane, but only the Hg17 mAb could be eluted with DNP alone. This study shows that polyreactive anti-laminin antibodies are produced during this autoimmune disease, and indicates that they may have pathogenic potential.

Using a panel of self antigens, IgM autoreactivities were clearly and constantly detected by enzyme immunoassay (EIA) in the sera of 29 normal human individuals. Similarly, IgM autoreactivities in sera were reproducibly detected by immunoblotting, using human organ extracts as the antigen sources. In contrast, IgG reactivities were low in whole sera but were considerably increased after affinity-chromatography purification on protein G-Sepharose. These increases differed from one individual IgG preparation to another and from one antigen to another (from 1-94 times) resulting in a unique IgG autoreactivity pattern for each subject.IgG reactivities diminished markedly when the IgG-depleted serum was added to the isolated autologous IgG. IgM antibodies isolated from sera on F(ab')2 IgG immunoadsorbent partially inhibited the binding of IgG to tubulin and myosin but not to actin. The individual IgG preparations examined separately exhibited, with all the autoantigens of the panel, higher autoreactivities than those of the same-but-pooled IgGs, which in turn were higher than those of a commercially available human IgG preparation obtained from approximately 8,000 healthy donors and used for intravenous injection. Depending upon the individual IgG sample, 31-65% of the IgG were bound to a DNP-Sepharose column and were eluted with DNP-glycine. The isolated anti-DNP antibodies were found to be polyreactive and possess higher autoreactivities than the original IgG preparation for all the antigens of the panel. Similarly, IgG antibodies analysed using an antibody exchange procedure were found to be essentially polyreactive but some apparently monospecific antibodies were also noted.These results suggest that the great majority of IgG present in normal humans are composed of polyreactive autoantibodies. IgG autoreactivities are only marginally expressed in these whole sera because of IgM-IgG, IgG-IgG and other, still unidentified, interactions.

SUMMARYSeveral hutiian monoclonal immunoglobtilins with the same auloatitibody activity have been shown to have cross-reactive idiotypes (CRI). In this study, using polyclonal anti-idiotypic antibodies, we fotind that 28% of hutnan monoclonal immunoglobulins with polyreaetive autoantibody activity from myeloma. Waldenstrom's macroglobulinaemia and eryoglobulinaemia patients shared common idiotype(s). Furthermore, the latter were expressed on human and murine natural MoAbs (respectively in 12% and 22% of the clones tested) and on human IgG preparations used for therapeutic intravenous injections (IVlg) and which contain natural antibodies. These findings suggest that monoclonal immunoglobulins could arise from the proliferation of a clotie that normally produces a natural antibody. The existenee of common idiotype(s) between monoclonal immunoglobulins and IVIg could be relevant to the improvement noted after treatment with IVIg in patients suffering from peripheral neuropathies associated with monoclonal gammopathy.

The characteristics of B cell lines isolated from patients with Alzheimer's disease (AD) and age-matched controls were investigated after having been transformed by Epstein-Barr virus (EBV). After isolation of mononuclear blood cells and in vivo or in vitro EBV infection, 35 and 21 lymphoblastoid cell lines (LCLs) were generated from 19 patients with AD (mean age 79.4 years) and 21 age-matched controls (mean age 80.0 years), respectively. B lymphocytes from AD patients were immortalised more easily than those from controls; the percentage of in vitro EBV infected LCLs (B95-LCLs) obtained in the AD group was significantly higher (76.2% versus 33.3% in the control group) and the mean time required for establishment was significantly lower (20.2 and 21.9 days versus 26.7 and 60.9 days in the control group). The EBV receptor and surface immunoglobulin (Ig) analyses showed no difference between the two groups. The expression of Epstein-Barr early antigens (EA) and viral capsid antigens (VCAs) revealed a tendency to higher viral replication in LCLs from AD patients; however, VCA expression remained limited to a small number of cells and did not affect overall cell growth. Finally, qualitative and quantitative differences were observed in the pattern of Ig production. Whereas spontaneously established LCLs from AD patients were generally monoclonal (80% of LCLs versus 33% in the control group), B95-LCLs were all polyclonal and secreted more IgM and IgA than those from controls; the mean IgM level was significantly higher in B95-LCLs from the AD group. These results suggest that B cells derived from AD patients seemed to be less differentiated than cells from age-matched controls.

The pathogenesis of gold-induced autoimmunity and membranous glomerulopathy is not well understood. HgCl2 and D-penicillamine, other chemicals known to trigger membranous glomerulopathy in humans, induce autoimmune manifestations in Brown-Norway (BN) rats but not in Lewis (LEW) rats. These chemicals trigger T-cell clones which are specific for self class II molecules from the major histocompatibility complex and are probably responsible for the polyclonal B-cell activation observed. The aim of this work was to test the effects of aurothiopropanolsulphonate (ATPS) in BN and LEW rats. In BN rats, ATPS induced a polyclonal B-cell activation marked by lymphoproliferation, hyperimmunoglobulinaemia affecting mainly IgE, and by the production of numerous autoantibodies. A glomerulonephritis occurred, initially due to anti-glomerular basement membrane antibody deposition, and later to the formation of granular deposits, occasionally resulting in a typical membranous glomerulopathy. Self class-II-specific T-cells were found that might be responsible for the polyclonal B-cell activation. Lewis rats were free of glomerulopathy but, like BN rats, exhibited an interstitial nephritis and some degree of polyclonal B-cell activation. These findings demonstrate that, depending on the strain, ATPS triggers different B-cell clones inducing different degrees of autoimmunity.

Immune complexes formed by mouse polyspecific natural autoantibodies and various structurally different antigens, such as DNA, tubulin and myosin, were analysed by rotary-shadowing electron microscopy. Each of the four natural IgM autoantibodies studied (E7, D23, 3C3 and M2-9) recognized multiple epitopes on the myosin molecule. These results, confirmed by immunoblotting experiments using myosin subfragments as antigens, strikingly contrasted with those obtained with an induced myosin-specific IgG antibody which interacted with a single myosin antigenic site.Based on the measurements of the antibody position on the antigen, made on a series of electron micrographs, two negatively charged myosin peptides were prepared by solid phase synthesis. Polymeric forms of one of the two peptides interacted with the positively charged CDR part of E7 and inhibited the binding of E7 and M2-9 to myosin. The importance of charge in the observed cross-reactivities was further supported by enzyme immunoassays showing that most, but not all, antigen/natural autoantibody interactions were sensitive to increasing concentrations of NaCl.

A total of 463 EBV Ig-secreting clones were derived from embryonic tissues, cord blood, and adult peripheral blood. Subcloning and analysis of the H and K loci (germline vs rearranged DNA status) of 44 primary clones insured clonality in at least 92% of cases. Whatever the cell origin, a somewhat constant proportion of clones (i.e., 11 to 16%) expressed polyspecific antibodies when tested on a panel of nine Ag, including self-Ag. The VH and VK repertoires have been studied using VH1-VH6 and VK1-VK4 family-specific probes. For all EBV clones the VH and VK utilization was similar to that of the normal untransformed population. A correlation was observed between the level of expression and the gene number for VH, whereas a clear distortion appeared for VK. Moreover, the usage pattern of VH and VK families of the polyspecific clones did not significantly differ from that of clones of unknown specificity, suggesting that polyspecificity was not linked to a restricted repertoire.

Pooled normal human polyspecific IgG (IVIg) contain anti‐idiotypes against a variety of autoantibodies from patients with autoimmune diseases and IgG autoantibodies present in IVIg. The present study indicates that IVIg may also react through idiotypic/anti‐idiotypic interactions with human natural IgM antibodies. Sixty‐four percent of IgM secreted by B lymphoid cell lines derived from B cells of healthy elderly donors and 18% of IgM secreted by cloned EBV‐transformed cord B cells that were tested, bound through their variable region to F(ab') 2 fragments of IVIg. The binding to 2,4,6‐trinitrophenyl (TNP) of a polyreactive IgM with anti‐TNP specificity, was inhibited by F(ab') 2 fragments from IVIg, indicating the presence in IVIg of anti‐idiotypes that may interfere with the antibody‐combining site of polyreactive IgM antibodies. The ability of IgM antibodies to interact with idiotypes on IVIg was not related to the degree of polyreactivity of natural antibodies. Our observations further document that IVIg contain antibody specificities against Ig from normal individuals and suggest that IgG originating from the physiologically expressed repertoire may modulate the expression of the potential B cell repertoire. The results may be relevant to the suppressive effect of IVIg in autoimmune diseases.

Sera from 19 patients with Alzheimer's disease (AD) and 21 control subjects were studied by immunofluorescence and enzyme immunoassay for antibody activity against various viruses and 12 self- and non-self-antigens. Total IgG mean level was significantly higher in the AD group; the IgG level was above 15 g/L in 52.8% of AD patients versus 14.3% of control subjects. Antiviral antibody titers showed no significant differences except for antibodies to herpes simplex virus-1, which were increased in control group. In contrast, autoantibodies were more frequently found in AD patients, and the prevalence of antibodies to spectrin, peroxidase, and thyroglobulin was significantly increased. Thus, in our series, autoimmune but not antiviral responses were heightened in at least 42% of AD patients (versus 9% of the control group) suggesting the existence of two subpopulations in the AD group.

Sera from rabbits before and after repeated injections of pig tubulin in complete Freund's adjuvant were examined for antibody activity against pig and rabbit tubulins and against a panel of antigens: actin, myosin, DNA, TNP/BSA. Antibody activity against all the antigens of the panel (PAg) increased moderately after the first but not after subsequent injections. Antibody activity against pig and rabbit tubulins strongly increased after the second immunization when the maximum was reached. Isolation of anti-tubulin antibodies from normal or immune sera on tubulin-immunoadsorbent demonstrated the presence of three different antibody populations: (1) polyspecific IgM reacting with the PAg and the tubulins, present in substantial amounts in normal sera and moderately increased in immune sera; (2) small amounts of polyspecific IgG detected only in immune sera; (3) high amounts of specific IgG reacting with pig and rabbit tubulins, present in immune but not normal sera. Western blot analysis of the specific IgG population showed that it contained antibodies reacting with both native pig and rabbit tubulins, as well as antibodies recognizing only the 30,000 proteolytic fragment of pig, but not that of rabbit tubulin. The results indicate that the immunization of rabbits with heterologous tubulin induced specific IgG anti-tubulin antibodies which recognize the self and non-self antigens differently.

Experimental Chagas' disease—infection of mice with the protozoan parasite Trypanosoma cruzi—has been shown to increase the number of Ia-bearing cells in the spleen and the lymph nodes. The majority of these Ia-positive cells were Ig+ and included in the large cell fraction of lymphoid organs from T. cruzi-infected animals indicating that they were activated B cells. These data are consistent with the polyclonal B-cell activation occurring during acute and chronic T. cruzi infection. The levels of secreted natural antibodies, of both IgM and IgG isotypes, were significantly increased in the sera of the infected animals. The present communication demonstrates that in vivo anti-Ia treatment of C3HHeJ mice infected with the CL strain of T. cruzi suppressed the polyclonal B-cell activation, affecting all the isotypes studied, including IgM, IgG2a and IgG2b, whose levels are predominantly increased during T. cruzi infection. In contrast to the decreased secretion of IgG autoantibodies, the levels of IgM autoantibodies were much less affected. The anti-Ia treatment totally abolished the specific anti-parasite response despite the fact that a pool of Ia-Ig positive cells remained after treatment.

Newborn BALB/c mice were repeatedly injected either with syngeneic (BALB/c) or xenogeneic (bovine) myosin, albumin, or actin in sterile physiological saline. The serum antibody response was evaluated by enzyme immunoassay 1 and 2 months after birth and after two booster injections. At 1 month, higher antibody titres were found in the sera of mice injected with syngeneic than with xenogeneic antigens. At 2 months and after boosting, anti-syngeneic actin antibodies were present in equal or higher amounts, anti-syngeneic albumin antibodies were not detected, and anti-syngeneic myosin antibodies were considerably decreased. Antibodies produced after booster injections of syngeneic actin were found to be highly specific and to belong mainly to the IgG isotype. These results suggest that newborn mice are better able than adult mice to respond to stimulation with self antigens, and that administration of self proteins during neonatal life may lead to the induction of immunological memory. They also indicate that one of the primary functions of the immune system in newborn mice is the recognition of self antigens.

Using a panel of antigens (actin, myosin, tubulin, albumin, transferrin, peroxidase, thyroglobulin, DNA, prolactin, TNP and myelin basic protein (MBP], we have tested the antibody activity of serum and cerebrospinal fluid (CSF) from healthy individuals, patients with multiple sclerosis (MS) and individuals with other neurological diseases. No differences in the concentrations and specificities of the serum antibodies were observed among the 3 groups. In contrast, we found that MS patients often had elevated CSF antibody levels against many antigens of the panel. The MS patients with local immunoglobulin production in the central nervous system (CNS) had the highest antibody levels. Restricted antibody activity against a given antigen of the panel was not observed. Compared to the two other groups, the MS group had equivalent titres of anti-MBP antibodies in the CSF. These results suggest that, in MS, a general immune dysregulation exists which leads to a local expansion of B lymphocytes producing autoantibodies with reactivities similar to those of serum natural autoantibodies.

Recent studies have shown that autoreactive B cells and autoantibodies are present in pathological as well as in normal situations. In the present study, we immortalized human B cell lines from normal individuals and from patients with malignant or benign dysglobulinemia with Epstein-Barr virus and examined, after cloning, the autoantibody reactivities of the immunoglobulins secreted by these cells. Forty-two supernatants were analyzed by enzyme-immunoassay on a panel of 13 self and non-self antigens: trinitrobenzenesulfonic acid (TNP), DNA, L- glutamine, L-alanine, L-tyrosine (GAT), actin, myosin, tubulin, albumin, renin, spectrin, transferrin, thyroglobulin, myoglobin, peroxidase, and by immunofluorescence in tissue sections. Fourteen (33%) of the immunoglobulin-secreting cell lines were found to have an autoantibody function; seven secreted IgM, six IgA, and one IgG. The light chains were of the kappa type in 11 cases. The vast majority of these clones reacted with more than five antigens of the panel and all of them reacted with TNP. No correlation was found between a given isotype and an antibody specificity. More than half of these antibodies also reacted with cellular antigens present in tissue sections. None of the four cell lines secreting monoclonal antiviral antibodies reacted with any of the antigens of the panel. The results indicate that immunoglobulins secreted by human monoclonal lymphoid cell lines can have polyspecific autoantibody functions, similar to those found in normal human polyclonal antibodies, in human monoclonal paraproteins and in natural monoclonal antibodies synthesized by murine or rat clones obtained from physiologically normal animals.

Click to increase image sizeClick to decrease image sizeKey Words: natural polyspecific autoantibodies