Stanford L. Peng

T cells in murine lupus: propagation and regulation of disease

MRL/Mp-lpr/lpr mice develop a spontaneous lupus syndrome, including hypergammaglobulinemia, autoantibodies, glomerulonephritis, and lymphadenopathy. To investigate the role of lymphocyte subsets in the pathogenesis of disease, lupus-prone MRL mice deficient in αβ T cells, γδ T cells, or both were generated. Mice deficient in αβ T cells developed a partially penetrant lupus syndrome, characterized by lymphadenopathy, elevated levels of class-switched immunoglobulins, an increased incidence of antinuclear antibodies, and immune deposits in kidneys which progressed to renal insufficiency over time. In comparison to wild type animals, γδ T cell-deficient animals developed an accelerated and exacerbated disease phenotype, characterized by accelerated hypergammaglobulinemia and enhanced autoantibody production and mortality. Repertoire analysis of these latter animals identified polyclonal expansion (Vβ) of αβ CD4+B220-cells. Mice lacking both αβ and γδ T cells failed to generate class-switched autoantibodies and immune complex renal disease. First, these findings demonstrate that murine lupus in the setting of Fas-deficiency does not absolutely require the presence of αβ T cells, and they also suggest that a significant basis for MRL/lpr disease, including renal disease, involves αβ T cell-independent, γδ T cell dependent, polyreactive B cell autoimmunity, upon which αβ T cell-dependent mechanisms aggravate specific autoimmune responses. Second, these data indicate that γδ T cells partake in the regulation of systemic autoimmunity, presumably via their effects on αβ CD4+B220-T cells that provide B cell help. Finally, these results demonstrate that MRL/lpr B cells, despite their intrinsic abnormalities, cannot per se cause tissue injury without T cell help.

Pathogenesis of autoimmunity in αβ T cell-deficient lupus-prone mice

Murine lupus in MRL mice has been strongly attributed to αβ T cell‐dependent mechanisms. Non‐αβ T cell‐dependent mechanisms, such as γδ T cells, have been shown to drive antibody and autoantibody production, but they have not been considered capable of inducing end‐organ disease. Here, we have expanded upon the findings of such previous work by examining the mechanism and extent of end‐organ disease attainable via γδ T cells and/or non‐αβ T cell‐dependent mechanisms, assessing two prototypical lupus lesions, renal and skin disease, in TCR α−/− MRL mice that possessed either functional or defective Fas antigen (Fas + or lpr). Observed to 1 year of age, TCR α−/− MRL mice developed disease characterized by increased mortality, overt renal disease and skin lesions. While delayed in onset and/or reduced in severity compared with TCR α+/+ MRL/lpr animals, renal and skin lesions in αβ T cell‐deficient animals were clearly increased in severity compared with age‐matched control non‐autoimmune mice. In contrast to TCR α+/+ MRL mice, whose disease reflected pan‐isotype immune complex deposition with significant complement fixation, renal disease in TCR α−/− MRL animals reflected predominantly IgG1 immune complex deposition, with poor complement fixation. Thus, this study demonstrates conclusively that non‐αβ T cell‐dependent mechanisms can induce renal and skin injury in murine lupus, but at least in the kidney, only via humoral autoimmunity of a relatively non‐pathological isotype which results in the delayed onset of end‐organ damage.

Autoreactive T cells in murine lupus

The conventional paradigm to explain systemic lupus erythematosus (SLE) is that disease results from tissue deposition of pathogenic autoantibodies and immune complexes, secondary to activation of autoreactive B cells in the context of help from αΒ T cells. Recent work in murine lupus has confirmed this notion and demonstrated that autoantigen-specific αΒ T cells are absolutely required for full penetrance of disease, with such autoreactive αΒ T cells, even in Fasintact mice, likely arising from defects in peripheral tolerance. These studies have also revealed a network of regulation that also involves nonclassical pathogenic and downregulatory αΒ and γδ T cells, suggesting that the lupus immune system involves more complex interactions than the conventional paradigm suggests.

Spliceosomal snRNPs Autoantibodies

Publisher Summary This chapter includes intensive efforts to standardize the assay, define associations with disease or clinical presentation, and characterize antigenic and biological properties of spliceosomal snRNPs autoantibodies. Autoantibodies against the snRNP particles recognize both protein and RNA epitopes, including the trimethylguanosine cap. The categorizations for anti-snRNP antibodies include soluble nuclear specificity termed Sm and ribonucleoprotein (RNP) antibodies. The anti-Sm specificity includes autoantibodies that target proteins of the common Sm core, and anti-RNP refers to anti-U 1 snRNP-specific autoantibodies that target the U1 RNA or the U1- specific proteins 70K, A or C. Detection of anti-snRNP antibodies includes the indirect immunofluorescent antinuclear antibody test (ANA). The antispliceosomal snRNP autoantibodies are definitive specificities found in various rheumatological diseases. The anti-snRNP antibodies are described in several connective tissue diseases, especially systemic lupus erythematosus and mixed connective tissue disease.

SYSTEMIC AUTOIMMUNITY IN LG/J MICE

Humoral and end-organ parameters of autoimmunity were investigated in LG/J mice, which have traditionally been considered normal, non-diseased animals. Surprisingly, LG/J mice were found to possess autoantibodies, including antinuclear antibodies and rheumatoid factor, and to develop renal disease, including glomerulonephritis, interstitial nephritis, and perivasculitis, but not hepatic or cutaneous disease. In contrast, age-matched, identically-housed control animals failed to develop autoantibodies or end-organ disease. These findings have complications for the genetic study of lupus erythematosus in the MRL murine model, which derives heavily from the LG/J background. Thus, the LG/J strain may provide a useful model in the analysis of autoimmunity.

MHC class I polymorphism in lupus-prone MRL/Mp mice.

Systemic autoimmunity in MRL/Mp ( H2k) mice closely resembles human systemic lupus erythematosus, including the development of autoantibodies and immune-complex end-organ disease (Andrews et al. 1978). Many studies have focused upon class II major histocompatibility complex (MHC) genes in lupus pathogenesis, since class IIrestricted CD4+ T cells have been widely implicated in the generation of pathogenic autoantibodies (Griffing et al. 1980; Jevnikar et al. 1994; Ogawa et al. 1990; Santoro et al. 1988; Schiffenbauer and Wegrzyn 1991). Although some investigations have found significant associations of MHC class II polymorphisms with autoimmunity (Owaga et al. 1990; Todd et al. 1988), MRL MHC class II antigens do not demonstrate significant sequence differences from normal mouse strains bearing the H2k haplotype (Schiffenbauer and Wegrzyn 1991). At the same time, however, the haplotype plays a significant role in the pathogenesis of mouse lupus (Eisenberg et al. 1989), suggesting that other regions of the MRL MHC locus may be involved in disease induction. Recent studies have implicated a role for MHC class I antigens, not previously investigated in detail, in mouse lupus pathogenesis (Christianson et al. 1996; Mozes et al. 1993). Consequently, we sequenced the peptide binding regions of the MRL MHC class I antigens H2-D and H2-K to determine whether they possess potentially significant polymorphisms. Total RNA was purified from spleens of MRL/Mp-+/+ and congenic MRL/Mplpr/lpr mice (Jackson Laboratory, Bar Harbor, ME) using a standard CsCl gradient protocol (Sambrook et al. 1989). Single-strand cDNA synthesis was performed using random hexamers and AMV reverse transcriptase, as directed (Boehringer Mannheim, Indianapolis, IN). Polymerase chain reaction was performed using primrs H284F (59-TACTACAACCAGAGC) and H2168R (5 9GAGCCACTCCACGCA), which corresponded to consensus regions at amino acids 84 and 168, respectively, of the MHC class I gene. The 250 base pair (bp) products were cloned using the TA Cloning  kit (Invitrogen, San Diego, CA), and plasmid DNA was sequenced using standard SP6 and T7 primers by dideoxy sequencing (Sequenase TM Version 2.0; US Biochemicals, Cleveland, OH). Nucleotide sequences were compared against reference H2-Dk andH2Kk alleles (Pullen et al. 1992). The MRL H2-K allele contained no differences from the known H2-Kk sequence (data not shown). The sequence of the MRL H2-D allele, however, contained a T ? A mutation at nucleotide 419, causing a phenylalanine to tyrosine change at amino acid 116, and a conservative C ? T mutation at nucleotide 364 (Fig. 1). This F 116 ? Y116 transition involves a charge difference in a position that has been suggested to play a significant role in peptide specificity (Latron et al. 1992; Matsumura et al. 1992; Pullen et al. 1992). Although Y 116 is present in other reference MHC class I molecules, including H2-K b and H2-Kk, no normal strain has been found to express an H2-D molecule with Y116 (Pullen et al. 1992). Consequently, this finding suggests that the MRL H2-D molecule presents a peptide repertoire different from normal H2-D k molecules, and raises the possibility that, as suggested for other utoimmune systems (Kimoto et al. 1993), defective binding of a protective peptide epitope, or enhanced binding of a pathogenic epitope, promotes systemic autoimmunity in MRL mice. This definition of a genetic polymorphism in MRL mice will therefore aid in future investigations of the pathogenesis of murine lupus.