Argyrios N. Theofilopoulos

Murine Models of Systemic Lupus Erythematosus

Publisher Summary This chapter reviews the histopathologic, serologic, lymphocytic, virological, hormonal, and genetic characteristics of murine models of systemic lupus erythematosus (SLE). The pathogenetic mechanisms underlying murine SLE are highly complex, apparently well-programmed genetically, but still diverse and their bases not as yet well defined. Significant serologic and cellular experimental data support the statement that the final immunopathologic perturbation in murine (and human) SLE is a B lymphocyte hyperactivity with corresponding enhancement of serum antibodies and autoantibodies, particularly IgG, and consequent formation of pathogenic antigen–antibody ICs. On the basis of the available data, it appears that this B cell hyperactivity is polyclonal but not necessarily panclonal in nature, because not only antibodies against a wide array of autoantigens but also antibodies against incidental antigens, such as haptens, can be detected. The presence of autoantibodies and of their idiotypes in some recombinant lupus x normal murine lines expressing the normal partners V genes, normal mice in which an exogenous or endogenous (Ipr gene) immunostimulator has been introduced, and unmanipulated normal mice indicate that lupus mice are not unique in their structural genetic Ig elements, whose presence determines the development of their autoimmune disease. The cause of B cell hyperactivity in lupus has not yet been fully elucidated. Autonomous B cell maturation does not appear likely, because B cell proliferation and differentiation in lupus mice was found to be dependent on the same number of accessory signals as in normal mice.

The Raji cell radioimmune assay for detecting immune complexes in human sera.

A sensitivie and simple procedure for the detection and quantitation of soluble complement (C)- fixing immune complexes in sera of patients with various disease states has been developed by utilizing C receptors on Raji cells. These cells lack membrane-bound immunoglobulin but have receptors for IgG Fc, C3b, C3d, and possibly with other C proteins. Uptake experiments showed that both aggregated human gamma globulin (AHG) and 7S IgG bound to receptors for IgG Fc; however, AHG reacted with C bound to cells only via receptors for C and this binding was much more efficient than via IgG Fc receptors. AHG was used as an in vitro model of human immune complexes and its uptake by Raji cells was quantitated by 125I-radiolabeled antihuman IgG. The limit of sensitivity of this test was 6 mug AHG/ml serum. The ability of Raji cells to detect AHG in serum depended on the amount of radioactive antibody used and the size of aggregates. The presence of an excess of C somewhat inhibited binding of AHG containing C to Raji cells. The efficient binding of AHG by receptors for C on Raji cells was used for the detection and quantitation of immune complexes in human sera. Raji cells were incubated with sera to be tested and then reacted with excess radiolabeled antihuman IgG; the amount of radioactivity bound to the washed cells was determined and referred to a standard curve of radioactive antibody uptake by cells previously incubated with increasing amounts of AHG in serum. Thereby immune complexes were detected and quantitated in serum hepatitis, systemic lupus erythematosus, vasculitis, subacute sclerosing panencephalitis, dengue hemorrhagic fever, and malignancies.

Type-I Interferon Receptor Deficiency Reduces Lupus-like Disease in NZB Mice

Indirect evidence suggests that type-I interferons (IFN-α/β) play a significant role in the pathogenesis of lupus. To directly examine the contribution of these pleiotropic molecules, we created congenic NZB mice lacking the α-chain of IFN-α/βR, the common receptor for the multiple IFN-α/β species. Compared with littermate controls, homozygous IFN-α/βR-deleted NZB mice had significantly reduced anti-erythrocyte autoantibodies, erythroblastosis, hemolytic anemia, anti-DNA autoantibodies, kidney disease, and mortality. These reductions were intermediate in the heterozygous-deleted mice. The disease-ameliorating effects were accompanied by reductions in splenomegaly and in several immune cell subsets, including B-1 cells, the major producers of anti-erythrocyte autoantibodies. Decreases of B and T cell proliferation in vitro and in vivo, and of dendritic cell maturation and T cell stimulatory activity in vitro were also detected. Absence of signaling through the IFN-α/βR, however, did not affect increased basal levels of the IFN-responsive p202 phosphoprotein, encoded by a polymorphic variant of the Ifi202 gene associated with the Nba2 predisposing locus in NZB mice. The data indicate that type-I IFNs are important mediators in the pathogenesis of murine lupus, and that reducing their activity in the human counterpart may be beneficial.

Etiopathogenesis of Murine SLE1

Autoimmunity has been considered to represent termination of the natural unresponsive state to self {Weigle 1977). It was originally thought that selftolerance is achieved by elimination of self-reactive lymphocyte clones early in ontogeny (Burnet 1959). However, we now know that such self-reactive clones are normally present in the intact immune system {Clagett & Weigle 1974, Cohen & Wekerle 1973, Bankhurst et al. 1973, Bankhurst & Williams 1975, Sawada et al. 1977, Wigzell 1977) and that self-recognition, based on the complementarity of interacting structures on T cells, B cells and macrophages, is the basis for immune responses {Katz 1977). The prevailing current concept is that self-recognition processes, essential to immune and possibly other physiologic systems, are normally controlled or suppressed by one or more mechanisms that counteract deleterious autoreactive consequences. A variety of events, either exogenous or endogenous, could tip the balance of such regulatory supervision, thereby upsetting its normal damping effect so that harmful autoreactivity is the outcome. Sometimes this imbalance is only transient, superceded by reestablishment of the normal damping mechanism and a return to control over the system. At other times, the defect is permanent, leading to a progressive increase in harmful autoreactivity. This sequence seems to account for the chronic autoimmune phenomena seen in some strains of mice and in certain human diseases, most notably systemic lupus erythematosus (SLE), the prototypic autoimmune disease.

The Biology and Detection of Immune Complexes

Publisher Summary This chapter describes the interaction of immune complexes (ICs) with complement and with the cells of the immune system. The effect of immune complexes depends to a great extent on their antigen–antibody ratio so that their influence, either stimulatory or suppressive, is itself modulated by quantitative aspects of the immune response. The development of numerous techniques for the detection and quantitation of immune complexes has stimulated clinically related research and expanded the list of diseases in which immune complexes appear to play an important role. An extension of this diagnostic technology is the ability to isolate immune complexes and, in turn, their antigenic component, thereby making it possible to identify the antigens involved in immune processes of a great many diseases, including those of unknown etiology. In vivo and in vitro experiments have clarified many factors involved in IC formation, removal, and localization as well as the mechanisms of IC-induced inflammatory reactions. An individual can make an immune response to a large number of exogenous and a smaller number of endogenous antigens. Depending upon the availability of antigen, the antibodies so produced form ICs, which for the most part serve the purpose of aiding the host in eliminating potential pathogens.

TLR-dependent and TLR-independent pathways of type I interferon induction in systemic autoimmunity

We formulate a two-phase paradigm of autoimmunity associated with systemic lupus erythematosus, the archetypal autoimmune disease. The initial Toll-like receptor (TLR)-independent phase is mediated by dendritic cell uptake of apoptotic cell debris and associated nucleic acids, whereas the subsequent TLR-dependent phase serves an amplification function and is mediated by uptake of TLR ligands derived from self-antigens (principally nucleic acids) complexed with autoantibodies. Both phases depend on elaboration of type I interferons (IFNs), and therapeutic interruption of induction or activity of these cytokines in predisposed individuals might have a substantial mitigating effect in lupus and other autoimmune diseases.

T cell homeostatic proliferation elicits effective antitumor autoimmunity

Development of tumor immunotherapies focuses on inducing autoimmune responses against tumor-associated self-antigens primarily encoded by normal, unmutated genes. We hypothesized that such responses could be elicited by T cell homeostatic proliferation in the periphery, involving expansion of T cells recognizing self-MHC/peptide ligands. Herein, we demonstrate that sublethally irradiated lymphopenic mice transfused with autologous or syngeneic T cells showed tumor growth inhibition when challenged with melanoma or colon carcinoma cells. Importantly, the antitumor response depended on homeostatic expansion of a polyclonal T cell population within lymph nodes. This response was effective even for established tumors, was characterized by CD8(+) T cell-mediated tumor-specific cytotoxicity and IFN-gamma production, and was associated with long-term memory. The results indicate that concomitant induction of the physiologic processes of homeostatic T cell proliferation and tumor antigen presentation in lymph nodes triggers a beneficial antitumor autoimmune response.

Interferon-gamma is required for lupus-like disease and lymphoaccumulation in MRL-lpr mice.

Congenic MRL-lpr mice homozygous and heterozygous for the IFN-gamma gene disruption were created to assess the role of this pleotropic cytokine on the lymphoaccumulation and lupus-like disease of Fas-defective mice. Early death was prevented, and glomerulonephritis severely reduced in IFN-gamma-/- mice. Hypergammaglobulinemia was maintained with a switch from IgG2a to IgG1 predominance, but the dramatic decrease in levels of the dominant IgG2a anti-dsDNA autoantibodies was not associated with a compensatory increase in TH2-associated IgG subclasses. Remarkably, early death and glomerulonephritis were also prevented in IFN-gamma+/- mice, although autoantibody levels and glomerular immune deposits were equivalent to IFN-gamma+/+ lpr mice, indicating the importance of additional locally-exerted disease-promoting effects of IFN-gamma. IFN-gamma-/- mice exhibited reduced lymphadenopathy concomitant to a decrease in DN B220(+) T cells. In vivo BrdU labeling showed reduced proliferation of DN B220(+) cells in IFN-gamma-/- vs. IFN-gamma+/+ lpr mice, while enhanced proliferation of all other T cell subsets was unaffected. Macrophages of IFN-gamma-/-lpr mice expressed markedly decreased levels of MHC class I and II molecules compared with controls. Moreover, the heightened expression of MHC class II molecules on proximal tubules of IFN-gamma+/+ lpr mice was significantly reduced in both IFN-gamma-/- and IFN-gamma+/- mice. The data indicate that IFN-gamma hyperproduction is required for lupus development, presumably by increasing MHC expression and autoantigen presentation to otherwise quiescent nontolerant anti-self T cells, and also by promoting local immune and inflammatory processes.

The Pathogenic Effect of IgG4 Autoantibodies in Endemic Pemphigus Foliaceus (Fogo Selvagem)

Endemic pemphigus foliaceus, or fogo selvagem, is an autoimmune blistering skin disease caused by IgG autoantibodies to a desmosome-associated glycoprotein. We studied the IgG subclasses with autoantibody activity in serum from 29 patients with active disease and in the skin lesions of 18 patients by immunofluorescence, using IgG-subclass-specific monoclonal antibodies. The predominant disease autoantibodies present in all patients were of the IgG4 subclass. IgG1 and IgG2 autoantibodies were detected in low titer in the 29 patients: IgG1 in 23 patients and IgG2 in 9. IgG3 autoantibodies were not detected in the serum of any patient. Direct immunofluorescence testing of skin lesions showed a preferential deposition of IgG4 on the keratinocyte surface. The pathogenic effect of IgG4 was demonstrated by the passive transfer of fractions containing IgG4 autoantibodies from the patients to neonatal BALB/c mice. The disease of the patients was reproduced clinically, histologically, and immunologically in these animals. Only IgG4 autoantibodies were detected by direct immunofluorescence, bound to the epidermis in the lesions of the mice, and by immunoelectron microscopy at the keratinocyte surface. IgG4 has previously been reported to be a blocking or protective antibody because it has poor effector functions in vitro, as compared with the other IgG subclasses. The finding that it is the pathogenic autoantibody in fogo selvagem raises the possibility that it may also be important in other autoimmune disease.

The basis of autoimmunity: Part I mechanisms of aberrant self-recognition

In this two-part series, Argyrios N. Theofilopoulos summarizes the current state of affairs in the field of autoimmunity. Part I integrates the collective mechanistic theories of autoimmune diseases. The most straightforward explanation to emerge with regard to organ-specific diseases is the concept that these are caused by inappropriate, yet conventional, immunological responses against self-antigens for which tolerance has never been established. A similar mechanism may be operative in systemic autoimmunity, but other abnormalities such as defects in the apoptosis machinery may also be invoked. Part II will address the genetic contributions predisposing to autoimmune syndromes.