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Objective. Patients characterized with antinuclear antibodies (ANA) and disease symptoms related to one organ system can be described as having incomplete systemic lupus erythematosus (SLE). Thr aim of this multicentre study was to describe the outcome of these so-called incomplete SLE patients. Two aspects of the outcome were studied: (i) the disease course, defined by the presence or absence of clinical symptoms: and (ii) the number of patients that eventually developed full SLE. Methods. Outcome parameters were the ACR criteria, the SLE: disease Activity Index (SLEDAI), the European Consensus Lupus Activity Measure (ECLAM) and the requirement for treatment. In 10 European rheumatology centres, patients who had been evaluated in the last 3 months of 1994 and had been diagnosed as having incomplete SLE on clinical grounds for at least yr were included in the study. All 122, patients who were included in the study were evaluated annually during 3 yr of follow-up. Results. Our results are confined to a patient cohort defined by disease duration of at least yr, being under clinical care at the different centres in Europe. These patients showed disease activity that was related mostly to symptoms of the skin and the musculoskeletal system, and leucocytopenia. During the follow-up, low doses of prednisolone were still being prescribed in 43%, of the patients. On recruitment to the study, 22 of the 112 incomplete SLE patients already fulfilled the ACR criteria for the diagnosis of SLE. Tn the 3 yr of follow-up only three patients developed SLE. Conclusions, A high proportion of patients in our cohort defined on clinical grounds as having incomplete SLE eventually showed disease activity defined by the SLEDAI as well as ECLAM. However, only three cases developed to SLE during the follow-up. This suggests that incomplete SLE forms a subgroup of SLE that has a good prognosis.

Increased titres of anti-dsDNA antibodies, especially if of high avidity, are associated with renal exacerbations in patients with systemic lupus erythematosus (SLE). One of the most reliable assays to measure anti-dsDNA antibodies, the Farr assay, is believed to detect preferentially high avidity antibodies. Purified non-complexed monoclonal antibodies (mAbs) against nucleosomes, obtained from mice with SLE, are not reactive in the Farr assay, but can become so once complexed to nucleosomes, These Farr-positive, nucleosome containing, immune complexes were also able to bind in vivo to the glomerular basement membrane (GBM), predominantly via heparan sulphate (HS). To evaluate whether in SLE patients the same kind of immune complexes are responsible for Parr reactivity, IgG from serum or plasma was isolated under dissociating and physiological conditions. We observed that after purification under dissociating conditions, Farr reactivity was significantly decreased (P

Objective. Most information available about the disease course of patients with systemic lupus erythematosus (SLE) is restricted to the first 5 yr after disease onset. Data about the disease course 10 yr after disease onset are rare. The aim of this multicentre study was to describe the outcome of SLE patients with a disease duration of >10 yr. Methods. Outcome parameters were the SLE Disease Activity Index (SLEDAI); the European Consensus Lupus Activity Measure (ECLAM), the Systemic Lupus International Collaborative Clinics/American College of Rheumatology Damage Index (SLICC/ACR), a global damage index (DI) and required treatment. In 10 different European rheumatology centres, all SLE patients who were evaluated in the last 3 months of 1994, and who had been diagnosed with SLE at least 10 yr ago, were included in the study. Results. It should be stressed that our results are confined to a patient cohort, defined by a disease duration of at least 10 yr, and who are still under clinical care at the different centres in Europe. These SLE patients still showed some disease activity, related to symptoms of the skin and musculoskeletal systems, next to the presence of renal involvement. A total of 72% of the patients needed treatment with prednisolone (less than or equal to 7.5 mg). The cumulative damage was overall related to clinical features of the central nervous system (14%) and renal involvement (14%), next to deforming arthritis (14%), osteoporosis (15%) and hypertension (40%). The prevalences of obesity, Cushing appearance and diabetes are highly suggestive that the ongoing treatment and that in the past might have had an impact on the total sum of endorgan damage. Conclusions. After 10 yr, a high proportion of patients in our cohort continued to show evidence of active disease, defined by the SLEDAI as well as ECLAM. The DI was related to the involvement of the central nervous system, renal involvement and the presence of hypertension.

Objective. We related soluble Fas (sFas) levels to the Systemic Lupus Erythematosus Disease Activity Index (SLEDAI) in a longitudinal series of plasma samples of patients with SLE to evaluate the relation between excessive production of sFas and disease activity. Methods. We generated 21 monoclonal antibodies against Fas. Two Of these were used to develop and validate a sensitive sandwich ELISA for the longitudinal analysis of sFas levels in plasma of 30 patients and 25 controls. Results. At the start of followup, a significant elevation (p

It has been suggested that binding of anti-double-stranded DNA antibodies to cell surfaces, followed by internalization and nuclear binding (so called in vivo ANA) is of pathophysiological significance for tissue damage in systemic lupus erythematosus. We have shown before that pathogenic antinuclear antibodies complexed to nucleosomal antigens can bind to heparan sulfate in the glomerular basement membrane in vivo. Because nucleosomes are also reported to bind to the cell surface, we hypothesized that in vivo ANA is a property of antinuclear antibodies bound to nucleosomal antigens. Therefore, we studied three antinucleosome monoclonal antibodies (mAb) that exhibit in vivo ANA as seen by immunofluorescence in mice inoculated intraperitoneally with the hybridoma producing the mAb. The same mAb complexed to nucleosomal antigens after intravenous injection into mice induced in vivo ANA, in contrast to purified noncomplexed mAb. To study this In more detail, we incubated complexed mAb with various cell lines and found binding to the cell surface and subsequent internalization into cytoplasmic vesicles. However, no binding to the nucleus was observed by immunoelectron microscopy (IEM) and confocal laser microscopy. Noncomplexed mAb did not bind to the cell surface, Next, from mice bearing the hybridomas producing the mAb intraperitoneally, a small part of the kidney was snap frozen in liquid N-2, fixed with acetone, and studied In immunofluorescence, whereas the remaining part of the kidney was fixed In vivo by renal perfusion with a mixture of 0.01 M sodium periodate, 0.075 M lysine HCI, 0.0375 M Na2HPO4, and 2% paraformaldehyde (PLP) and studied in both immunofluorescence and IEM. In the acetone-fixed kidney sections obtained without in vivo fixation we again observed in vivo ANA. However, after in vivo PLP perfusion fixation, no nuclear binding was found. in IEM, localization in cytoplasmic vesicles was seen. in conclusion, antinucleosome antibodies complexed to nucleosomal antigens can bind to the cell surface and are transported into the cytoplasm, but do not bind to the nucleus. The reported nuclear localization of antinuclear antibodies is caused by a fixation artifact.

The presence of anti-heparan sulphate (HS) reactivity in serum is closely related to the occurrence of nephritis in patients with systemic lupus erythematosus (SLE). Since patients with lupus nephritis in general also have high titres of anti-DNA antibodies, we wanted to clarify the relationship between anti-HS and anti-DNA reactivity in serum. Therefore, we studied longitudinally six patients with lupus nephritis who experienced 12 exacerbations of their disease, and five SLE patients without nephritis experiencing 10 periods of non-renal disease exacerbations. In addition, we tested single serum samples of another 24 patients obtained during a renal disease exacerbation and 22 sera of patients without nephritis. The sera of all patients were tested for anti-DNA (Farr assay) and anti-HS reactivity (ELISA). We confirmed that SLE patients during renal exacerbations have a significantly higher anti-HS reactivity than patients without nephritis (P

Lupus nephritis is regarded as an immune complex mediated disease. Since anti-DNA antibodies are present in the circulation and in diseased glomeruli of patients with lupus nephritis, these antibodies have been assigned a pivotal role in the initiation of lupus nephritis. It remains however unclear how these antibodies become localized in the glomerulus. Contrary td the classical concept of glomerular deposition of DNA/anti-DNA complexes, it has been suggested that anti-DNA antibodies can interact with intrinsic glomerular antigens. Some anti-DNA antibodies can cross-react with heparan sulphate (HS), which is such an intrinsic constituent of the glomerular basement membrane (GBM). Serum HS reactivity coincides with the occurrence of lupus nephritis. It was found that this HS reactivity was exhibited by anti-DNA antibodies complexed to nucleosomes and not by the antibody itself. Nucleosomes are DNA/histone complexes, present in the nucleus, which are released by dying cells. The histone part of the nucleosome is responsible for the binding to the GBM. Recently, it has become clear that also anti-nucleosome antibodies can bind to HS in the GBM via nucleosomes. These nucleosome-containing immune complexes exhibit anti-DNA reactivity in ELISA and Farr assay. It is now thought that nucleosomes released by dying cells bind to anti-DNA or anti-nucleosome antibodies in the circulation, giving rise to nephritogenic immune complexes. Alternatively, nucleosomes may bind to the GBM and serve then as planted antigen for subsequent binding of antibodies via an in situ mechanism. Binding of antibodies via both mechanisms leads to complement activation and damage of the GBM. The recent finding of histones and DNA in glomerular depositions in lupus nephritis is in line with this hypothesis.

Autoantibodies reacting with a great variety of autoantigens are characteristic for the autoimmune disease systemic lupus erythematosus (SLE). Although reactivity with heparan sulfate (HS) in sera of patients with SLE is found in association with the occurrence of nephritis, the aetiological significance of this association is not clear. The assay which is generally used to measure anti-HS reactivity is subject to false-positive results, as a consequence of the binding of negatively charged moieties within immune complexes to the precoat employed (protamine sulfate). Therefore, we have developed a new ELISA in which photobiotinylated HS is efficiently and reproducibly bound to streptavidin-coated wells. We compared the new ELISA with the classical anti-HS ELISA by testing culture supernatants of 20 murine monoclonal antibodies (mAb) to DNA (containing free anti-DNA and anti-DNA/nucleosome immune complexes) and preparations of these mAb (containing only free anti-DNA), purified under dissociating conditions. In the classical anti-HS ELISA, 14 out of 20 of the culture supernatants reacted positively with HS; after purification no reactivity remained. The discrepancy must be due to anti-DNA/nucleosome immune complexes present in the culture supernatants. In the new ELISA only four out of 20 culture supernatants and one of the purified preparations reacted with HS. This latter reactivity is probably not specific, since this mAb also reacted with streptavidin alone. To find out whether there is a correlation between the occurrence of nephritis and anti-HS reactivity, measured in this new anti-HS ELISA, we tested sera of patients with a renal- or non-renal exacerbation of SLE in the newly developed anti-HS ELISA. We observed a correlation between anti-HS reactivity and nephritis.

Histones can mediate the binding of DNA and anti-DNA to the glomerular basement membrane (GBM). Zn ELISA histone/DNA/anti-DNA complexes are able to bind to heparan sulfate (HS), an intrinsic constituent of the GBM. We questioned whether histone containing immune complexes are able to bind to the GBM, and if so, whether the ligand in the GBM is HS. Monoclonal antibodies (mAbs) complexed to-nucleosomal antigens and noncomplexed mAbs were isolated from culture supernatants of four IgG anti-nuclear mAbs. All noncomplexed mAbs showed strong anti-nucleosome reactivity in ELISA. One of them showed in addition anti-DNA reactivity in noncomplexed form. The other three mAbs only showed anti-DNA reactivity when they were complexed to nucleosomal antigens. After renal perfusion a fine granular binding of complexed mAbs to the glomerular capillary mall and activation of complement was observed in immunofluorescence, whereas noncomplexed mAbs did not bind. Immuno-electron microscopy showed binding of complexes to the whole width of the GBM. When HS in the GBM was removed by renal heparinase perfusion the binding of complexed mAb decreased, but did not disappear completely. We conclude that anti-nucleosome mAbs, which do not bind DNA, become DNA reactive once complexed to nucleosomal antigens. These complexed mAbs can bind to the GBM. The binding ligand in the GBM is partly, but not solely, HS. Binding to the GBM of immune complexes containing nucleosomal material might be an important event in the pathogenesis of lupus nephritis.