Matrix-degrading protease ADAMTS-5 cleaves inter-α-inhibitor and releases active heavy chain 2 in synovial fluids from arthritic patients.

Destruction of the cartilage matrix in joints is an important feature of arthritis. Proteolytic degradation of cartilage glycoproteins can contribute to the loss of matrix integrity. Human inter-α-inhibitor (IαI), which stabilizes the extracellular matrix, is composed of the light-chain serine proteinase inhibitor bikunin and two homologous heavy chains (HC1 and HC2) covalently linked through chondroitin 4-sulfate. Inflammation promotes the transfer of HCs from chondroitin 4-sulfate to hyaluronan by tumor necrosis factor-stimulated gene-6 protein (TSG-6). This reaction generates a covalent complex between the heavy chains and hyaluronan that can promote leukocyte invasion. This study demonstrates that both IαI and the HC-hyaluronan complex are substrates for the extracellular matrix proteases ADAMTS-5 and matrix metalloprotease (MMP) -3, -7, and -13. The major cleavage sites for all four proteases are found in the C terminus of HC2. ADAMTS-5 and MMP-7 displayed the highest activity toward HC2. ADAMTS-5 degradation products were identified in mass spectrometric analysis of 29 of 33 arthropathic patients, indicating that ADAMTS-5 cleavage occurs in synovial fluid in arthritis. After cleavage, free HC2, together with TSG-6, is able to catalyze the transfer of heavy chains to hyaluronan. The release of extracellular matrix bound HC2 is likely to increase the mobility of the HC2/TSG-6 catalytic unit and consequently increase the rate of the HC transfer reaction. Ultimately, ADAMTS-5 cleavage of HC2 could alter the physiological and mechanical properties of the extracellular matrix and contribute to the progression of arthritis.

Inhibitory effect of Triperygium wilfordii polyglucoside on dipeptidyl peptidase I in vivo and in vitro.

BACKGROUD: Dipeptidyl peptidase I (DPPI), a lysosomal cysteine protease is derived from granule immune cells including mast cell, neutrophils, and toxicity T cells. DPPI can activate serine proteases by removal of dipeptides from N-termini of the pro-proteases, resulting in granule immune cells activation which involved in physiological or pathological responses. Triperygium Wilfordii Polyglucoside (TWP) is one of the traditional Chinese medicines, and commonly used in rheumatoid arthritis (RA) treatment. The present study intended to evaluate the effects of TWP on DPPI activity. METHODS: In vivo and in vitro studies were carried out to investigate the functions of TWP or triptolide (TP) on DPPI activities in serum, tissues of CIA rats. Rats were divided into five groups randomly: normal group, untreated CIA rat group, TWP treatment CIA groups (the low dose 2.5mg/100g body-weight and high dose 5mg/100g body-weight), and TP treatment CIA group (4µg/100g body-weight). Arthritis development was monitored visually, and joint pathology was examined radiologically. Total protein concentrations in synovial fluids (SFs) were determined by BCA method. Serums and tissue homogenates from CIA rats were collected and DPPI activities were detected using fluorescence substrate GF-AFC. The in vitro interactions between DPPI in serums or in tissue homogenates and TWP or TP were assessed. RESULTS: TWP-treated CIA rats showed a significant improvement in bone erosion. TWP significantly suppressed paw swelling and total protein concentration in the SFs of CIA rats compared with untreated CIA rats. The elevated activities of DPPI in serums or tissues of CIA rats were significantly inhibited by TWP, but not by TP in vivo. The inhibitory effects of TWP on DPPI activities were also confirm by in vitro study. CONCLUSION: One of the therapeutic functions of TWP in RA treatment could be inhibiting DPPI activity in serums and synovial tissue produced during RA development, and then reducing inflammatory serine proteases activities and further recovering CIA rats from RA symptoms.

The regulatory role of Dipeptidyl peptidase I on the activation of immune granulocytes.

Dipeptidyl peptidase I (DPPI), a lysosomal cysteine protease, required for activation of serine proteases of granulocytes including mast cells (MCs), neutrophils (NPs) and others, which were found in synovial tissue of patients with rheumatoid arthritis (RA). But, the role of DPPI associated with those cells in RA development is unclear. In this study, the collagen-induced-arthritis (CIA) rat-model was employed to investigate the expression and activity levels of DPPI and its association with RA progress. Primary granulocytes were freshly extracted from bone-marrows of normal or CIA rats, human mast cell line LAD-2 and primary neutrophils, human-recombinant-DPPI, DPPI-inhibitor Gly-Phe-CHN2 , LTB4, anti-IgE antibody, calcium ionophore were used to study the regulatory role of DPPI in cell activations. The increased DPPI activities in synovial fluids, serum, and bone-marrow homogenates of CIA rats associated with RA severities progress were observed after injections. MMP2/9 expressions in SFs and bone-marrow were in different patterns. Regular-Blood-Tests have shown the high leveled DPPI activities associated with granulocytes differentiations in-vivo in blood of CIA rats. In-vitro cell models, DPPI up-regulated the proliferation of primary bone-marrow granulocytes of normal rats, but inhibited that of CIA rats. DPPI up-regulated and Gly-Phe-CHN2 down-regulated MCs intracellular DPPI and chymase activities. Gly-Phe-CHN2 also inhibited the LTB4 -activated-NPs and NP-elastase activities. Following stimulation of calcium ionophore, the net-releases of DPPI and ß-hexosaminidase from MCs were increased over a time-course, while Gly-Phe-CHN2 down-regulated MCs and NPs activation. Our findings demonstrate the role of DPPI in regulating MCs and NPs activation, and modulating proteolysis in the process of RA.

[Age-related traits of cytogenetic abnormalities in synovial cells of knee-joint in residents of the North of Siberia with arthritis of different etiology depending on the GSTM1 gene polymorphism of glutathion-S-transferase.]

It was studied the frequency of cells with cytogenetic abnormalities in the synovial fluid cells of the knee joint in patients of different age groups suffering from chronic arthritis associated with Lyme borreliosis (CAALB) or post-traumatic arthritis (PTA), depending on the polymorphism of the GSTM1 gene of glutathione-S-transferase. The study included 135 residents of the north of the Tomsk and Tyumen regions, 68 of whom suffered from CAALB, and the rest of the 67 patients who made up the control group were diagnosed with PTA. The results of this study have demonstrated that there are significant age-related differences in the frequency of cytogenetic abnormalities of the synovial fluid cells of the knee joint between young and elderly patients of СAALB. The integrative assessment of clinical and cytogenetic parameters in the group of elderly СAALB patients with mutant GSTM1 (0/0) allele, as compared with the other groups, enable to conclude that there are significant positive correlations between the indices of the severity disruption of articular locomotor function and the frequency of synovial fluid cells with trisomy of chromosome 7.

Characterization of a new mPGES-1 inhibitor in rat models of inflammation.

Microsomal prostaglandin E synthase (mPGES)-1 inhibition has been proposed as an alternative to cyclooxygenase (COX) inhibition in the treatment of pain and inflammation. This novel approach could potentially mitigate the gastro-intestinal and cardiovascular side effects seen after long-term treatment with traditional non-steroidal anti-inflammatory drugs (NSAIDs) and Coxibs respectively. Several human mPGES-1 inhibitors have been developed in the recent years. However, they were all shown to be considerably less active on rodent mPGES-1, precluding the study of mPGES-1 inhibition in rodent models of inflammation and pain. The aim of this study was to characterize the new mPGES-1 inhibitor compound II, a pyrazolone that has similar potency on rat and human recombinant mPGES-1, in experimental models of inflammation. In cell culture, compound II inhibited PGE2 production in synovial fibroblasts from patients with rheumatoid arthritis (RASF) and in rat peritoneal macrophages. In vivo, compound II was first characterized in the rat air pouch model of inflammation where treatment inhibited intra-pouch PGE2 production. Compound II was also investigated in a rat adjuvant-induced arthritis model where it attenuated both the acute and delayed inflammatory responses. In conclusion, compound II represents a valuable pharmacological tool for the study of mPGES-1 inhibition in rat models.

Ability of curcuminoid compared to diclofenac sodium in reducing the secretion of cycloxygenase-2 enzyme by synovial fluid's monocytes of patients with osteoarthritis.

AIM: to assess the ability of curcuminoid from Curcuma domestica Val in reducing the cycloxygenase-2 secretion by synovial fluid's monocytes compared to diclofenac sodium in patients with osteoarthritis. METHODS: this was a prospective randomized open end blinded evaluation (PROBE) study. The subjects were patients with knee osteoarthritis who were divided randomly into two groups, the first group received 30 mg 3 times daily of curcuminoid and the second group received 25 mg 3 times daily of diclofenac sodium. The joints aspiration was done and the secretion of cycloxygenase-2 enzyme by synovial fluid's monocytes was evaluated by scoring method before and after 4 weeks of treatments. RESULTS: a total of 80 patients with knee osteoarthritis were enrolled. In curcuminoid group the average scores were 1.84±0.37 and 1.15±0.28 respectively (p<0.001). In diclofenac group the average scores were 1.79±0.38 and 1.12±0.27 respectively (p<0.001). In curcuminoid group the decreasing score of cycloxygenase-2 secretion was 0.70±0.51 while in diclofenac group was 0.67±0.45. There was no significant difference in decreasing the score of cycloxygenase enzyme secretion between both treatment groups (p=0.89). CONCLUSION: the ability of curcuminoid from Curcuma domestica Val. rhizome extract was not significantly different compared to diclofenac sodium in suppressing the secretion of cycloxygenase-2 enzyme by synovial fluid's monocytes.

Over-expression of extracellular superoxide dismutase in mouse synovial tissue attenuates the inflammatory arthritis.

Oxidative stress such as reactive oxygen species (ROS) within the inflamed joint have been indicated as being involved as inflammatory mediators in the induction of arthritis. Correlations between extracellular- superoxide dismutase (EC-SOD) and inflammatory arthritis have been shown in several animal models of RA. However, there is a question whether the over-expression of EC-SOD on arthritic joint also could suppress the progression of disease or not. In the present study, the effect on the synovial tissue of experimental arthritis was investigated using EC-SOD over-expressing transgenic mice. The over-expression of EC- SOD in joint tissue was confirmed by RT-PCR and immunohistochemistry. The degree of the inflammation in EC-SOD transgenic mice was suppressed in the collagen-induced arthritis model. In a cytokine assay, the production of pro-inflammatory cytokines such as, IL-1ß, TNFα, and matrix metalloproteinases (MMPs) was decreased in fibroblast-like synoviocyte (FLS) but not in peripheral blood. Histological examination also showed repressed cartilage destruction and bone in EC-SOD transgenic mice. In conclusion, these data suggest that the over-expression of EC-SOD in FLS contributes to the activation of FLS and protection from joint destruction by depressing the production of the pro-inflammatory cytokines and MMPs. These results provide EC-SOD transgenic mice with a useful animal model for inflammatory arthritis research.

The role of human xanthine oxidoreductase (HXOR), anti-HXOR antibodies, and microorganisms in synovial fluid of patients with joint inflammation.

This work is to investigate the levels of human xanthine oxidoreductase (HXOR), its antibodies, and microorganisms in synovial fluid of patients with untreated rheumatoid joint diseases. Synovial fluids were collected from sixty-four patients with rheumatoid joint diseases. Sixty-four age-matched individuals were included as control. Xanthine oxidoreductase (XOR) proteins level and anti-XOR antibodies were determined in the blood and synovial fluid, using human XOR as antigen, by enzyme-linked immunosorbent (ELISA) assay. Synovial fluids were cultured for bacteria and fungi. The titers of XOR protein in the synovial fluid of patients with rheumatoid arthritis were 90.43 ± 23.37 µg/ml (mean ± SD, n = 29) and up to 62.42 ± 8.74 µg/ml (mean ± SD, n = 35) in other joint inflammation. Anti-HXOR antibodies titers in patients were 167.72 ± 23.64 µg/ml, n = 64, which was significantly higher in rheumatoid arthritis patients. The results indicated that anti-HXOR antibodies in synovial fluids have a protective role as high concentrations against XOR were detected in inflammatory arthritis. These antibodies play a role in eliminating XOR from synovial fluids. However, immune complex formation could activate complement and participate in propagating the inflammatory cycle. Synovial aspirate ordinary microbial cultures were negative for any bacteria or fungi, but that does not exclude organisms of special culture requirements.

Matrix metalloproteinase activity in stifle synovial fluid of cranial cruciate ligament deficient dogs and effect of postoperative doxycycline treatment.

This prospective clinical study investigated the activity of matrix metalloproteinases (MMPs) in stifle synovial fluid (SF) of 13 dogs with acute cranial cruciate ligament (CCL) rupture, and the effect of a postoperative doxycycline treatment. MMP-2, 3, 9 and 13 activities were compared with respect to the time of sampling (preoperatively or 1 month after surgical stabilisation) and the type of postoperative adjuvant treatment (doxycycline or not). No significant activity was detected for both MMP-3 and MMP-13. MMP-2 and MMP-9 activities were found to be significantly highly increased in SF of CCL ruptured stifles compared to control stifles of unaffected dogs. No significant effect from surgical stabilisation and postoperative doxycycline treatment on MMP-2 and MMP-9 activities was found, indicating that doxycycline may not be an appropriate postoperative medical treatment after CCL rupture.

Over-expression of extracellular superoxide dismutase in mouse synovial tissue attenuates the inflammatory arthritis

Oxidative stress such as reactive oxygen species (ROS) within the inflamed joint have been indicated as being involved as inflammatory mediators in the induction of arthritis. Correlations between extracellular-superoxide dismutase (EC-SOD) and inflammatory arthritis have been shown in several animal models of RA. However, there is a question whether the over-expression of EC-SOD on arthritic joint also could suppress the progression of disease or not. In the present study, the effect on the synovial tissue of experimental arthritis was investigated using EC-SOD over-expressing transgenic mice. The over-expression of EC-SOD in joint tissue was confirmed by RT-PCR and immunohistochemistry. The degree of the inflammation in EC-SOD transgenic mice was suppressed in the collagen-induced arthritis model. In a cytokine assay, the production of pro-inflammatory cytokines such as, IL-1beta, TNFalpha, and matrix metalloproteinases (MMPs) was decreased in fibroblast-like synoviocyte (FLS) but not in peripheral blood. Histological examination also showed repressed cartilage destruction and bone in EC-SOD transgenic mice. In conclusion, these data suggest that the over-expression of EC-SOD in FLS contributes to the activation of FLS and protection from joint destruction by depressing the production of the pro-inflammatory cytokines and MMPs. These results provide EC-SOD transgenic mice with a useful animal model for inflammatory arthritis research.

Formation of invadopodia-like structures by synovial cells promotes cartilage breakdown in collagen-induced arthritis: involvement of the protein tyrosine kinase Src.

OBJECTIVE: Invasive synovial fibroblasts are suggested to be the major effectors of cartilage and bone destruction, and this aggressive phenotype can lead to irreversible damage. In cancer cells, invasion across tissue boundaries and metastasis have recently been shown to depend on the capacity of the cells to breach the basement membrane, a process that was linked to the formation of the actin-rich cell protrusions called invadopodia. This study was undertaken to investigate whether arthritic synovial cells use invadopodia to invade and degrade cartilage components. METHODS: Fibroblast-like synoviocytes (FLS) from control rats or rats with collagen-induced arthritis (CIA) were cultured on fluorescent matrix in the presence of Src inhibitors or were transfected with wild-type or variants of Src kinases. The in vivo effect of Src inhibition on cartilage degradation and invasion was studied in a rat model of CIA. RESULTS: FLS from rats with CIA produced more invadopodia-like structures than did FLS from control rats, leading to increased extracellular matrix degradation. Furthermore, c-Src activation was increased in synovial cells from rats with CIA, and Src activity was found to mediate the formation of invadopodia. Pharmacologic blockade of Src activity by PP2 or intraarticular expression of a c-Src-specific short hairpin RNA in the CIA model reduced synovial membrane hyperplasia and cartilage degradation, an event linked to decreased invadopodia formation by synovial fibroblasts. CONCLUSION: This study demonstrates that inhibition of invadopodia formation in arthritic synovial cells leads to a direct effect on extracellular matrix degradation in vitro and in vivo, making invadopodia a relevant therapeutic target for interfering with this process.

Clinical study on the treatment of knee osteoarthritis of Shen-Sui insufficiency syndrome type by electroacupuncture.

OBJECTIVE: To study the clinical effificacy of electroacupuncture (EA) on treating knee osteoarthritis (KOA) of Shen ()-Sui () insuffificiency (SSI) syndrome type. METHODS: A total of 245 patients (279 knees) of KOA-SSI were randomly assigned to two groups by lottery: 141 knees in the treatment group and 138 knees in the control group. The treatment group was managed with EA at the dominant points of Neixiyan (Ex-LE4) and Waixiyan (Ex-LE5) as well as the conjugate points of Xuanzhong (GB39) and Taixi (KI3) for 30 min, once a day, with 15 days as one course; 2 courses were applied with a 5-day interval in between. The control group was treated with intra-articular injection of 2 mL hyaluronic acid into the affected joint every 7 days for 5 times in total. The clinical effects on the patients in different stages were observed, and their symptom scores of knee and contents of cytokines, including interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), prostaglandin E(2alpha) (PGE(2alpha)) and matrix metalloproteinases-3 (MMP-3), in the knee joint fluid were measured before and after treatment. RESULTS: The study was completed in 235 patients (263 knees); four patients (7 knees) in the treatment group and six patients (9 knees) in the control group dropped out. Comparison of therapeutic effects (excellent and effective rates) between the two groups showed insignificant differences (P>0.05). Symptom scores of knee and contents of cytokines in the knee flfluid after treatment were lowered signifificantly in the patients of stage I-III in both groups (P<0.05 or P<0.01). However, the lowering of the total symptom score of knee in the patients of stage III in the treatment group was more signifificant (P<0.05). CONCLUSIONS: EA could effectively alleviate the clinical symptoms in KOA patients of stage III, showing an effect superior to that of hyaluronic acid. EA also shows action in suppressing the secretion of IL-1, IL-6, TNF-alpha, PGE(2alpha) and MMP-3 in the knee flfluid.

Group IIA secretory PLA2 inhibition by ursolic acid: a potent anti-inflammatory molecule.

Ursolic acid (3beta-hydroxy-urs-12-en-28-oic acid) isolated from many medicinal plants has diverse pharmacologically important properties, including strong anti-inflammatory activity. However its interaction with pro-inflammatory PLA2 is not known. Ursolic acid inhibited secretory PLA2 (sPLA2) enzymes purified from Vipera russelli, Naja naja venom and human pleural fluid and synovial fluid. IC50 values determined for these enzymes ranged from 12 to 18 microM. Group II secretory PLA2 from both venoms & human inflammatory source were found to be sensitive to inhibition in comparison with group I cobra venom sPLA2. Variation in Ca2+ concentration from 2.5-15 mM did not alter the level of inhibition. Similarly sPLA2 inhibition by ursolic acid is independent of substrate concentration. Ursolic acid interacts with purified venom sPLA2 enzymes and enhances relative fluorescence intensity in a dose dependent manner. In the presence of ursolic acid apparent shift in the far UV-CD spectra of sPLA2 was observed, indicating a direct interaction with the enzyme and formation of enzyme-ursolic acid complex. This complex results in irreversible inhibition of sPLA2 as evident by dialysis study. Inhibition of sPLA2 induced mouse paw edema and indirect hemolytic activity confirmed its sPLA2 inhibitory activity in vivo and in situ respectively. These studies revealed that the strong anti-inflammatory activity of ursolic acid is by inhibiting sPLA2 enzymes.

Oxidant and anti-oxidant systems of synovial fluid from patients with knee post-traumatic arthritis.

It has been suggested that patients with knee post-traumatic arthritis (PA), associated or not to haemarthrosis (HA), display altered oxidant and anti-oxidant systems in their synovial fluid. This study aimed to establish whether this is really the case. Synovial fluid samples were obtained by transdermal arthrocentesis from 69 patients with PA (36 of them had HA) and 22 control subjects. The activities of synovial fluid zinc-copper superoxide dismutase (ZnCuSOD) and manganese superoxide dismutase (MnSOD) isoenzymes, catalase (CAT), glutathione peroxidase (GPX), glutathione reductase (GR) and glutathione-S-transferase (GST) enzymes, and malondialdehyde (MDA) concentration and synovial fluid viscosity were measured in the study groups. Patients with PA had significantly increased activities of all antioxidant enzymes, except CAT, and MDA concentration than did the controls. However, synovial fluid viscosity was found to be decreased in the study group, mainly in the HA subgroup. Results suggest that excessive free radicals production may exist in synovial fluid of PA patients and may contribute to knee joint destruction.

Effects of bee venom on protease activities and free radical damages in synovial fluid from type II collagen-induced rheumatoid arthritis rats.

The effect of bee venom acupuncture (BVA) (api-toxin) on the development of type II collagen (CII)-induced arthritis (CIA) in rats has been studied. We have compared the levels of activity of a comprehensive range of cytoplasmic, lysosomal and matrix protease types, together with the levels of free radical-induced protein damage (determined as protein carbonyl derivative) in synovial fluid from CIA-treated, BVA-treated and normal rats. Many protease types showed significantly increased activity in CIA compared with normal rats. BVA (5 and 10 microl/100g) significantly reduced these enzyme activities by some 80% each, but levels of plasma proteases activity (including those enzyme types putatively involved in the immune response, such as dipeptidyl aminopeptidase IV and proline endopeptidase) in CIA, BVA (5 microl/100g)-treated and normal plasma samples were not significantly different. The level of free radical induced damage to synovial fluid proteins was approximately three-fold higher in CIA compared with normal rats. However, BVA (5 microl/100g) significantly decreased the level of reactive oxygen free radical species (ROS) induced oxidative damage to synovial fluid proteins. It was concluded that activation of proteolytic enzymes and free radicals are likely to be of equal potential importance as protein damaging agents in the pathogenesis of rheumatoid arthritis (RA), and the development of novel therapeutic strategies for the latter disorder should include both protease inhibitory and free radical scavenging elements. In addition, the protease inhibitory element should be designed to inhibit the action of a broad range of enzymatic mechanistic types (cysteine, serine, metallo proteinases and peptidases). In conclusion, BVA is considered to be an effective RA modulator, inhibiting protease activities and removing ROS.

A novel approach to measure the contribution of matrix metalloproteinase in the overall net proteolytic activity present in synovial fluids of patients with arthritis.

Despite decades of research, only a very limited number of matrix metalloproteinase (MMP) inhibitors have been successful in clinical trials of arthritis. One of the central problems associated with this failure may be our inability to monitor the local activity of proteases in the joints since the integrity of the extracellular matrix results from an equilibrium between noncovalent, 1:1 stoichiometric binding of protease inhibitors to the catalytic site of the activated forms of the enzymes. In the present work, we have measured by flow cytometry the net proteolytic activity in synovial fluids (SF) collected from 95 patients with osteoarthritis and various forms of inflammatory arthritis, including rheumatoid arthritis, spondyloarthropathies, and chronic juvenile arthritis. We found that SF of patients with inflammatory arthritis had significantly higher levels of proteolytic activity than those of osteoarthritis patients. Moreover, the overall activity in inflammatory arthritis patients correlated positively with the number of infiltrated leukocytes and the serum level of C-reactive protein. No such correlations were found in osteoarthritis patients. Members of the MMP family contributed significantly to the proteolytic activity found in SF. Small-molecular-weight MMP inhibitors were indeed effective for inhibiting proteolytic activity in SF, but their effectiveness varied greatly among patients. Interestingly, the contribution of MMPs decreased in patients with very high proteolytic activity, and this was due both to a molar excess of tissue inhibitor of MMP-1 and to an increased contribution of other proteolytic enzymes. These results emphasize the diversity of the MMPs involved in arthritis and, from a clinical perspective, suggest an interesting alternative for testing the potential of new protease inhibitors for the treatment of arthritis.

MMP protein and activity levels in synovial fluid from patients with joint injury, inflammatory arthritis, and osteoarthritis.

OBJECTIVE: To determine protein and activity levels of matrix metalloproteinases 1 and 3 (MMP-1 and MMP-3) in synovial fluid of patients with knee joint injury, primary osteoarthritis, and acute pyrophosphate arthritis (pseudogout). METHODS: Measurements were done on knee synovial fluid obtained in a cross sectional study of cases of injury (n = 283), osteoarthritis (n = 105), and pseudogout (n = 65), and in healthy controls (n = 35). Activity of MMP-1 and MMP-3 in alpha(2) macroglobulin complexes was measured using specific low molecular weight fluorogenic substrates. ProMMP-1, proMMP-3, and TIMP-1 (tissue inhibitor of metalloproteinase 1) were quantified by immunoassay. RESULTS: Mean levels of proMMP-1, proMMP-3, and TIMP-1 were increased in injury, osteoarthritis, and pseudogout compared with controls. MMP-1 activity was increased in pseudogout and injury groups over control levels, whereas MMP-3 activity was increased only in the pseudogout group. The increase in MMP-1 activity coincided with a decrease in TIMP-1 levels in the injury group. CONCLUSIONS: Patients with joint injury have a persistent increase in proMMP-1 and proMMP-3 in synovial fluid and an increase in activated MMPs, which are not inhibited by TIMP. The differences in activation and inhibition patterns between the study groups are consistent with disease specific patterns of MMP activation and/or inhibition in joint pathology.

Matrix metalloproteinase 9, apoptosis, and vascular morphology in early arthritis.

OBJECTIVE: To examine matrix metalloproteinase 9 (MMP-9) in the synovial fluid (SF) and synovial membrane (SM) in relation to vascular endothelial cell (EC) apoptosis, vascular endothelial growth factor (VEGF), and SM vascular pattern. METHODS: Thirty-four patients underwent needle arthroscopy of the knee joint; 12 had early rheumatoid arthritis (RA), 12 had early psoriatic arthritis (PsA), and 10 had osteoarthritis (OA). The early RA and early PsA patients were matched for disease activity. SF levels of MMP-9 and VEGF were measured by an enzyme-linked immunosorbent assay, and EC apoptosis was measured by TUNEL assay. MMP-9 expression was examined in SM by immunohistochemistry. Synovial tissue explants were stimulated with VEGF, and MMP-9 levels were measured in the supernatants. The synovial vascular pattern was recorded. RESULTS: SF MMP-9 levels were significantly higher in early PsA patients than in early RA patients; OA patients had minimal levels. MMP-9 levels correlated with blood vessel morphology and SF VEGF levels. MMP-9 expression was greater in early PsA SM than in early RA SM, but the difference was not significant. In contrast however, EC apoptosis was greater in early RA SM than in early PsA SM. MMP-9 levels increased 2-fold and 9-fold, respectively, in SM explant culture supernatants on day 7 in response to stimulation with 25 ng/ml and 50 ng/ml of VEGF. CONCLUSION: SF MMP-9 levels correlate with the pattern of SM neovascularization and SF VEGF levels in early inflammatory arthritis, and VEGF increases MMP-9 production by SM. Endothelial cell apoptosis, however, appears to be more prevalent in early RA. This combination of factors may explain the pattern of differential angiogenesis in these arthritides.

[Practical significance of cytokine determination in joint fluid in patients with arthroses or rheumatoid arthritis]. / Praktische Bedeutung der Zytokinbestimmung im Gelenkpunktat von Patienten mit Arthrosen oder rheumatischen Arthritiden.

OBJECTIVE: To determine whether the activity of cartilage-degrading enzymes in the synovial fluid (SF) of patients with rheumatoid arthritis and other joint diseases is correlated with the concentration of cytokines in the SF. METHODS: Cytokines and cartilage-degrading enzymes were determined in the SF of 97 patients with various disorders involving the knee joints (rheumatoid arthritis (RA) n 44; osteoarthritis (OA) n 35; meniscal trauma (Men) n 10; reactive arthritides (ReA) n 8). In these samples we measured the concentrations of interleukin-1 alpha and beta, IL-1-receptor antagonist (IL-1ra), IL-6, IL-8, tumor necrosis factor alpha (TNF alpha; all by ELISA), collagenase-activity and caseinase-activity (by substrate assays). RESULTS: With the exception of IL-1 alpha and IL-6, cytokine-concentrations were significantly higher in RA than in OA SF-samples (p < 0.05; ANOVA on ranks). IL-1ra, IL-6, and IL-1 beta were correlated best with the collagenase-activity in the SF (r = 0.63; 0.57; 0.55; Spearman's rank correlation), while IL-1 beta (r = 0.53) and IL-1ra (r = 0.52) were best correlated with the caseinase-activity in the samples. The SF-concentration of IL-1ra was well correlated with the levels of IL-6, IL-1 beta, II-8, and TNF alpha (r from 0.73 to 0.66; all p < 0.005), but not with IL1 alpha. The molar ratio of IL-1 to IL-1ra in the SF was neither correlated with the activity of collagenase nor caseinase. IL-1 beta and IL-1ra in the SF were positively correlated with the erythrocyte sedimentation rate (ESR). CONCLUSIONS: The determination of IL-1 beta and IL-1ra in the SF of patients with joint disorders as examined in this study seems to allow to a certain extent a prediction of the collagenase- and caseinase-activity contained in the diseased joint. We would favor.

[Change in phospholipid composition of synovial fluid in children with various types of arthritis]. / Izmenenie fosfolipidnogo sostava sinovial'noi zhidkosti u detei s razlichnymi artritami.

Phospholipid composition of synovial fluid was studied in patients with reactive, rheumatoid and juvenile chronic forms of arthritis as compared with that of normal synovial fluid. The most pronounced alterations of the synovial fluid phospholipid composition (additional phospholipid fractions, increase in content of lyso-derivatives) were found in the patients with rheumatoid and juvenile chronic arthritis, which appear to occur due to activation of endogenous phospholipases A2, C and lysophospholipase A1.