Identification and expression of iron regulators in human synovium: evidence for upregulation in haemophilic arthropathy compared to rheumatoid arthritis, osteoarthritis, and healthy controls.

Recurrent joint bleeding is the most common manifestation of severe haemophilia resulting in haemophilic arthropathy (HA). Iron plays a central role in the pathogenesis of the two main features of HA: synovitis and cartilage destruction. The aim of this study was to investigate the synovial presence of the iron regulator proteins ferroportin (FPN), hepcidin, haemoglobin scavenger receptor CD163 (CD163), feline leukaemia virus subgroup C (FLVCR), and heme carrier protein 1 (HCP-1). A comparison of the expression in HA with rheumatoid arthritis (RA), osteoarthritis (OA), and healthy controls (HC) is made. Synovial expression of iron regulators was investigated by immunohistochemistry in human synovial tissue and in a murine haemophilia model. We demonstrate for the first time the synovial presence of the investigated iron regulator proteins. Expression of the iron regulator proteins FPN, CD163, FLVCR, and HCP-1 was enhanced in HA in comparison to RA, OA, and HC synovium. In addition, in a murine haemophilia model of acute joint bleeding, synovial expression of FPN, CD163, and HCP-1 was increased. In both human and murine experiment, synovial expression of hepcidin was not altered. These findings indicate the presence of iron regulator proteins in the synovium, demonstrate an enhanced expression of FPN, CD163, FLVCR, and HCP-1 in HA, and suggest a synovial adaptation mechanism to maintain synovial iron homeostasis in HA.

IL-4 alone and in combination with IL-10 protects against blood-induced cartilage damage.

OBJECTIVE: It has been reported that interleukin (IL)-10 limits blood-induced cartilage damage. Our aim was to study the effect of IL-4 alone and in combination with IL-10 on blood-induced cartilage damage. DESIGN: Healthy human full thickness cartilage explants were cultured for 4 days in the presence of 50% v/v blood. IL-4, IL-10, or a combination of both cytokines was added during blood exposure. Cartilage matrix turnover was determined after a recovery period; additionally cytokine production, chondrocyte apoptosis, and expression of the IL-4 and IL-10 receptors were analyzed directly after exposure. RESULTS: Blood-induced damage to the cartilage matrix was limited by IL-4 in a dose-dependent way (P<0.05). Also IL-10 limited this damage, although to a lesser extent (P<0.03). The effect of IL-4 plus IL-10 was more pronounced and protective than IL-10 alone (P<0.05). Production of IL-1ß and tumor necrosis factor (TNF)-α was limited by both IL-4 and IL-10 (P<0.05), but more strongly by IL-4. Blood-induced apoptosis of chondrocytes was limited by IL-4 and the combination, and not by IL-10 alone. No direct beneficial effect of IL-4 or IL-10 on cartilage was found, however, the chondrocyte receptor expression of both cytokine receptors was upregulated by exposure to blood. CONCLUSIONS: This study demonstrates that IL-4 alone and in combination with IL-10 prevents blood-induced cartilage damage. Expectedly, anti-inflammatory effects on monocytes in the blood fraction and protective effects on chondrocytes are both involved. IL-4 in combination with IL-10 might be used to prevent blood-induced joint damage as a result of trauma or surgery.

Very rapid clearance after a joint bleed in the canine knee cannot prevent adverse effects on cartilage and synovial tissue.

OBJECTIVE: Joint bleeding leads to joint destruction. In vitro exposure of human and canine cartilage to blood results in long-lasting severe adverse changes in cartilage. An in vivo joint haemorrhage in the canine knee joint demonstrates similar adverse effects although significantly less outspoken. As a possible explanation for this discrepancy, we studied the clearance rate of blood from the canine knee joints. METHODS: Blood was injected into the knee joint of Beagle dogs either 48 h, 24h or 15 min before termination. The amount of red blood cells (RBC) and white blood cells (WBCs) present in the joint cavity was determined. Chondrocyte activity and cartilage matrix integrity as well as cartilage destructive activity of synovial tissue were determined biochemically. Additionally, synovial tissue was analyzed by use of histochemistry. RESULTS: The amount of blood was decreased to <5% within 48 h. Within this time period the cartilage was negatively affected and the synovial tissue showed cartilage destructive activity. Evaluation of the synovial tissue 15 min post-injection revealed countless numbers of intact RBC that were almost completely disappeared after 48 h without significant recruitment of macrophages. CONCLUSIONS: Blood is cleared very rapidly from the canine knee joint, but already has adverse effects on both cartilage and synovial tissue within that short time span. This rapid clearance can play a role in the discrepancy between long-term in vitro and in vivo effects of blood-induced joint damage since more than 10% v/v blood present for at least 48 h is needed to induce long-term adverse effects in vitro.

Pretreatment macrophage infiltration of the synovium predicts the clinical effect of both radiation synovectomy and intra-articular glucocorticoids.

OBJECTIVE: To explore whether pretreatment features of synovial tissue in patients with gonarthritis could predict the clinical effect of radiation synovectomy with yttrium-90 (90Y) and glucocorticoids or with intra-articular glucocorticoids alone. METHODS: A synovial biopsy was carried out blindly 2 weeks before treatment in 66 patients with persistent gonarthritis, who were randomised to treatment either with 90Y and triamcinolone or with placebo and triamcinolone. Immunohistochemistry was used to detect T cells, macrophages, B cells, plasma cells, fibroblast-like synoviocytes, adhesion molecules and pro-inflammatory cytokines. Stained sections were evaluated by digital image analysis. Individual patient improvement was expressed using a composite change index (CCI; range 0-12). Successful treatment was defined as CCI > or = 6 after 6 months. RESULTS: Patients with rheumatoid arthritis, psoriatic arthritis, undifferentiated arthritis and other causes of gonarthritis were included. The overall response rate was 47%. Clinical efficacy in both therapeutic groups was similar and not dependent on diagnosis. No significant differences were noted between baseline microscopic features of synovial tissue inflammation in patients with rheumatoid arthritis and in those with non-rheumatoid arthritis (ie, all diagnoses other than rheumatoid arthritis). The number of macrophages in the synovial sublining was significantly higher in responders than in non-responders (p = 0.002), independent of treatment group and diagnosis. The clinical effect was positively correlated with pretreatment total macrophage numbers (r = 0.28; p = 0.03), sublining macrophage numbers (r = 0.34; p = 0.005) and vascular cell adhesion molecule 1 expression (r = 0.25; p = 0.04). CONCLUSION: The observations support the view that intra-articular treatment either with 90Y and glucocorticoids or with glucocorticoids alone is especially successful in patients with marked synovial inflammation.

Glucocorticoid receptor up-regulation in early rheumatoid arthritis treated with low dose prednisone or placebo.

OBJECTIVE: Low or medium dose prednisone in early rheumatoid arthritis (RA), albeit with significant variation in clinical efficacy, reduces the progression of joint damage. The glucocorticoid receptor (GR) number in peripheral mononuclear cells (PBMC) might be helpful to predict which patients will respond to low or medium dose prednisone and therefore do not or will not need higher doses. With this in mind we determined in a double blind, placebo controlled study at baseline and yearly the GR number in PBMC. METHODS: Eighty-one early RA patients (disease duration less than one year) were included. All patients fulfilled the ACR criteria and were disease modifying antirheumatic drugs (DMARD) and glucocorticoid-naive. They were randomly assigned to treatment with 10 mg prednisone daily or placebo. From all patients disease activity (CRP, number of tender and swollen joints), the radiological joint score, bone mineral density, and the GR number in PBMC were measured annually. RESULTS: In females the GR number was up-regulated over time in both the prednisone and the placebo group. The same trend was observed in males. No correlations were found between the GR number in the prednisone users at the start of their treatment and changes in radiological scores or bone density after 2 years of treatment. No correlations were found between the GR number at the start and the clinical characteristics after a follow-up of 2 years. CONCLUSION: The GR number in the PBMC of early RA patients did not predict which patients would be prednisone responders based on clinical or radiological parameters. However, the up-regulation of the GR number in PBMC in early RA patients towards the GR number of healthy subjects during the first two years of their disease course seems to reflect a recovery or compensatory mechanism as a response to an ongoing inflammatory process. This recovery may be not enough to efficiently control the inflammatory situation.

Down regulation of glucocorticoid receptors in early-diagnosed rheumatoid arthritis.

OBJECTIVE: In patients with rheumatoid arthritis (RA) of longer duration, glucocorticoid receptor (GR) down-regulation has been reported without any change in cortisol levels. This phenomenon might play a role in the aetio-pathogenesis of RA. Therefore we studied GR expression, as well as the serum cortisol levels, in patients with recently diagnosed RA. METHODS: In 81 early diagnosed RA patients with disease duration < 1 year (52F/29M; mean (SD) age 63 (13) years) and in 39 age and sex matched controls (23F/16M; mean age 63 (15) years) blood samples were taken between 8-10 h AM. GR expression (GR-number and GR-affinity), serum cortisol levels, ESR, CRP, painful and swollen joints were measured. RESULTS: A significantly lower GR-number was found in the female patients compared with female controls: 7.0 versus 9.8fmol/million cells, respectively (difference: 2.8, 95% CI 1.1 - 4.6). Interestingly, also serum cortisol levels were significantly lower in the female patients compared with the female controls: 0.21 versus 0.41 micromol/l, respectively (difference: 0.20, 95% CI 0.12 - 0.28). However, between the male patients and male controls no difference was found in GR expression nor in serum cortisol levels. Neither in female nor in male patients were correlations found between GR expression and parameters of disease activity nor was there a relation between GR expression and serum cortisol levels. CONCLUSIONS: Changes in GR expression as well as serum cortisol were not a general phenomenon in early diagnosed RA patients, being present only in females and not related to disease activity. Therefore it seems unlikely that GR expression per se is causally involved in the pathogenesis of RA. We cannot preclude that it may be involved in the incidence, severity and course of RA, as this may be differentially regulated in males and females.

Joint distraction in treatment of osteoarthritis (II): effects on cartilage in a canine model.

OBJECTIVE: From a clinical point of view, joint distraction as a treatment for osteoarthritis (OA) of hip and ankle has been demonstrated to be very promising. Pain, joint mobility and functional ability, the most important factors for a patient with severe OA, all improved. Although radiographic joint space enlargement in a significant number of patients suggested cartilage repair, actual cartilage repair remains difficult to evaluate. Therefore the present study was initiated to evaluate the actual effects of joint distraction on cartilage. METHODS: For this purpose a canine model for OA, anterior cruciate ligament transection (ACLT) was used. Sixteen weeks after ACLT articulating Ilizarov joint distraction of the knee was carried out. Absence of mechanical contact between articular surfaces and presence of intra-articular intermittent fluid pressure, characteristics of Ilizarov joint distraction, were confirmed. Twenty-five weeks after ACLT joint tissue of the dogs was analyzed. RESULTS: Biochemical analysis showed that after joint distraction the abnormal cartilage proteoglycan (PG) metabolism, characteristic for OA, had changed to a level found in control joints. Moreover, a mild degree of inflammation, present after ACLT, was reduced upon joint distraction. PG-content and histological cartilage degeneration had not (yet) improved within the time of treatment. DISCUSSION: Results suggest that the promising clinical results of Ilizarov joint distraction in patients with OA are accompanied by changes in cartilage metabolism. A change in proteoglycan turnover, indicating normalization of overall chondrocyte function, might in the long term, with normal joint use, lead to actual repair of cartilage.

Experimentally induced stress in rheumatoid arthritis of recent onset: effects on peripheral blood lymphocytes.

OBJECTIVE: To examine the effects of experimentally-induced stress on the mobilization of peripheral blood lymphocytes (PBL) in patients with rheumatoid arthritis (RA) of recent onset. METHODS: Twenty-two (16 F, 6 M) patients (mean age 57.6 yrs.) and 23 (15 F, 8 M) healthy subjects (mean age 54.7 yrs.) were subjected to experimental stressors. The numbers of T-cells, B-cells, and NK-cells were determined before and after the completion of tasks inducing physical and mental effort. RESULTS: The change in PBL in response to stress was about equal for patients and healthy subjects (p > 0.75 in all PBL subsets). In patients as well as in healthy subjects, the correlations between PBL and cortisol changes in response to stress tended to be positive, while the correlations between PBL and cardiovascular changes were positive in healthy subjects, but zero or negative in patients. Moderate to high (0.32 < or = r < or = 0.55) correlations between PBL changes and pain were observed. CONCLUSION: Experimentally-induced changes in PBL (as well as cortisol) are normal in patients with early RA who are receiving long term medication, but correlations between these changes and autonomic nervous system responses are zero or negative. This apparent shift in the control of the change in PBL in response to stress is observed in particular in patients with more pain. The pathophysiological significance of these findings should be clarified in longitudinal studies.

Iron deposits and catabolic properties of synovial tissue from patients with haemophilia.

Haemophilic arthropathy is characterised by iron deposits in synovial tissues. We investigated the suggestion that iron plays an important role in synovial changes. We obtained synovial tissue from six patients with haemophilia during arthroplasty, finding that brown haemosideritic tissue was often adjacent to tissue with a macroscopically normal appearance in the same joint. Samples from both types of synovial tissue were analysed histologically and biochemically to determine catabolic activity. Macroscopically haemosideritic synovium showed a significantly higher inflammatory activity than that with a normal appearance. Cultures of abnormal synovial tissue gave a significantly enhanced production of IL-1, IL-6 and TNF alpha compared with cultures of synovial tissue with a normal appearance. In addition, the supernatant fluids from the cultures showed greater catabolic activity from haemosideritic tissue, as determined by the inhibition of the synthesis of articular cartilage matrix. We conclude that in patients with haemophilic arthropathy, local synovial iron deposits are associated with increased catabolic activity.

Degenerated human articular cartilage at autopsy represents preclinical osteoarthritic cartilage: comparison with clinically defined osteoarthritic cartilage.

OBJECTIVE: To investigate whether macroscopically fibrillated human articular knee cartilage observed at autopsy can be considered an early, preclinical phase of osteoarthritis (OA). METHODS: Histological and biochemical characteristics of 3 types of articular knee cartilage were compared: macroscopically degenerated knee cartilage obtained at autopsy (6 donors) from donors without clinical history of OA, normal healthy knee cartilage obtained at autopsy (6 donors), and OA cartilage obtained during joint replacement surgery from patients (n = 6) with clinically defined OA of the knee. From the same donors synovial tissue and synovial fluid were obtained and analyzed for features of inflammation. RESULTS: Histological changes of OA were comparable for degenerated and OA cartilage and significantly different from normal cartilage. Content and synthesis of proteoglycans showed intermediate levels for degenerated tissue compared to normal and OA cartilage. Analysis of synovial tissue revealed a low, mild, and moderate degree of inflammation for joints with normal, degenerated, and OA cartilage, respectively. The same sequence was found for metalloproteinase activity in synovial fluid. CONCLUSION: In general, all changes observed in OA joints were, to a lesser extent, observed in the joints with degenerated cartilage and were significantly different from joints with normal cartilage. We conclude that cartilage degeneration observed at autopsy can be considered a preclinical phase of OA, suitable for studying the process of cartilage degeneration in OA.