anti-ssDNA Antibodies Research Articles

Effect of Dietary Unsaturated Fatty Acids on Senile Amyloidosis in Senescence-Accelerated Mice

Effects of dietary oils on aging were investigated in senescence-accelerated mice. For 26 weeks, mice were fed purified diets containing 4% olive oil, safflower oil, perilla oil, or fish oil. Serum total, high-density lipoprotein cholesterol, and apolipoprotein A-II (ApoA-II) were significantly lower in the fish oil group than in the perilla oil group, and these were significantly lower than in the olive oil or safflower oil group. The olive oil and safflower oil groups had significantly fewer ApoA-II amyloid fibril (AApoAII) deposits and anti-single-strand DNA (ssDNA) antibodies than the fish oil or perilla oil group, and the fish oil diet induced significantly more AApoAII deposits and anti-ssDNA antibodies than did the perilla oil diet. Survival decreased earlier in the fish oil group than in the other groups (as seen in the survival curve). The results suggest that greater the degree of unsaturation of dietary fatty acids, greater is the tendency for accelerated senescence.

Recurrent apnea induces neuronal apoptosis in the guinea pig forebrain

Obstructive sleep apnea (OSA) and sleep-disordered breathing (SDB) can result in impaired cognition and mental acuity, and the generation of mood disorders, including depression. However, the mechanisms of neuronal damage for these complications have not been elucidated. Accordingly, using immunohistochemical technique with monoclonal antibody against single-stranded DNA, we examined the morphological effects of chronic recurrent apnea on neurons in the hippocampus and related forebrain sites in guinea pigs. Our results show that a large number of neurons labeled by anti-ssDNA antibody were present in the cingulate, insular and frontal cortices, the hippocampus and the amygdala in conjunction with periods of recurrent apnea. However, no labeling was observed in comparable regions of the brain in control guinea pigs. In the cortices of experimental animals, labeled neurons were detected mainly in the superficial layers (II–III) in the frontal, insular and cingulate cortex. In the hippocampus, most labeled neurons were located in the CA1 region, in which most of stained neurons were observed in strata pyramidal, while only a few positive neurons were located in the strata radiatum and the strata oriens. In addition, a large number of labeled neurons were also detected in the central nucleus of amygdala in the guinea pigs underwent recurrent periods of apnea. The present data indicate that recurrent apnea results in cell death in the hippocampus and related forebrain regions via mechanisms of apoptosis, which may represent the basis for the clinical complications of obstructive sleep apnea and sleep-disordered breathing.

Autostimulatory effects of IL-6 on excessive B cell differentiation in patients with systemic lupus erythematosus: analysis of IL-6 production and IL-6R expression.

Introducing avidin-biotin complex ELISA for anti-DNA antibody, the mechanism of in vitro production of anti-ssDNA antibody as well as of polyclonal immunoglobulin mediated by an IL-6-IL-6R loop was studied in patients with systemic lupus erythematosus (SLE). Regardless of the presence or absence of T cells, B cells from SLE patients could produce IgG anti-ssDNA antibody as well as total IgG without any stimulation. Low density B cells obtained by Percoll gradient density centrifugation responded to rIL-6 to produce IgG and IgG anti-ssDNA antibody. rIL-2 and rIL-4 had lesser effects on the differentiation of low density B cells. In fact, IL-6R was preferentially expressed on low density B cells from active SLE patients, as detected by anti-IL-6R MoAb, MT18, which did not inhibit IL-6 binding. SLE B cells, especially high density B cells, produced greater amounts of IL-6 in culture supernatants than did T cells, regardless of whether disease was active or inactive. Normal T cells and B cells did not produce significant amounts of IL-6. Thus, endogenous IL-6 produced by high density B cells bound to the IL-6R preferentially expressed on the low density B cells, and drove them into terminal differentiation, especially in active SLE patients. Further, addition of polyclonal anti-IL-6 or anti-IL-6R MoAb (PM1), which inhibited IL-6 binding, both inhibited IgG anti-ssDNA antibody as well as total IgG production by SLE B cells in a dose-dependent manner. These results suggest that interruption of the autocrine IL-6 loop would be of therapeutic value in SLE.

Endothelial cell binding by human polyclonal anti-DNA antibodies: relationship to disease activity and endothelial functional alterations.

Polyclonal anti-dsDNA and anti-ssDNA antibodies (PoAb) that showed significant binding to human umbilical vein endothelial cells (HUVEC) were isolated from eight patients with systemic lupus erythematosus (SLE). Anti-dsDNA PoAbs from five patients and anti-ssDNA PoAbs from seven patients demonstrated enhanced binding to HUVEC during active disease, compared with PoAbs obtained from corresponding patients during remission. Reduction of the DNA content in the PoAb preparations by DNase treatment was associated with enhanced binding to HUVEC in 20 of 32 PoAbs tested, which included 75% 'active disease' PoAbs, and with reduced binding to HUVEC in three of 32 PoAbs tested, all obtained during remission. Such altered endothelial cell binding was reversed with DNA reconstitution. Binding of the remaining nine PoAbs to HUVEC was not altered by variations in their DNA content. Induced plasma membrane expression of E-selectin, but reduced expression of vascular cell adhesion molecule-1 (VCAM-1) by HUVEC, was observed following incubation of HUVEC with 'active disease' PoAbs from three and two of the eight patients, respectively. PoAbs and serum samples from two of the eight patients during active disease induced von Willebrand factor release from HUVEC, which was not observed during remission. We conclude that anti-DNA antibodies from selected patients with SLE can bind to endothelial cells. Correlation between cellular binding and disease activity suggests that such binding of anti-DNA antibodies to endothelial cells could be of pathogenic significance. Preliminary data also suggest that the expression of adhesion molecules and haemostatic factor(s) by endothelial cells may be modified following their binding by anti-DNA antibodies.

A study of immune responses to myelin and cardiolipin in patients with systemic lupus erythematosus (SLE)

Sera from 39 patients with SLE, 20 patients with cerebrovascular disease with no evidence of SLE, and 20 normal controls were tested for antibodies to cardiolipin (CL), brain total upper (UPG) and lower phase (LPG) glycolipids, myelin basic protein (MBP), myelin, and single strand DNA (ssDNA) by ELISA. Binding to the glycolipids and MBP was negative or negligible in all the groups, but significant binding was observed against CL, myelin and ssDNA in some of the SLE patients. Many sera from SLE patients with cerebral disorders and high CL binding also demonstrated high binding to myelin. These sera also labelled cell surface antigens on neonatal mouse neurons and astrocytes by immunofluorescence in tissue culture. A correlation was found to exist between anti-CL and antimyelin antibodies in SLE patients with cerebral lesions, but not between anti-ssDNA and anti-CL antibodies. As much as 80-90% of the specific activity of these antibodies could be absorbed out by the relevant antigens but only partially by the other antigens. In the control groups binding was low and no specific absorption could be demonstrated.

Autoimmune syndrome after induction of neonatal tolerance to alloantigens: analysis of the role of donor T cells in the induction of autoimmunity

The injection of (C57BL/6 x BALB/c) F1 spleen cells into BALB/c newborn mice leads to activation of persisting F1 donor B cells and development of a lupus-like syndrome in tolerized BALB/c mice. This syndrome is characterized by hypergammaglobulinaemia, high levels of anti-DNA and anti-Sm antibodies, circulating immune complexes and deposits of immunoglobulin in renal glomeruli. The role of donor T cells in this model was investigated by injecting the newborn mice with F1 cells depleted in different T cell subsets by using specific monoclonal antibodies (MoAbs). Tolerance, as shown by an absence of H-2b-specific CTL alloreactivity and persistence of immunoglobulin bearing the donor allotype were observed in mice injected with F1 cells previously depleted in the CD4+ and/or CD8+ T cell subsets as well as in those which received Thy-1+-depleted F1 spleen cells. In these mice, a typical autoimmune syndrome was found, including splenomegaly and lymphadenopathy, anti-ssDNA and anti-aortic myosin IgG antibodies and renal deposition of immunoglobulin. However, some quantitative changes were seen: the levels of anti-aortic myosin antibodies were lower in mice tolerized with CD4+-depleted F1 cells than in those receiving untreated F1 cells. Conversely, higher levels of these autoantibodies were observed in mice tolerized with CD8+-depleted F1 cells. These results suggest that mature donor T cells are not necessary neither for the establishment of neonatal tolerance to alloantigens nor for the activation of F1 donor B cells in the production of the autoimmune syndrome in tolerant mice, but they may contribute in the regulation of the expression of autoreactive B cell clones.

Regulation of B cell tolerance by 129‐derived chromosome 1 loci in C57BL/6 mice

ObjectiveSystemic lupus erythematosus is a multifactorial disease with a strong genetic component. Previous studies have shown that a 129-derived chromosome 1 interval (Sle16) on the C57BL/6 (B6) background is sufficient to induce humoral autoimmunity. The aim of the present study was to elucidate the mechanisms by which this locus contributes to the loss of peripheral tolerance.MethodsAnti–single-stranded DNA (anti-ssDNA)–knockin transgenic mice (VH3H9R/Vκ8R and VH3H9R) were crossed with a B6 congenic line named B6.129chr1b that carries the Sle16 locus. A parallel study of a gene-targeted animal, whose mutated gene is located within the 129chr1b interval on chromosome 1, was also performed.ResultsThe combination of VH3H9R/Vκ8R with the 129chr1b interval resulted in impaired B cell anergy, and transgenic IgM and IgG anti-ssDNA antibodies were found in the circulation. The presence of IgG2aa anti-ssDNA and IgMa anti-Sm antibodies in sera indicated that the autoreactive transgenic B cells underwent class switching and epitope spreading. The 129chr1b locus appeared to have a dominant effect, since transgenic antibodies were also detected in mice carrying a single allele. The gene-targeted animals showed a similar phenotype.ConclusionThe presence of a single 129chr1b locus on the B6 background impaired B cell anergy, prevented deletion of anti-DNA transgenic B cells, and induced receptor revision. The findings of this study also emphasize that the autoimmune phenotype observed in mice with targeted genes located on chromosome 1 may simply arise from epistatic interactions between the 129 and B6 parental strains.

Anti-α-Actinin Antibodies Cross-react with Anti-ssDNA Antibodies in Active Autoimmune Hepatitis

Recently, we have observed that the association of anti-F-actin autoantibodies (Abs) with anti-alpha-actinin Abs (double reactivity) was correlated with liver disease activity in autoimmune hepatitis type 1 (AIH-1; Guéguen et al., J Clin Immunol 26:495-505, 2006). As anti-alpha-actinin Abs are also associated with anti-single-stranded (ss) DNA in 16 of 50 (32%) AIH-1 patients, compared with 16 of 401 (4.0%) liver disease control patients, we sought to characterize these Abs in AIH-1. When associated, anti-ssDNA Abs cross-react with anti-alpha-actinin Abs, but neither with anti-double-stranded DNA Abs nor with anti-F-actin Abs. Furthermore, anti-alpha-actinin Abs are associated with high-avidity anti-ssDNA Abs in AIH-1. In addition, double reactivity against alpha-actinin and ssDNA was correlated with clinical and histological activity of the disease and untreated AIH-1 patients.

C1q deficiency promotes the production of transgenic-derived IgM and IgG3 autoantibodies in anti-DNA knock-in transgenic mice

C1q-deficient mice have been shown to develop a lupus-like disease and to display an impaired clearance of apoptotic cells that are enriched in lupus autoantigens. However, the role of C1q in the regulation of autoreactive B cells remains debatable. To explore this we crossed MRL/Mp C1q-deficient mice with knock-in transgenic (Tg) mice expressing an anti-ssDNA antibody (VH3H9R and VH3H9R/VLκ8R). Analysis of the VH3H9R mice showed that in the absence of C1q higher titres of Tg-derived IgM and IgG3 anti-ssDNA antibodies were detectable. In contrast, in the VH3H9R/VLκ8R C1q-deficient animals no increase in Tg antibody levels was observed. In both models the lack of C1q induced a marked reduction of marginal zone B cells and this was paralleled by a significant increase in the percentage of plasmocytes. Thus, one could postulate that in the absence of C1q the failure to clear efficiently dying cells provides an additional stimulus to the autoreactive Tg B cells resulting in their emigration from the marginal zone B cell compartment with subsequent increase in plasmocytes. However, the lack of C1q led to an increased production of Tg IgM and IgG3 antibodies only in VH3H9R mice indicating that additional genetic susceptibility factors are required to break self-tolerance.

Highly Specific Novel Method for Isolation and Purification of Lupus Anti‐DNA Antibody via Oligo‐(dT) Magnetic Beads

A novel method for isolation and purification of anti-ssDNA antibodies from human sera is developed. The process involves: antibody purification based on their affinity for single-stranded sequence of thymidines and removal of remaining components via protein G coated magnetic beads, with high affinity for only IgG subclass. The high degree of purity and molecular weights of healthy versus lupus anti-ssDNA antibodies were confirmed by SDS-PAGE and silver staining. Western blot confirmed IgG isotype. This novel technique allows for diagnostic purposes, structural and functional analysis of anti-DNA antibodies, and studies of their role in autoimmune diseases.

A Potential Trigger of Nephritogenic Anti‐DNA Antibodies in Lupus Nephritis

Anti-DNA antibodies play an essential role in the pathogenesis of lupus nephritis. Mammalian DNA alone, however, is poorly immunogenic. We speculated that the antigenic trigger for the production of human nephritogenic anti-DNA antibodies is a non-DNA substance. The cDNA library from peripheral blood lymphocytes (PBLs) of a patient with active lupus nephritis was screened using the single-chain Fv of a human monoclonal nephritogenic O-81 anti-DNA antibody in a two-hybrid system. A clone containing the gene of an endoplasmic reticulum (ER) stress-inducible protein, Herp, was obtained: The O-81 antibody bound to recombinant Herp protein synthesized by Escherichia coli. Immunization with Herp elicited both anti-double-stranded DNA (anti-dsDNA) and anti-single-stranded (anti-ssDNA) antibodies in BALB/c mice and formed deposits of IgG in renal glomeruli. Anti-DNA antibodies purified from SLE sera bound to Herp. Moreover, anti-Herp antibodies showed specific binding to DNA. Herp was spontaneously expressed in PBLs of patients with active SLE, but not in PBLs of healthy subjects. These results imply that an inducible intracellular self-protein represents a candidate trigger for human nephritogenic anti-DNA autoantibodies. Any cell stress causing ER stress, such as viral infection, ultraviolet radiation, and chemicals, might be responsible for anti-DNA antibody production via Herp.

Structural Modeling of Sequence Specificity by an Autoantibody against Single-Stranded DNA

11F8 is a sequence-specific pathogenic anti-single-stranded (ss)DNA autoantibody isolated from a lupus prone mouse. Site-directed mutagenesis of 11F8 has shown that six binding site residues (R31VH, W33VH, L97VH, R98VH, Y100VH, and Y32VL) contribute 80% of the free energy for complex formation. Mutagenesis results along with intermolecular distances obtained from fluorescence resonance energy transfer were implemented here as restraints to model docking between 11F8 and the sequence-specific ssDNA. The model of the complex suggests that aromatic stacking and two sets of bidentate hydrogen bonds between binding site arginine residues (R31VH and R96VH) and loop nucleotides provide the molecular basis for high affinity and specificity. In part, 11F8 utilizes the same ssDNA binding motif of Y32VL, H91VL, and an aromatic residue in the third complementarity-determining region to recognize thymine-rich sequences as do two anti-ssDNA autoantibodies crystallized in complex with thymine. R31SVH is a dominant somatic mutation found in the J558 germline sequence that is implicated in 11F8 sequence specificity. A model of the mutant R31S11F8.ssDNA complex suggests that different interface contacts occur when serine replaces arginine 31 at the binding site. The modeled contacts between the R31S11F8 mutant and thymine are closely related to those observed in other anti-ssDNA binding antibodies, while we find additional contacts between 11F8 and ssDNA that involve amino acids not utilized by the other antibodies. These data-driven 11F8.ssDNA models provide testable hypotheses concerning interactions that mediate sequence specificity in 11F8 and the effects of somatic mutation on ssDNA recognition.

Repertoire diversification in mice with an IgH-locus-targeted transgene for the rearranged VH domain of a physiologically selected anti-ssDNA antibody

To test the fate of developing B cells with autoreactive receptor components, we studied mice homozygous for a knock-in transgene coding the VH domain of an IgM ssDNA-binding antibody. The transgene has unmutated C57 BL/6 V gene segments. Homozygous knock-in mice developed normal numbers of spleen and bone marrow B cells and normal serum Ig concentrations, and had the same low level of serum anti-ssDNA antibody as non-transgenic mice. Mature B cells expressed the transgene, and it underwent mutation and class switching. In young knock-in animals, nearly all IgM and some IgG cDNA clones from bone marrow and spleen contained the transgene V HD HJ H, with few or no mutations. In many IgM clones from older animals, however, and many IgG clones from both young and old mice, VH domains were revised by productive replacement with a new V HD H segment. VL segments were diverse. Immunized homozygous knock-in mice produced serum antibodies to polysaccharide, nucleic acid and protein antigens. Monoclonal IgM and IgG antibodies to nucleic acids used either transgenic or revised VH domains; but all of 20 IgG monoclonal antibodies to thyroglobulin used revised VH domain genes. Thus, B cells expressing an autoreactive (ssDNA-binding) VH domain did progress through development and were precursors for cells producing IgM and IgG, but underwent extensive VH gene revision in diversification of antibody responses.

Cyclooxygenase-2 regulates the degree of apoptosis by modulating Bcl-2 protein in pleomorphic adenoma and mucoepidermoid carcinoma of the parotid gland

ConclusionThese results suggest that COX-2 and bcl-2 protein were overexpressed and that apoptosis was reduced in MEC compared to PMA, and that COX-2 may regulate the degree of apoptosis by modulating bcl-2 protein in PMA and MEC.ObjectiveCyclooxygenase (COX)-2 plays a crucial role in tumorigenesis and overexpression of COX-2 in vitro accompanied by overexpression of bcl-2 protein has been shown to reduce apoptosis. The purpose of this study was to verify that COX-2 regulates the degree of apoptosis by modulating bcl-2 protein in benign and malignant parotid gland tumors.Material and methodsWe examined archival formalin-fixed, paraffin-embedded tissue sections of 10 pleomorphic adenomas (PMAs) and 10 mucoepidermoid carcinomas (MECs) by immunostaining with anti-COX-2, anti-bcl-2 and anti-single-stranded DNA (ssDNA) antibodies. Labeling indices of the three antibodies were calculated using computer-assisted image analysis.ResultsLabeling indices (mean±SD) of anti-COX-2 antibody in PMA and MEC were 2.05±1.30 and 11.2±2.95, respectively (p<0.001), those of anti-bcl-2 antibody were 2.00±1.28 and 9.68±4.05, respectively (p<0.001) and those of anti-ssDNA antibody were 8.06±2.54 and 2.08±1.47, respectively (p<0.001). Correlation coefficients between the labeling indices of anti-COX-2 antibody and anti-bcl-2 antibody, anti-bcl-2 antibody and anti-ssDNA antibody and anti-COX-2 antibody and anti-ssDNA antibody were 0.88, −0.75 and −0.76, respectively (p<0.001).

Genetic origin of lupus in NZB/SWR hybrids: Lessons from an intercross study

(SWR x NZB)F(1) (or SNF(1)) hybrid mice succumb to lupus nephritis. A previous analysis of SNF(1) x NZB backcross mice revealed the existence of 4 SWR loci (H2 on chromosome 17, Swrl-1 on chromosome 1, Swrl-2 on chromosome 14, and Swrl-3 on chromosome 18) and 2 NZB loci (Nba1 and Lbw2/Sbw2, both on chromosome 4). A second study focusing on SNF(1) x SWR backcross offspring uncovered 5 suggestive loci for antinuclear antibody formation, consisting of 3 dominant NZB contributions (Nba4 on chromosome 5, Lbw4 on chromosome 6, and Nba5 on chromosome 7) and 2 recessive SWR contributions (Swrl-1 on chromosome 1 and Swrl-4 on chromosome 10). The present intercross study was executed to replicate the earlier findings, using an independent panel of (SWR x NZB)F(2) offspring. A panel of (NZB x SWR)F(2) hybrids were phenotyped (for renal disease, early mortality, and a variety of autoantibodies) and genotyped (using 95 microsatellite primers positioned across all 19 autosomes and the X chromosome). Linkage analysis was conducted using the derived phenotype and genotype data, with the interval-mapping program MapManager. Four suggestive loci were mapped: Swrl-5 on chromosome 1 (peak at 106 cM), linked to hypergammaglobulinemia; an NZB locus on chromosome 5 (Nba4; peak at 15 cM), linked to IgG anti-single-stranded DNA (anti-ssDNA) antibodies, IgG anti-doubled-stranded DNA (anti-dsDNA) antibodies, and glomerulonephritis; an NZB locus on chromosome 13 (Nba6; peak at 28 cM), linked to IgG anti-dsDNA antibodies; and an SWR locus on chromosome 14 (Swrl-2; peak at 30 cM), linked to IgG anti-ssDNA antibodies. Eight additional loci revealed linkage at P < 0.01, of which 7 co-mapped with lupus susceptibility loci previously identified in other models. Considering all 3 mapping studies together, lupus in SWR/NZB hybrids appears to be the epistatic end product of several distinct loci, of which 3 SWR-derived loci (Swrl-1, Swrl-2, and Swrl-3) and 5 NZB-derived loci (Nba1, Nba3, Nba4, Nba5, and Lbw4) have been independently confirmed. The immunologic functions and molecular identities of these loci await elucidation.

Efficacy of topical tacrolimus in the treatment of various cutaneous manifestations of lupus erythematosus

Efficacy of topical tacrolimus in the treatment of various cutaneous manifestations of lupus erythematosus

Research Articles Mechanisms of Hexachlorobenzene-Induced Adverse Immune Effects in Brown Norway Rats

Hexachlorobenzene (HCB) is an environmental pollutant that induces adverse immune effects in humans and rats. Brown Norway rats (BN) appear to be very susceptible to HCB-induced immune effects. Oral exposure causes inflammatory skin and lung lesions, enlarged spleen and lymph nodes (LN) and elevated humoral responses. This review describes recent experiments that were performed to elucidate the mechanisms underlying these adverse immune effects. The metabolites tetrachlorohydroquinone and tetrachlorobenzoquinone are capable of generating neoantigen-specific T-cells in the mouse reporter antigen popliteal lymph node assay with 2,4,6-trinitrophenyl-Ficoll. Additional experiments in BN rats have shown that T-cells are involved in the development of HCB-induced skin lesions, the increase in auricular LN weight, lung eosinophilia, and humoral responses. However, splenomegaly and macrophage infiltration in the spleen and lung occur independent of T-cells. Remarkably, HCB does not induce T-cell sensitization in BN rats, suggesting that T-cells are activated nonspecifically, for example by macrophages. Transcriptome profiles obtained after subchronic HCB exposure to BN rats reveal that HCB induces a systemic inflammatory response in which several innate immune cells, such as macrophages, and pro-inflammatory cytokines are involved. Additional experiments have shown that macrophages are implicated in HCB-induced skin lesions, in particular in the aggravation of skin effects. Also, HCB-induced lung eosinophilia and elevation of anti-ssDNA IgM antibodies are less pronounced after macrophage elimination. Presumably, HCB activates macrophages, thereby generating pro-inflammatory adjuvant signals that induce a systemic inflammatory response. Subsequently, this may lead to polyclonal activation of T- and B-cells, eosinophilia, and eventually visible clinical effects.

Development of autoimmune arthritis with aging via bystander T cell activation in the mouse model of Sjögren's syndrome

A wide spectrum of extraglandular manifestations may occur in patients with Sjogren's syndrome (SS), but the mechanisms responsible for in vivo progression are still obscure. We undertook this study to evaluate the age-related changes during the development of extraglandular autoimmune lesions, including arthritis, in the murine model of primary SS, and to evaluate the possible relationship between age-related disturbance of activation-induced cell death and the in vivo kinetics against autoantigens. A total of 126 NFS/sld mice were investigated at ages 2, 4, 6, 10, 12, 18, 20, and 24 months. Cytokine production was tested using culture supernatants from anti-CD3 monoclonal antibody-stimulated T cells. Anti-single-stranded DNA (anti-ssDNA) antibodies, Ig isotypes (IgG1, IgG2a), rheumatoid factor (RF), and anti-type II collagen (anti-CII) antibodies were detected by enzyme-linked immunosorbent assay. Proliferative T cell responses against each of 3 recombinant alpha-fodrin proteins and against CII were analyzed. Autoimmune arthritis developed in SS model mice until age 24 months. Significant elevations in serum levels of RF, anti-ssDNA antibodies, and anti-CII antibodies were found in aging SS model mice. A high titer of serum autoantibodies against alpha-fodrin fragments (containing different epitopes that were originally identified in primary SS model mice) was frequently detected in young and aged SS model mice. Moreover, we found that alpha-fodrin autoantigen induced Th1 immune responses and accelerated disturbance of Fas-mediated T cell apoptosis in aged SS model mice. These results indicate that age-related disturbance of activation-induced cell death via bystander T cell activation may play a crucial role in the development of autoimmune arthritis in a murine model of SS.

Prolonged membranous lupus nephritis with change of anti-ssDNA antibody titer and repeated renal relapse

We report a case of a 44-year-old woman with nephrotic syndrome who underwent renal biopsy three times. On each occasion, light microscopy showed membranous nephropathy with mild to moderate thickening of the glomerular capillary walls. Immunofluorescence microscopy showed predominant deposition of immunoglobulin (Ig) G, IgG1, IgG2, IgG3, and IgG4; C3; and C1q along the glomerular capillary walls and deposition of IgM and IgA in some parts of the walls. Electron microscopy revealed the accumulation of electron-dense deposits in the mesangium and the subepithelial area of the glomerular basement membrane. Virus-like particles were detected in the subendothelial cells in all three biopsy specimens. A definitive diagnosis of systemic lupus erythematosus (SLE) was made at the time of the second admission, when she was 31 years old. A diagnosis of membranous lupus nephritis was then made on the basis of the pathological and clinical findings. A change in anti-single-stranded (ss)DNA antibody titers was of particular interest in this patient. Occasional small increases in anti-double-stranded (ds)DNA antibody were found, but increased anti-ssDNA antibody titers occurred before there was any elevation of urinary protein during renal relapse, and a sustained increase in the titers was shown subsequently. Hypocomplementemia occurred in parallel with the increase of anti-ssDNA antibody. Immunosuppressive therapy with steroid promptly eliminated anti-dsDNA antibody, but anti-ssDNA antibody remained positive. The patient had normocomplementemia and proteinuria was absent. Later, anti-ssDNA antibody decreased. Renal function has remained in the normal range for 20 years.

An analysis of apoptosis in lymphoid organs and lupus disease in murine systemic lupus erythematosus (SLE).

Apoptosis is a programmed cell death process that helps to regulate both T cell and B cell development. In this study, we have investigated the levels of apoptotic death in cells of the thymuses and spleens (white matter) of autoimmune MRL-lpr/lpr mice with progressive lymphadenopathy and SLE disease activity; we also examined the renal pathology in these animals. Fas is a cell surface receptor, which when activated initiates the sequence of events that lead to apoptosis. In MRL-lpr/lpr mice Fas is defective, so the competency for apoptosis may be reduced. In young animals of advancing age the thymuses enlarged until in 5-month-old females the average weight was three times that at 1 month, and spleen and kidney weights also increased in size disproportionately. At light microscope level apoptotic cells in tissue sections were counted using both routine eosin and haematoxylin staining (to identify them by their morphology) and in situ end-labelling of cells with DNA strand breaks; their presence was further confirmed by electron microscopy. As the mice aged, the numbers of apoptotic cells in thymic cortex, thymic medulla and spleen white pulp areas reduced significantly (P < 0.01-0.001), whereas in BALB/c normal controls they increased significantly (P < 0.05). These changes were coincident with the development of severe lupus, whose activity was assessed by measuring serum anti-ssDNA and anti-dsDNA antibody titres and urinary protein (albumin) level which were elevated significantly by 5 months of age (P < 0.001 for both ssDNA and dsDNA and P < 0.01 for urine albumin) compared with their younger counterparts. Thus, lymphoid organ enlargement, decrease in apoptotic indices, elevated serum anti-ssDNA and anti-dsDNA antibody levels, and impaired renal function coincided with the onset and severity of lupus disease in lpr mice. It seems likely that there is a causal relationship between defective deletion of autoreactive lymphoid cells, imperfect Fas-mediated apoptosis and development of murine SLE.