Production Of Natural Autoantibodies Research Articles

Revisiting Cell Death Responses in Fibrotic Lung Disease: Crosstalk between Structured and Non-Structured Cells.

Fibrosis is a life-threatening disorder caused by excessive formation of connective tissue that can affect several critical organs. Innate immune cells are involved in the development of various disorders, including lung fibrosis. To date, several hematopoietic cell types have been implicated in fibrosis, including pro-fibrotic monocytes like fibrocytes and segregated-nucleus-containing atypical monocytes (SatMs), but the precise cellular and molecular mechanisms underlying its development remain unclear. Repetitive injury and subsequent cell death response are triggering events for lung fibrosis development. Crosstalk between lung structured and non-structured cells is known to regulate the key molecular event. We recently reported that RNA-binding motif protein 7 (RBM7) expression is highly upregulated in the fibrotic lung and plays fundamental roles in fibrosis development. RBM7 regulates nuclear degradation of NEAT1 non-coding RNA, resulting in sustained apoptosis in the lung epithelium and fibrosis. Apoptotic epithelial cells produce CXCL12, which leads to the recruitment of pro-fibrotic monocytes. Apoptosis is also the main source of autoantigens. Recent studies have revealed important functions for natural autoantibodies that react with specific sets of self-antigens and are unique to individual diseases. Here, we review recent insights into lung fibrosis development in association with crosstalk between structured cells like lung epithelial cells and non-structured cells like migrating immune cells, and discuss their relevance to acquired immunity through natural autoantibody production.

Toll-Like Receptor Mediated Activation of Natural Autoantibody Producing B Cell Subpopulations in an Autoimmune Disease Model.

Altered expression and function of the Toll-like receptor (TLR) homologue CD180 molecule in B cells have been associated with autoimmune disorders. In this study, we report decreased expression of CD180 at protein and mRNA levels in peripheral blood B cells of diffuse cutaneous systemic sclerosis (dcSSc) patients. To analyze the effect of CD180 stimulation, together with CpG (TLR9 ligand) treatment, on the phenotype defined by CD19/CD27/IgD/CD24/CD38 staining, and function (CD69 and CD180 expression, cytokine and antibody secretion) of B cell subpopulations, we used tonsillar B cells. After stimulation, we found reduced expression of CD180 protein and mRNA in total B cells, and CD180 protein in B cell subpopulations. The frequency of CD180+ cells was the highest in the CD19+CD27+IgD+ non-switched (NS) B cell subset, and they showed the strongest activation after anti-CD180 stimulation. Furthermore, B cell activation via CD180 induced IL-6 and natural autoantibody secretion. Treatment with the combination of anti-CD180 antibody and CpG resulted in increased IL-6 and IL-10 secretion and natural autoantibody production of B cells. Our results support the role of CD180 in the induction of natural autoantibody production, possibly by NS B cells, and suggest an imbalance between the pathologic and natural autoantibody production in SSc patients.

AB0734 Prevalence and co-occurrence of autoantibodies in blood donors

BackgroundIncidence of autoimmune diseases, including systemic lupus erythematosus (SLE) and rheumatoid arthritis, has continuously increased over the past decades [1]. Different autoimmune diseases seem to co-occur at greater than expected...

Aerobic exercise training as a potential source of natural antibodies protective against human immunodeficiency virus‐1

Despite the effectiveness of HAART in controlling HIV-1 replication, the emergence of drug-resistant viruses in infected patients and the severe side effects caused by the currently used drug regimens and the lack of an effective vaccine necessitate the continued search for new therapeutic strategies for prevention and therapy of HIV disease. Previously we reported that natural autoantibodies, recognizing peptide FTDNAKTI (peptide NTM1) derived from the C2 domain of HIV-1 gp120, contribute to the control of HIV disease. Here we demonstrated that sera from well-trained athletic (HIV-negative) subjects showed high reactivity with peptide NTM1. This result confirms that aerobic exercise training stimulates production of natural autoantibodies, which recognize peptide NTM1. Bioinformatics analysis indicates that these natural autoantibodies could slow down disease progression by blocking the superantigenic site on HIV-1 gp120. The results suggest that aerobic exercise training may be a promising non-toxic and inexpensive adjunctive anti-HIV therapy.

Augmented TLR9-induced Btk activation in PIR-B–deficient B-1 cells provokes excessive autoantibody production and autoimmunity

Pathogens are sensed by Toll-like receptors (TLRs) expressed in leukocytes in the innate immune system. However, excess stimulation of TLR pathways is supposed to be connected with provocation of autoimmunity. We show that paired immunoglobulin (Ig)-like receptor B (PIR-B), an immunoreceptor tyrosine-based inhibitory motif–harboring receptor for major histocompatibility class I molecules, on relatively primitive B cells, B-1 cells, suppresses TLR9 signaling via Bruton's tyrosine kinase (Btk) dephosphorylation, which leads to attenuated activation of nuclear factor κB p65RelA but not p38 or Erk, and blocks the production of natural IgM antibodies, including anti-IgG Fc autoantibodies, particularly rheumatoid factor. The autoantibody production in PIR-B–deficient (Pirb−/−) mice was further augmented in combination with the Faslpr mutation, which might be linked to the development of autoimmune glomerulonephritis. These results show the critical link between TLR9-mediated sensing and a simultaneously evoked, PIR-B–mediated inhibitory circuit with a Btk intersection in B-1 cells, and suggest a novel way toward preventing pathogenic natural autoantibody production.

The V(lambda)-J(lambda) repertoire of patients with systemic lupus erythematosus manifests characteristics of the natural antibody repertoire.

To understand in detail the mechanisms of autoantibody production in patients with systemic lupus erythematosus (SLE), we performed a comprehensive analysis of the normal human immunoglobulin light chain V(lambda) repertoire and compared it with the V(lambda) repertoire in SLE patients. The SLE V(lambda) repertoire of B cells obtained from 3 SLE patients was analyzed and compared in detail with the V(lambda) repertoire of IgM+ B cells obtained from 3 human fetal spleens and IgM+,CD5+ B cells obtained from 2 normal adults. Conventional IgM+,CD5- B cells obtained from normal adults were used as controls. V(lambda)-J(lambda) rearrangements were amplified from the genomic DNA of individual B cells by polymerase chain reaction. The expressed V(lambda) repertoire of SLE patients contained several similarities with the expressed repertoire of the fetus and the adult CD5+ B cells. The V(lambda) genes 3L and 1G were overexpressed in the fetus, the adult CD5+ B cells, and the patients with SLE. The selection for rearrangements with restricted junctional diversity by utilization of homology-mediated joining, together with diminished N nucleotide addition, was a prominent feature of fetal, adult CD5+, and SLE B cell repertoires. Furthermore, profound expansion of V(lambda) clones with identical third complementarity-determining regions was observed in the adult CD5+, fetal, and SLE B cell repertoires. Notably, significant numbers of expanded adult CD5+ B cells, fetal, and SLE V(lambda) clones utilized homology-mediated joining at the V(lambda)-J(lambda) junctions. These data demonstrate that the SLE V(lambda)-J(lambda) repertoire manifests characteristics of normal adult IgM+,CD5+ and fetal B cell populations that are known to be enriched for the production of natural autoantibodies.

The Diabetes-Prone NZO/HlLt Strain. I. Immunophenotypic Comparison to the Related NZB/BlNJ and NZW/LacJ Strains

New Zealand Obese (NZO)/HlLt male mice exhibit a polygenic obesity and approximately 50% develop type 2 diabetes. This strain is known to produce a variety of autoantibodies, including autoantibodies to the insulin receptor. Because of their relatedness to the autoimmune-predisposed New Zealand Black (NZB) and New Zealand White (NZW) inbred strains, we compared NZO to its two related strains for shared hematologic and immunologic characteristics. Comparison of the three strains by serotyping and genotyping methods indicated that NZO shared with NZW the rare (recombinant) H2z haplotype at the major histocompatibility complex. Similar to the NZB and NZW strains, spleens from NZO mice contained increased numbers of CD19+CD43+ IgM+ B-1 B cells, a phenotype associated with natural autoantibody production. NZO mice developed a progressive microcytic anemia that was distinguished from NZB hemolytic anemia by absence of demonstrable antierythrocyte antibodies in the former. Outcross of NZO females with NZB males accelerated development of obesity and diabetes in F1 males. NZO males made B-lymphocyte–deficient by a disrupted immunoglobulin heavy chain gene did not become diabetic. These results suggest that NZO mice should be useful to investigators interested in studying the genetic contributions to autoimmunity made by the related NZW and NZB strains. Further, these results, combined with the pancreatic histopathology contained in the companion manuscript, suggest that B lymphocytes may be important contributors to diabetes pathogenesis in the NZO mouse.

The Pathologic Significance of the Immunoglobulins Expressed by Chronic Lymphocytic Leukemia B-cells in the Development of Autoimmune Hemolytic Anemia

The increased number of CD5+ B-cells in some human autoimmune diseases, the frequent commitment of CD5+ B-cells to the production of natural autoantibodies, and the apparent involvement of these cells in the pathogenesis of the autoimmune hemolytic anemia (Aiha) in certain mouse models suggests a causal relationship between the CD5+ chronic lymphocytic leukemia (CLL) B-cell and the Aiha which frequently develops in this malignant disorder. In support of this conclusion is our recent finding that the VH region gene repertoire of the leukemic B-cells from CLL patients with Aiha is rather biased and characterised by the over-representation of the 51 p1 VH gene. On the other hand, it appears relatively certain that the pathogenic anti-erythrocyte antibodies in CLL patients with Aiha are produced by remnant normal B-cells, and that the antibodies expressed by the leukemic CD5+ B-cells do not directly bind red blood cells (RBC). Of interest, the antibodies produced by the leukemic B-cells from CLL patients with Aiha might have in common rheumatoid factor (RF) activity. These data indicate that the antibodies produced by the leukemic B-cells from CLL patients with Aiha are not directly involved in red blood cell destruction, but may be involved in the induction or amplification of a polyclonal anti-RBC response. Finally, we discuss the possible clinical implications of our finding that CLL patients with leukemic cells expressing the 51 p1 VH gene may be at a higher risk to develop autoimmune hemolytic anemia.

Beneficial effect of polyclonal immunoglobulins from malaria-infected BALB/c mice on the lupus-like syndrome of (NZB x NZW)F1 mice.

We previously reported that infection of BALB/c mice with the parasite Plasmodium chabaudi induces high production of natural autoantibodies. Here we demonstrate that such an infection of lupus-prone (NZB x NZW)F1 (B/W) mice retards the development of their autoimmune disease. Survival and disease hallmarks (high-grade proteinuria and IgG anti-DNA antibodies) were delayed for 6 months when parasite inoculation was given at either 3 or 7 months of age, i.e. before or after the onset of the clinical symptoms. Similar beneficial effects, although less pronounced, were obtained when mice were treated with a total of 800 micrograms of IgG (P-IgG) or IgM (P-IgM) or 300 micrograms of cryoglobulin preparations isolated from P. chabaudi-infected BALB/c mice while similarly prepared fractions from uninfected mice had little effect. Compared to these fractions, P-IgG and P-IgM contained higher levels of natural antibodies bearing the D23 idiotype characteristic of polyreactive natural autoantibodies with enhanced activity against Fab and Fc fragments of IgG. In surviving mice, the level of anti-DNA antibodies, particularly those of IgG1 isotype, were significantly decreased. Flow cytometric analysis of various T cell subsets showed that the number of cells expressing gamma delta T cell receptor (TcR) antigens which did not vary with age was not modified after P-IgG or P-IgM treatment. In contrast, the number of T cells expressing V beta 8.1,2,V beta 10 and V beta 14 TcR antigens, which increased with age, were significantly reduced. Taken together, these results indicate that parasite infection of mice induces the synthesis of populations of IgM and IgG natural autoantibodies with immunoregulatory properties and that these antibodies attempt, at least transitorily, to rescue a natural autoantibody network that is deficient in B/W mice.

4 Biology of the neoplastic lymphocyte in B-CLL

Immunological markers have identified the proliferating lymphocyte in CLL as a mature B lymphocyte which, unlike lymphocytes in other B cell malignancies, expresses low amounts of surface membrane immunoglobulin (smIg), forms rosettes with mouse erythrocytes and expresses the CD5 marker. It has been postulated that Ly1 B cells (the murine counterpart of human CD5+B cells) constitute a separate B cell lineage. Whether the CD5 marker defines a discrete lineage or is a maturation marker is one of the main issues that might be solved in the near future. Another recent advance has been the discovery that the B lymphocyte in CLL is in an activated state and can be induced to differentiate. Using B cell mitogens and somatic hybridization, it has been demonstrated that the B-CLL lymphocyte is frequently involved in the production of natural autoantibodies and expresses a restricted set of genes. These results may provide a basis for passive immunotherapy using anti-idiotypic antibodies. Hypogammaglobulinaemia is a distinct feature of B-CLL, observed in 60% of patients. It may result from impaired function of residual normal B cells. This could occur as a consequence of progressive dilution of normal non-clonal B cells, or because normal B cells are downregulated by an unknown mechanism. This decrease in or inhibition of normal CD5-B cells could also explain the classical inability of patients with B-CLL to respond to new antigenic challenges, since Ly1 B cells have been claimed to be unable to respond to exogenous antigens. Although regulatory abnormalities in T cells may play a role in the induction of hypogammaglobulinaemia, data concerning helper, suppressive, NK and ADCC cells are contradictory and fail to establish firmly the contribution of these cells in the development of hypogammaglobulinaemia. Associated autoimmune phenomena are a prominent complication in CLL. They are related to the presence of autoantibodies directed mainly against blood components, which in most cases are not the product of the malignant clone. The relationship between autoimmune phenomena and hypogammaglobulinaemia is not definitively substantiated. That hypogammaglobulinaemia may determine the loss of some anti-idiotypic antibodies designed to antagonize autoimmune clones is an attractive hypothesis, which needs to be substantiated. Several recurrent chromosomal abnormalities, such as trisomy 12, structural aberrations of the 13q14 and 14q32 bands, are frequently observed in B-CLL. Less frequently, alterations of chromosomes 11, 6, 18, 3, 17, 7 and 8 have been reported.(ABSTRACT TRUNCATED AT 400 WORDS)

Surface markers, heavy chain sequences and B cell lineages.

A unifying theory of B cell development and lineage commitment is presented. There are two firmly established B lineages: cells which normally arise only from fetal sources and lack N insertions in their rearranged heavy chains; and N-containing cells which arise from adult bone marrow precursors (and perhaps from late fetal sources). Commitment to the expression of CD5 and the capacity for long-life (or self-renewal) are induced as a consequence of sIg cross-linking, typically by a repeating epitope, thymus independent type two antigen. Alternatively, activation resulting from cognate interaction with a helper T cell does not induce CD5 but results in lower expression of J11d. In this case activation occurs in the absence of sIg cross-linking. It is further proposed that differences in the Ig repertoire make it highly likely that fetal/neonatal, but not adult derived B cells will be induced to express CD5. The model offers a plausible explanation for the correlation of CD5 expression and natural autoantibody production by neonatal B cells. Possible sources of pathogenic autoantibody are discussed in the context of this model.

Evidence that chronic lymphocytic leukemia B lymphocytes are frequently committed to production of natural autoantibodies

CD5+ B lymphocytes have been postulated to be primarily involved in autoantibody production. To investigate whether CD5 B lymphocytes from chronic lymphocytic leukemia (CLL) patients display the same function, we fused B lymphocytes from 23 CD5+ B-CLL with nonsecreting murine myeloma X-63 cells. Hybrids were derived from 18 of the 23 patients, but only hybrids from 13 patients were found to secrete immunoglobulin (Ig) (IgM kappa 4, IgM lambda 8, and only kappa light chain 1). Supernatants of these hybrids were tested against an extensive panel of antigens by enzyme-linked immunosorbent assay, indirect immunofluorescence, and hemagglutination. At the same time, peripheral blood mononuclear cells (PBMC) from 15 of these patients, including most patients from whom secreting hybrids had not been obtained, were stimulated with phorbol myristate acetate (PMA). Our results indicated that secreting heterohybrids from CLL B lymphocytes are frequently obtained; however, this is highly dependent on the light chain phenotype, since 8 of 9 lambda-expressing CLL produced hybrids secreting complete Igs, as compared with 4 of 12 kappa-expressing CLL. In addition, B lymphocytes of most patients from whom we failed to derive secreting hybrids also failed to secrete Ig on PMA stimulation. Secondly, CD5+ B-CLL lymphocytes were frequently committed to the production of natural autoantibodies, since in 8 of 15 cases (including hybrids and PMA stimulation) anti-Fc activity was found; three of these also displayed a multispecific binding pattern.

Evidence That Chronic Lymphocytic Leukemia B Lymphocytes Are Frequently Committed to Production of Natural Autoantibodies

CD5+ B lymphocytes have been postulated to be primarily involved in autoantibody production. To investigate whether CD5 B lymphocytes from chronic lymphocytic leukemia (CLL) patients display the same function, we fused B lymphocytes from 23 CD5+ B-CLL with nonsecreting murine myeloma X-63 cells. Hybrids were derived from 18 of the 23 patients, but only hybrids from 13 patients were found to secrete immunoglobulin (Ig) (IgM kappa 4, IgM lambda 8, and only kappa light chain 1). Supernatants of these hybrids were tested against an extensive panel of antigens by enzyme-linked immunosorbent assay, indirect immunofluorescence, and hemagglutination. At the same time, peripheral blood mononuclear cells (PBMC) from 15 of these patients, including most patients from whom secreting hybrids had not been obtained, were stimulated with phorbol myristate acetate (PMA). Our results indicated that secreting heterohybrids from CLL B lymphocytes are frequently obtained; however, this is highly dependent on the light chain phenotype, since 8 of 9 lambda-expressing CLL produced hybrids secreting complete Igs, as compared with 4 of 12 kappa-expressing CLL. In addition, B lymphocytes of most patients from whom we failed to derive secreting hybrids also failed to secrete Ig on PMA stimulation. Secondly, CD5+ B-CLL lymphocytes were frequently committed to the production of natural autoantibodies, since in 8 of 15 cases (including hybrids and PMA stimulation) anti-Fc activity was found; three of these also displayed a multispecific binding pattern.

T cell receptor alpha/beta expressing double-negative (CD4-/CD8-) and CD4+ T helper cells in humans augment the production of pathogenic anti-DNA autoantibodies associated with lupus nephritis.

It is generally accepted that human Th cells express the surface glycoproteins CD4 and alpha/beta-chain heterodimer of the TCR whereas cytotoxic/suppressor cells are usually CD8+ and alpha/beta TCR+. Another minor set of T cells found in the periphery are CD4-/CD8- (double negative) and express the gamma/delta TCR; these cells can manifest MHC-restricted or nonrestricted cytotoxicity but no helper function. Herein we describe the existence of an unusual Th population in the peripheral blood of humans that are CD4-/CD8- and alpha/beta TCR+. These double-negative Th were markedly expanded in patients with the autoimmune disease SLE and along with CD4+ Th, they induced production of the pathogenic variety of anti-DNA autoantibodies that are IgG in class and cationic in charge. The cationic anti-DNA antibodies induced by the Th were markedly restricted in spectrotype indicating that an oligoclonal population of B cells were committed to produce the pathogenic autoantibodies in active lupus. IL-2-dependent T cell lines were also derived from the patients with active lupus nephritis but the majority of those T cell lines lacked pathogenic autoantibody-inducing capability. Only 4 out of 42 T cell lines from a lupus patient could induce the production of cationic IgG class anti-DNA autoantibodies. The phenotypes of the pathogenic autoantibody-inducing Th lines were similar to the Th subsets: CD4+, alpha/beta TCR+ or CD4-/CD8-, alpha/beta TCR+. These studies suggest that production of pathogenic autoantibodies in human lupus is mediated by mechanisms that are distinct from the generalized, nonspecific polyclonal B cell hyperactivity that leads to excessive production of natural autoantibodies.

Estrogen induces normal murine CD5+ B cells to produce autoantibodies.

Females have better humoral immune responses and are more susceptible to autoimmune diseases than males. Normal female mice (C57BL/6J, C3H/HeJ, and NZW) have significantly increased spontaneous autoimmune plaque-forming cells (APFC) to mouse erythrocytes pretreated with bromelain (Br-ME) in spleen, peritoneal exudate cell, and bone marrow compared to their male counterparts. A minor subpopulation of B cells, CD5+ B, is thought to produce this autoantibody. As determined by dual color flow cytometry, increased APFC to Br-ME in females is not due to quantitative increase of CD5+ B cells. Rather, it is due to increased numbers or percentages) of CD5+ B cells producing these autoantibodies, because CD5+ B cells from females produced greater numbers of APFC to Br-ME than equal numbers of cells derived from males. The increased autoantibody production in females can be attributed to the effect of estrogen on the immune response because this hormone markedly augments APFC to Br-ME in intact or orchidectomized males. Male hormone had little effect. Importantly, estrogen did not increase the numbers of B or CD5+ B cells but augmented the ability of B cells to produce this response. This was verified when a T cell-depleted B cell fraction or fluorescence-activated cell sorter purified CD5+ B cells from estrogen-treated mice proved more efficient in the production of APFC to Br-ME. These results suggest that the number of CD5+ B cells committed to produce autoantibodies to Br-ME is increased under the influence of estrogen. This is the first demonstration that estrogen can augment the production of natural autoantibodies in normal mice. The overall augmented humoral immune responses in females and the B cell hyperactivity in female predominant autoimmune diseases appears to be due to estrogen.

Newborn Xid mice carry the genetic information for the production of natural autoantibodies against DNA, cytoskeletal proteins, and TNP.

Spleen cells from 6-day-old unimmunized male and female (CBA/N X BALB/c)F1 mice, fused with the nonsecreting hybrid SP2/0, gave 184 hybrids secreting immunoglobulins (183 IgM class; 115 from males and 69 from females) which were screened for antibody (Ab) activity against actin, tubulin, myosin, TNP-BSA, dsDNA, and denatured DNA. Eleven hybrids from the male series (9.65%) and eight hybrids from the female series (11.55%) secreted immunoglobulins with significant Ab activity against at least one of the six antigens tested. Four of these clones reacted with a single antigen, while 15 others reacted simultaneously with at least two antigens. Three hybrid clones exhibited Ab activity against all of the antigens tested. The monoclonality and specificity of the immunoglobulins from tissue culture supernatants were respectively assessed by two-dimensional gel electrophoresis after biosynthetic labeling and competitive enzyme-linked immunosorbent assay. These results confirm our previous findings with newborn and adult BALB/c mice, demonstrating a high frequency of this auto-Ab repertoire, especially in newborn mice, and widespread polyspecificity found among these natural auto-Ab. Xid mice are characterized by the absence of a Lyb-3+,5+ B cell population; nevertheless, these studies suggest--but do not prove--that the Lyb-3-,5- lymphocyte population seems to carry genetic information for the production of natural auto-Ab, because no marked differences were observed in the secretory patterns of hybrids obtained from male mice bearing the Xid defect and from normal female mice. Hence, we postulate that the Xid gene might be a regulatory gene because Xid mice, in contrast with normal mice, are unable to secrete anti-DNA auto-Ab on mitogenic stimulation, despite carrying the genetic information.