Exposure to nuclear antigens contributes to the induction of humoral autoimmunity during tumour necrosis factor alpha blockade.

OBJECTIVE: Type I interferons and apoptotic particles contribute to antinuclear autoimmunity in experimental models. This study assessed whether similar mechanisms contribute to break peripheral B-cell tolerance in humans by studying the induction of antinuclear antibodies by tumour necrosis factor blockade in spondyloarthritis. METHODS: 40 spondyloarthritis patients treated with infliximab or etanercept and 20 renal cell carcinoma patients treated with sorafenib were studied. Serum antinucleosome IgM and nucleosomes were measured by ELISA. Type I interferon serum activity was measured using a functional reporter cell assay. Synovial apoptosis was assessed by terminal transferase nick end-labelling (TUNEL) assay and anti-active caspase-3 immunostaining. Complement was measured by nephelometry. RESULTS: Despite a similar clinical improvement and reduction of synovial inflammation, antinucleosome IgM were induced by infliximab but not etanercept. This induction did not correlate with type I interferon activity, which was transiently downmodulated by infliximab but persistently upregulated by etanercept. In contrast, antinucleosome IgM levels did correlate with serum nucleosome levels, which were significantly upregulated by infliximab but not by etanercept treatment. This increase in serum nucleosome levels was not directly related to massive cell death, but rather to a decrease of complement 3 and 4 serum levels during infliximab treatment. CONCLUSION: Infliximab and etanercept have a differential effect on both type I interferon activity and nucleosome levels. Only elevated serum nucleosomes relate to the induction of antinucleosome antibodies after infliximab treatment.

Melanoma inhibitory activity, a biomarker related to chondrocyte anabolism, is reversibly suppressed by proinflammatory cytokines in rheumatoid arthritis.

OBJECTIVE: In mice, melanoma inhibitory activity (MIA) is a chondrocyte-specific molecule with similar regulation to collagen type II. As MIA is a small secreted protein, its value as cartilage biomarker in human inflammatory arthritis was assessed. METHODS: MIA tissue distribution was studied by quantitative PCR and immunohistochemistry. The regulation of MIA production was studied in vivo in rheumatoid arthritis (RA) (n = 37) and spondyloarthritis (SpA) (n = 30) synovial fluid (SF), and in vitro in alginate embedded human chondrocytes. Therapeutic modulation of serum MIA was evaluated during tumour necrosis factor (TNF)alpha and interleukin (IL)1 blockade in RA. RESULTS: MIA was primarily expressed by chondrocytes in the human joint. SF MIA levels were lower in RA than in SpA despite similar levels of overall synovial inflammation. Further analysis indicated that these levels were inversely correlated with the degree of joint inflammation in RA, but not in SpA, and that the levels of TNFalpha and IL1beta were significantly increased in RA versus SpA. Accordingly, these proinflammatory cytokines suppressed MIA mRNA and protein in cultured chondrocytes. This suppression was paralleled by suppression of cartilage anabolism as assessed by collagen type 2 and aggrecan mRNA. Treatment of patients with RA with TNF blockade or IL1 blockade induced an increase of serum MIA levels. CONCLUSION: The decreased levels of MIA in the inflamed RA joint and the coregulation of MIA and cartilage matrix molecules by proinflammatory cytokines indicate that joint inflammation in RA not only drives accelerated cartilage degradation but also suppresses cartilage anabolism. This inflammation-driven suppression is reversible in vivo.

Anti-citrullinated protein/peptide antibodies (ACPA) in rheumatoid arthritis: specificity and relation with rheumatoid factor.

Anti-citrullinated protein/peptide antibodies (ACPA) are highly specific and sensitive markers for rheumatoid arthritis (RA). For instance, for the anti-CCP2 assay, sensitivities ranging from 55% to 80% and specificities ranging from 90% to 98% have been reported. Despite their high specificity, recent reports have suggested that ACPA may be found in some patients with other rheumatic autoimmune diseases, including psoriatic arthritis, systemic lupus erythematosus and Sjögren's syndrome. Also, the differences between the classical rheumatoid factor (RF) and ACPA, as well as the complementarity between both tests have recently been demonstrated more clearly. Indeed, both antibody systems have a different association with specific RA features like extra-articular manifestations, a different association with the HLA shared epitope and, behave differently following anti-TNF therapy.

Chemical agents and enzymes used for the extraction of gut lymphocytes influence flow cytometric detection of T cell surface markers.

Chemical agents (DTT, EDTA) and enzymes (collagenase, DNAse) are often used for the generation of a single cell suspension from tissue samples. In this study, flow cytometry was used to examine the effect of chemical agents and enzymes on the expression of cell membrane markers of T lymphocytes from tonsils and peripheral blood. Expression of CD4, CD8, CD25, CD38, L-selectin, CD44, alphaEbeta7 and alpha4beta7 were studied. Incubation of lymphocytes with DTT and EDTA resulted in a decrease of CD38, alphaEbeta7 and alpha4beta7 expression. Incubation with collagenase A and DNAse resulted in a decrease of CD25, L-selectin, alphaEbeta7 and alpha4beta7. The results of this study indicate that a careful interpretation is necessary for phenotypic descriptions of lymphocyte populations obtained by enzymatic isolation techniques.

Focal up-regulation of E-cadherin-catenin complex in inflamed bowel mucosa but reduced expression in ulcer-associated cell lineage.

The E-cadherin-catenin complex is important for the maintenance of epithelial architecture. We studied its expression in Crohn disease, ulcerative colitis, acute ileitis, and controls. Immunohistochemical stainings for E-cadherin, alpha-catenin, beta-catenin and gamma-catenin were performed. E-cadherin messenger RNA (mRNA) was detected using riboprobes. In active inflammation, there was up-regulation of the complex. In particular, epithelium adjacent to ulcers showed increased expression of protein and mRNA, but in ulcer-associated cell lineage, the intensity of staining was weak to negative. In focal inflammation, up-regulation was found in affected areas. Reparative epithelium growing over denuded areas showed weaker expression. Since structural or functional perturbation in any of the molecules of the E-cadherin-catenin complex results in loss of intercellular adhesion, the preexistent epithelium may benefit from up-regulation to try to maintain its normal architecture under inflammatory conditions. Reduced expression in reparative epithelium and ulcer-associated cell lineage could facilitate the motility of these cells.

Bowel inflammation and the spondyloarthropathies.

The concept of spondyloarthropathies gathers together a group of chronic diseases in which not only the locomotor system is involved but also other organs, especially the gastrointestinal tract. In humans, ileocolonoscopic studies demonstrated the presence of inflammatory gut lesions in all the diseases in the spondyloarthropathy group; their presence varied in the different diseases between 20% and 70%. The inflammation could be related to specific disease features in the spondyloarthropathies. Further research supports the hypothesis of subclinical inflammatory bowel disease in some patients with spondyloarthropathy, in which the locomotor inflammation was the only clinical manifestation. The link between gut inflammation and arthropathy has also been demonstrated in animal models, notably the human leukocyte antigen B27 transgenic rats. The temporal relationship between activity and severity of colonic involvement and flares of peripheral arthritis directs treatment of choice. For all forms of enterogenic arthropathies, nonsteroidal anti-inflammatory drugs remain the acute treatment form. Caution is in order, however, because of their possible harmful effects on intestinal integrity, permeability, and even on gut inflammation.

Short term variation of human immunoglobulin levels with an estimation of the day to day physiological variability.

In this work serum Ig levels were determined daily for 14 days in 12 subjects, every two days for 14 days in 19 subjects, every week for 1 month in 11 subjects and fortnightly in 11 subjects for 6 months. The serum IgG, IgM and IgA levels were quantitated by the linear plate method. The day-to-day reproducibility of the technique was checked for a period of one months on seven different samples. The mean coefficient of variation in the reproducibility experiments was found to be 4.8% for IgG, 8.2% for IgM and 5.2% for IgA. Since the observed variation in all groups of subjects was found to be higher than the methodological variation, it appears that physiological variability occurs in the serum Ig levels of an individual. No difference was found between the physiological variances of IgG, IgM and IgA observed in the group with daily analysis and those with determinations every two days. The physiological variations increase when longer periods are considered between consecutive determinations, but the variability of IgG and IgA in the group with fortnightly determinations is less important than that observed in the group with weekly analyses. Some evidence is presented indicating a periodic fluctuation of serum IgG and IgA levels with an amplitude of nearly 20% and with a recurrence rate of two weeks.

Intra-articular injection pentosanpolysulphate results in increased hyaluronan molecular weight in joint fluid.

The influence of an oversulphated glycosaminoglycan, pentosanpolysulphate, on hyaluronan metabolism of the synovial lining cell was studied in vivo in human volunteers. Significant increases in the mean degree of polymerisation of the hyaluronan chains were observed after a series of four to six intra-articular injections of this glycosaminoglycan. No increases in hyaluronan synthesis rates were observed. Repeated administration of the drug did not cause any inflammation or bleeding in the joint cavity.

Pigmented villonodular synovitis of the hip--association with osteochondromatosis.

Pigmented villonodular synovitis, diffuse or nodular in nature, is most commonly seen in the knee but occasionally involves the hip. The authors reviewed the medical records of seven patients with a pathological diagnosis of unilateral pigmented villonodular synovitis of the hip. Characteristic findings were proliferation of the synovial lining cells and subsynovial invasion of fibroblast or polyhedral mesenchymal cells in a nodular pattern. Multiple initiating factors have been advanced to account for the occurrence of this monoarticular chronic arthritis, but the etiology and pathogenesis still remain obscure. On three occasions the authors found an association with free floating osteochondromas and well-delineated foci of metaplastic cartilage under the synovial lining cells. Two benign lesions may occur simultaneously and are possibly related entities involving the synovial membranes as a reaction to a common etiologic factor. Although the histopathology of pigmented villonodular synovitis of the hip and the knee is identical, localization in the former joint has its typical clinical, radiological, differential diagnostic and therapeutic aspects.

Goldsalts, levamisole and D-penicillamine as first choice slow-acting antirheumatic drugs in rheumatoid arthritis--a long-term follow-up study.

In the present study, 345 rheumatoid arthritis patients were treated using goldsalts, D-Penicillamine or levamisole as the slow-acting antirheumatic drug of first choice. Goldsalts were given to 182 patients, levamisole to 139 and D-Penicillamine to 24. At the time of the present evaluation, 83 patients were still on goldsalts (44.6%), 63 on levamisole (45.2%) and 11 on D-Penicillamine (45.9%). Adverse reactions required interruption of treatment in 64 patients on goldsalts (35.2%), in 44 on levamisole (31.7%) and in 5 on D-Penicillamine (20.8%). Inefficacy was responsible for withdrawal of 33 patients receiving goldsalts (18.1%), 30 receiving levamisole (21.6%) and 8 receiving D-Penicillamine (33.3%). The duration of treatment was 4.6 years for goldsalts, 3.6 years for levamisole and 3.6 years for D-Penicillamine. In the present analysis none of the compounds was found to have a definite advantage over the others. The rather favourable treatment continuation rates in this study can be attributed to the fact that the slow-acting antirheumatic drugs were given at an early stage of the disease, preferably before the occurrence of radiological lesions.

Reparative response of human articular cartilage in tissue culture. Comparison between a normal and an osteoarthritic knee of the same donor.

Some evidence is presented that the metabolism of visually intact cartilage taken from a knee with focal osteoarthritic lesions, may already be affected. Proteoglycan metabolism of articular cartilage from the macroscopically unaffected right knee and the osteoarthritic left knee of the same donor was assayed in long-term tissue culture. A good reparative response was found in the right knee with an increase in the relative amount of radiolabelled proteoglycan and proteoglycan aggregates, and in the total amount of labelled proteoglycan per mg dry tissue. No significant differences were found when the anteromedial, anterolateral and posterolateral areas were compared. Visually intact samples taken from the left knee, with a deep osteoarthritic cartilage erosion on the medial condyle, showed no accumulation of radiolabel into proteoglycan and proteoglycan aggregates for the enzymatically pretreated samples. This was interpreted as a repair failure.

Osteomyelitis in a patient with a chronically infected arterial graft and longstanding hypertrophic osteoarthropathy of the leg.

A patient is reported with hypertrophic osteoarthropathy of the right leg who developed an osteomyelitis; this appeared to be related to an infected aortic bifurcation prosthesis with increased uptake on leukocyte scan and chronic gas formation at the proximal aortic anastomosis on computed tomography. There have been previous reports of hypertrophic osteoarthropathy as a sign of arterial prosthesis infection. In this patient antibiotic therapy was successful in controlling the osteomyelitis but eradication of the arterial prosthesis infection could not be confirmed and an elective arterial operation involved too great a risk.

Subclinical involvement of the gut in undifferentiated spondylarthropathies.

In order to evaluate the frequency of subclinical gut involvement in the seronegative spondylarthropathies, ileocolonoscopy with biopsies of the colon, ileocecal valve and ileum were performed on 211 patients with ankylosing spondylitis (AS), reactive arthritis (ReA) and undifferentiated spondylarthropathies. Inflammatory gut lesions were detected in a large number of these patients. It was concluded that the group of undifferentiated spondylarthropathies could be split into fairly equal subgroups: one subgroup suffering from subclinical inflammatory bowel disease associated with peripheral joint symptoms, a second subgroup presenting a form of enterogenic ReA, and a third subgroup in which no relation with the gut could be demonstrated. Ankylosing spondylitis, which has to be considered as a form of undifferentiated seronegative spondylarthropathy, could be subdivided into the same subgroups. These findings confirm the existence of subclinical gut involvement in patients with seronegative spondylarthropathies.

The influence of hydrocortisone on aggrecan metabolism in human articular chondrocyte cultures: comparison of two different matrices.

OBJECTIVE: Numerous reports of negative effects as well as protective effects of glucocorticoids on articular cartilage convinced us to study the influence of hydrocortisone on aggrecan synthesis of isolated phenotypically stable human articular chondrocytes cultured in two different matrices. METHODS: Macroscopically normal human articular cartilage was obtained from femoral condyles within 24 hours postmortem. Chondrocytes were isolated and cultured in gelled agarose or in alginate. After 14 days in culture, hydrocortisone was added for 5 days at concentrations ranging from 0.005 microgram to 1 mg/ml for the agarose cultures and from 0.005 microgram to 1 microgram/ml for the alginate culture system. Aggrecan synthesis was measured by the incorporation of 35Sulphate, and the proportion of neosynthesized aggrecan that bound to hyaluronan to form aggrecan aggregates was analyzed by gel chromatography. RESULTS: At concentrations from 0.005 to 1 microgram/ml, hydrocortisone was found to produce a similar dose-dependent stimulation of aggrecan synthesis in both matrices. The synthesis of aggrecans remained at the same level for concentrations of 1 microgram/ml up to 100 micrograms/ml of hydrocortisone. When supraphysiological concentrations of hydrocortisone were added the aggrecan synthesis rate plateau declined. Simultaneously with the increase in aggrecan synthesis, the proportion of low-molecular weight 35S-proteoglycans decreased in favour of 35S-aggrecan aggregates and monomers in the agarose system. The chondrocytes cultured in alginate showed this increase of aggrecan aggregates and monomeric aggrecans in both the cell-associated and the inter-territorial matrix. CONCLUSION: Hydrocortisone is a stimulator of aggrecan synthesis by normal human articular chondrocytes cultured in vitro. The two culture systems (agarose and alginate) tested in this experiment showed a comparable aggrecan synthesis rate, increasing under the influence of hydrocortisone at concentrations up to 1 microgram/ml. The proportions of 35Sulphate incorporated in aggrecan aggregates and monomeric aggrecan were also higher under the influence of hydrocortisone.

Influence of the gut and cytokine patterns in spondyloarthropathy.

Clinical and histological studies have previously shown that spondyloarthropathy (SpA) patients can have subclinical gut inflammation. This gut inflammation is related to enterocolitis in Crohn's disease (CD) and may evolve to overt inflammatory bowel disease in a subset of these patients. Moreover, there is an intriguing clinical link between gut inflammation and peripheral joint inflammation. In order to explore immunologically these concepts, recent studies have characterized phenotypically and functionally the inflammatory cells in both the gut and the synovium of SpA patients and have provided a number of new insights. Firstly, they confirm histological and pre-histological alterations of the gut immune system in SpA, which are redundant of CD and which are linked to alterations of the peripheral joints. Secondly, both the acquired and the innate immune system contribute to these alterations, with an important role for both T cells and macrophages and their cytokines. Thirdly, interpretation of these data support the hypothesis that gut and joint inflammation in SpA are induced by the combination of an impaired anti-bacterial host defence and an uncontrolled pro-inflammatory response of the innate immune system. The insights provided by the study of the gut immunology in SpA have contributed to develop new therapeutic strategies, with TNFalpha blockade as prototype.

Influence of polysulphated polysaccharides and hydrocortisone on the extracellular matrix metabolism of human articular chondrocytes in vitro.

OBJECTIVE: To evaluate the influence of hydrocortisone and two polysulphated polysaccharides (xylosan polysulphate and chondroitin polysulphate) on the extracellular matrix metabolism of chondrocytes cultured in gelled agarose. METHODS: Isolated chondrocytes from normal femoral cartilage of the knee joints of 7 donors were cultured in gelled agarose to maintain their differentiated phenotype. After two weeks of culture, hydrocortisone (0.2 microgram/ml), xylosan polysulphate (10 micrograms/ml) and chondroitin polysulphate (10 micrograms/ml) were added to the culture media supplemented with or without interleukin (IL)-1 beta. After one week of incubation, the cells were liberated from the agarose with agarase. Isolated cells were labelled with antibodies against aggrecan and type II collagen, as well as biotinylated hyaluronic acid binding protein to analyse the extracellular matrix (ECM) molecules in the cell-associated matrix (CAM). The levels of matrix metalloproteinase (MMP)-1, -3, and -13, as well as tissue inhibitor of metalloproteinase (TIMP)-1 and -3 were determined after the cells had been permeabilised and stained with the appropriate antibodies. Triplicate samples were analysed with flow cytometry. RESULTS: IL-1 beta decreased the accumulation of aggrecan, hyaluronan and type II collagen in the CAM and increased intracellular MMP-1, -3 and -13 at a concentration of 100 pg/ml. Xylosan polysulphate and chrondroitin polysulphate restored the expression of these CAM molecules in these IL-1 beta-treated cultures. Hydrocortisone stimulated the accumulation of CAM aggrecan and hyaluronan whether or not under the exposure to IL-1 beta. Intracellular MMP-1, -3, -13 and TIMP-1 and -3 of IL-1 beta-treated cells was downregulated after treatment with hydrocortisone. CONCLUSION: Both hydrocortisone and the two polysulphated polysaccharides could stimulate the accumulation of CAM macromolecules of IL-1 beta-treated chondrocytes. This effect probably resulted in part from the downregulation of MMPs. These agents showed cartilage structure modifying effects in vitro.

Churg-Strauss syndrome presenting as polymyositis.

Churg-Strauss syndrome is a vasculitis-based pathology, predominantly characterized by asthma and eosinophilia. Histopathologically, the vessel wall infiltration contains a substantial proportion of eosinophils and extravascular granulomata can be demonstrated. We report a case of Churg-Strauss syndrome presenting as polymyositis.

Etiological factors and underlying conditions in patients with leucocytoclastic vasculitis.

This study concerns a retrospective analysis of 63 consecutive patients presenting with leukocytoclastic vasculitis at the Departments of Dermatology or Rheumatology of the University Hospital Ghent (Belgium) (period 1988-1993). The diagnosis of leukocytoclastic vasculitis was confirmed by histopathology in all cases. All patients were screened for underlying causes, including drugs, infection, systemic autoimmune disease or neoplasia. In 34 patients, an etiological factor was identified: drugs (5 patients), infection (6 patients), drugs or infection (4 patients), systemic autoimmune disease (10 patients), Henoch Shönlein (6 patients), neoplasia (2 patients) and cryoglobulinemia (1 patient). In the group of patients with leukocytoclastic vasculitis in the context of systemic autoimmune disease, 4 patients suffered from systemic lupus erythematosus, 2 from Wegener's disease, 2 from Behçet's disease, 1 from polyarteritis nodosa and 1 from rheumatoid arthritis. In the remaining 29 patients, no cause for the vasculitis could be identified.

Destructive lesions of small joints in seronegative spondylarthropathies: relation to gut inflammation.

Destructive lesions of small joints were found in 40 out of 211 patients suffering from seronegative spondylarthropathies (SpA) on whom ileocolonoscopy with biopsies of the ileum and colon were performed. The destructive lesions of small joints, radiologically only distinguishable from rheumatoid arthritis lesions by the pauciarticular and asymmetrical involvement, the rare tendency to fusion and the rare occurrence of periosteal hypertrophy, were observed more frequently in patients presenting subclinical inflammatory gut lesions, predominantly of the chronic type, than in patients without gut inflammation.

Broadening of the T cell receptor spectrum among rheumatoid arthritis synovial cell-lines in relation to disease duration.

OBJECTIVE: The aim of the study was to evaluate the T cell receptor (TCR) family usage in T cell-lines from subcutaneous nodules and synovium from patients with rheumatoid arthritis (RA), with specific reference to the duration of symptoms. In vitro adherence characteristics of nodular T cells was studied as well. METHODS: Monoclonal antibodies were used to determine the distribution of TCR families in T cell-lines from synovium of patients with early and long-standing RA, from rheumatoid nodules and control tissues. An in vitro binding assay with T cell-lines from 2 rheumatoid nodules was performed. RESULTS: In early RA synovium, a restricted TCR family usage was observed in 5 out of 8 patients, contrary to long-standing disease, peripheral blood, ileum and colon. In RA nodules, a similar degree of restriction was noted. Moreover, the same TCR family was overexpressed by T cell-lines from different nodules derived from the same patient. T cell-lines from rheumatoid nodules demonstrated a preferential in vitro adherence to rheumatoid synovium and rheumatoid nodules, while no binding was observed on skin or tonsil. CONCLUSION: The TCR spectrum among RA synovial cell-lines broadens in relation to the disease duration. The overexpression of the same TCR family in different rheumatoid nodules from the same patients, and the in vitro adherence of T cell-lines from rheumatoid nodules may be indicative for recirculation between the different disease manifestations in RA.