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Objective: The objective of this study was to investigate CD44 and proteoglycan metabolism in patients suffering from Kashin-Beck Disease (KBD), an endemic osteoarthropathy that affects 2.5 million of 30 million people living in the KBD regions of China.Methods: Immunohistochemical analyses of cluster of differentiation-44 (CD44), BC-13 and 3-B-3(-) expression were performed in cartilage sections harvested from KBD and normal patients. In addition, the serum levels of soluble CD44 (sCD44), interleukin-1beta (IL-1beta), tumor necrosis factor-alpha (TNF-alpha) and matrix metalloproteinase-1 were determined using a sandwich enzyme linked immunosorbent assay.Results: Hematoxylin & eosin and toluidine blue staining indicated that there was cell necrosis and proteoglycan loss in cartilage from both KBD children and adult cartilage. Strong immunohistochemical staining for CD44, BC-13 and 3-B-3(-) occurred in the majority of adult KBD patients and most KBD children. Furthermore, statistically significant elevated levels of sCD44, IL-1beta and TNF-alpha were found in the sera of both adult and child KBD patients when compared to the levels of normal adult and child controls. Interestingly, IL-1beta and TNF-alpha serum levels were all high in normal children from KBD regions when compared to normal children from non-KBD regions suggesting that unidentified factors (e.g., a genetic predisposition) may protect some people from KBD pathology.Conclusion: Our results demonstrate that altered CD44, IL-1beta and TNF-alpha metabolism occurs in the pathogenesis of KBD and there is an increased aggrecanase-generated proteoglycan loss from KBD adult and child cartilage. These primary metabolic changes are likely to be significant contributing factor causing pathological joint formation and instability that leads to secondary osteoarthritis in KBD patients. [ABSTRACT FROM AUTHOR]

Objective: Oxidative stress occurs when the metabolic balance of a cell is disrupted through exposure to excess pro-oxidant. Whilst it is known that unregulated production or exposure to exogenous sources of pro-oxidants induces chondrocyte cell death and degrades matrix components in vitro, relatively little is known of the effects of pro-oxidants on articular cartilage in situ. The objective of this study was to determine if a single exposure to the pro-oxidant hydrogen peroxide (H(2)O(2)) induces a degenerative phenotype.Methods: Articular cartilage explants were obtained from skeletally mature bovine steers and exposed to a single dose of hydrogen peroxide (0.1-1.0 mM) and cultured for up to 21 days. Cell death, and sulfated glycosaminoglycan loss into the medium and gene expression were quantitatively determined. Adoption of an abnormal chondrocyte phenotype was analyzed through the expression of 3B3(-), nitrotyrosine and procollagen type IIA epitopes in cartilage explants.Results: Cell death occurred primarily at the surface zone of cartilage in a dose-dependent manner in H(2)O(2) treated explants, and supplementation of standard serum-free medium with insulin-selenium-transferrin significantly reduced cell death (>fourfold). Nitric oxide synthase-2 gene expression and proteoglycan loss increased in oxidant treated explants in a concentration-dependent manner. Antibody labeling to 3B3(-), procollagen type IIA and nitrotyrosine was present in all treated explants but absent in untreated explants.Conclusions: This study demonstrates that a single exposure to high levels of pro-oxidant causes the expression of genes and antibody epitopes that are associated with early degenerative changes observed in experimental osteoarthritis. [ABSTRACT FROM AUTHOR]

Summary Objective To evaluate the potential of ADAMTS-4 (aggrecanase -1) activity in synovial fluid (SF) as a biomarker of knee injury and joint disease. Design We have measured ADAMTS-4 activity in the synovial fluid of 170 orthopaedic patients with different degrees of joint pathology, using a commercial ADAMTS-4 fluorescence resonance energy transfer (FRET) substrate assay. Patients were classified at arthroscopy as (i) macroscopically normal, (ii) with an injury of the meniscus, anterior cruciate ligament or chondral/osteochondral defects or (iii) with osteoarthritis, and the influence of independent factors (age, patient group, effusion and synovial inflammation) on ADAMTS-4 activity levels was assessed. Results In most patients (106/170) ADAMTS-4 activity was undetectable; ADAMTS-4 ranged from 0 to 2.8 ng/mL in synovial fluid from patients with an injury, 0–4.1 ng/mL in osteoarthritic patients and 4.0–12.3 ng/mL in patients with large effusions. Four independent variables each significantly influenced ADAMTS-4 activity in synovial fluid (all P < 0.001): age (concordance = 0.69), presence of osteoarthritis (OA) (concordance = 0.66), level of effusion (concordance = 0.78) and inflammation (concordance = 0.68). Not only did effusion influence the amount of ADAMTS-4 activity most strongly, but it also did this in an ordered manner ( P < 0.001). Conclusions The main finding of this study is that ADAMTS-4 levels in synovial fluid are most strongly correlated with inflammation and severity of effusion in the knee. Further study is required to determine if it could provide a useful tool to aid clinical diagnoses, indicate treatment, to monitor progression of joint degeneration or OA or alternatively the success of treatment. [ABSTRACT FROM AUTHOR]

Summary Objective Kashin-Beck Disease (KBD) is a rare and severe osteoarthropathy endemic to China. We evaluated the frequency and patterns of hand radiographic osteoarthritis (rOA) in adults with and without KBD. Methods Han Chinese ( N = 438) from Yongshou County of central China underwent right hand radiography for determining case status. Presence of KBD was based on characteristic radiographic deformities of articular ends of bones including articular surface depression, carpal crowding, any subchondral bone deformities in the proximal end of phalanges or first metacarpal bone, or the distal ends of metacarpal bones 2–5, and any bony enlargement with deformity of the distal ends of phalanges. Hand rOA severity was determined by osteophyte (OST), joint space narrowing (JSN), and Kellgren and Lawrence (KL) grades. Results This study included 127 KBD and 311 non-KBD adults of similar mean age (39 years) and body mass index (BMI) (21 kg/m 2 ). Inter- and intra-rater reliability for radiographic determination of case status and rOA features was high (kappa 0.72–0.96). Compared to non-KBD, KBD adults had significantly more severe hand rOA of the thumb, distal interphalangeal (DIP), proximal interphalangeal (PIP) and metacarpophalangeal (MCP) joints. Only KBD adults had end-stage carpometacapal (CMC) disease. In KBD, DIPs and PIPs were more affected than MCPs and the frequency of OSTs was significantly higher in PIPs than DIPs. Conclusions Compared with age-matched adults from the same area and farming occupation, KBD hand rOA was more widespread and severe, particularly of PIPs and CMCs. The ability to differentiate adult KBD from non-KBD hand rOA will facilitate genetic analyses of the vast majority of affected individuals. [ABSTRACT FROM AUTHOR]

Objectives: To identify changes in the expression patterns of enzymes involved in chondroitin sulfate (CS) glycosaminoglycan (GAG) metabolism in articular cartilage proteoglycan (PG) isolated from adolescent patients with Kashin-Beck disease (KBD). Methods: Samples of articular cartilage were divided into two groups: Control samples (from five normal children), and KBD samples (from five KBD children) aged 3-12 years old. The morphology and pathology of hand joint cartilage were examined by histochemical staining. The localization and expression patterns of enzymes involved in CS GAG metabolism (i.e., PAPS synthetase 2 (PAPSS2), PAPS transporter 1 (PAPST1), Carbohydrate (N-acetylgalactosamine 4-sulfate 6-O) sulfotransferases 15 (CHST15), Arylsulfatase B (ARSB) and N-acetylgalactosamine-6-sulfate sulfatase (GALNS)) were performed using immuno-histochemical analyses. Positive immunostaining in articular cartilage was semi-quantified. Results: Reduced aggrecan staining was observed in KBD samples compared with the control samples. The percentages of positive staining for the anabolic enzymes PAPSS2, PAPST1 and CHST15 in the upper and middle zones of KBD samples were significantly lower than that found in the Controls. In contrast, the percentages of positive staining in KBD samples for the catabolic enzymes ARSB and GALNS were significantly higher than the control samples. However, the staining for all of these GAG metabolism enzymes were hardly observed in the deep zones of KBD cartilage, suggesting that significant cell death and necrosis had occurred in this region. Conclusions: Our results indicate that alterations of enzymes involved in articular cartilage CS GAG metabolism on PGs in the articular cartilage play an important role in the onset and pathogenesis of KBD in adolescent children. [ABSTRACT FROM AUTHOR]

Abstract: Musculoskeletal complaints are the second most frequent reason for medical treatments. Within these diseases rheumatoid arthritis (RA) and, especially, osteoarthritis (OA) are common. Although the causes of arthritis are multifactorial and not fully understood, clinical trials have generally shown benefit from dietary n-3 polyunsaturated fatty acids. This has usually been attributed to their anti-inflammatory properties. Recently we have used in vitro model systems to study the molecular mechanism(s) by which n-3 PUFAs may act to alleviate the symptoms of arthritis. These experiments showed that n-3 PUFAs reduce expression of cartilage-degrading proteinases, cyclooxygenase-2 and inflammatory cytokines. Eicosapentaenoic acid (EPA) was more effective than docosahexaenoic acid (DHA) or alpha-linolenic acid. The data provide a scientific rationale for the consumption of n-3 fatty acids as part of a healthy diet and perhaps in treating arthritis. [Copyright &y& Elsevier]

Summary: Objective: To assess the relative efficacy of three different omega-3 (n-3) polyunsaturated fatty acids (PUFAs) in suppressing the mRNA levels for important proteins involved in the etiology of osteoarthritis (OA). Methods: A model cell culture system (bovine chondrocytes) was used. Inflammatory factors and enzymes involved in OA were induced by exposure of the chondrocyte cultures to interleukin-1α (IL-1α). The effect of pre-incubating cultures with various amounts of exogenous fatty acids on subsequent levels of mRNAs was assessed by reverse transcription-polymerase chain reactions (RT-PCR). Results: Exposure of cultures to IL-1α induced expression of the cartilage proteinases A Disintegrin And Metalloproteinase with ThromboSpondin motifs (ADAMTS)-4 and ADAMTS-5, cyclooxygenase (COX)-2, the matrix metalloproteinase (MMP)-3 and the inflammatory cytokines IL-1α, interleukin-1β (IL-1β) and tumour necrosis factor-α (TNF-α). n-3 PUFAs were able to reduce the levels of mRNA for ADAMTS-4, ADAMTS-5, MMP-3, MMP-13, COX-2 (but not COX-1), IL-1α, IL-1β and TNF-α. Eicosapentaenoic acid (EPA) was the most effective, followed by docosahexaenoic (DHA) and then α-linolenic (ALA) acid. The n-6 PUFA, arachidonic acid (AA) had no effect. Conclusion: These results show that omega-3 (n-3) PUFAs cause a reduction in the mRNA levels for various proteins known to be important in the pathology of OA. They provide a molecular explanation, at least in part, for beneficial effects of dietary omega-3 PUFAs for the amelioration of symptoms of the disease. The relative efficacy of EPA suggests that this omega-3 PUFA may be especially useful for dietary supplementation in patients with OA. [Copyright &y& Elsevier]

Summary: Objective: To determine whether chondroitin sulphate (CS) impedes the migration of primary articular chondrocytes. Design: Articular chondrocytes were isolated from young and skeletally mature bovine animals. Boyden chambers were used to quantify chondrocyte migration on aggrecan in the presence and absence of CS chains. A novel in vitro model of cell migration into articular cartilage explants was designed to visualise and quantify the migration of labelled chondrocytes into cartilage matrix which had been treated with chondroitinase ABC to remove CS chains present. Results: A consistent trend of increased migration with both age groups of a sub-population of chondrocytes was demonstrated on aggrecan in the absence of CS. These data were supported by results from the in vitro model of chondrocyte migration which demonstrated increasing numbers of a chondrocyte sub-population from both age groups of cartilage migrating into the chondroitinase ABC digested cartilage explants with time in culture. Minimal migration of these chondrocytes was demonstrated into phosphate buffered saline (PBS) treated control explants. Conclusions: We confirm that a sub-population of chondrocytes isolated from both young and skeletally mature articular cartilages have the ability to migrate. We also demonstrate that CS chains inhibit the migration of these articular chondrocytes and that their removal by chondroitinase ABC digestion enhances the migration of these chondrocytes. Such findings may provide a clinical application for improving cell-based cartilage repair strategies by enhancing integration between endogenous and repair tissue. [Copyright &y& Elsevier]

Summary: Objective: The objective of this study was to investigate CD44 and proteoglycan metabolism in patients suffering from Kashin–Beck Disease (KBD), an endemic osteoarthropathy that affects 2.5 million of 30 million people living in the KBD regions of China. Methods: Immunohistochemical analyses of cluster of differentiation-44 (CD44), BC-13 and 3-B-3(−) expression were performed in cartilage sections harvested from KBD and normal patients. In addition, the serum levels of soluble CD44 (sCD44), interleukin-1beta (IL-1β), tumor necrosis factor-alpha (TNF-α) and matrix metalloproteinase-1 were determined using a sandwich enzyme linked immunosorbent assay. Results: Hematoxylin & eosin and toluidine blue staining indicated that there was cell necrosis and proteoglycan loss in cartilage from both KBD children and adult cartilage. Strong immunohistochemical staining for CD44, BC-13 and 3-B-3(−) occurred in the majority of adult KBD patients and most KBD children. Furthermore, statistically significant elevated levels of sCD44, IL-1β and TNF-α were found in the sera of both adult and child KBD patients when compared to the levels of normal adult and child controls. Interestingly, IL-1β and TNF-α serum levels were all high in normal children from KBD regions when compared to normal children from non-KBD regions suggesting that unidentified factors (e.g., a genetic predisposition) may protect some people from KBD pathology. Conclusion: Our results demonstrate that altered CD44, IL-1β and TNF-α metabolism occurs in the pathogenesis of KBD and there is an increased aggrecanase-generated proteoglycan loss from KBD adult and child cartilage. These primary metabolic changes are likely to be significant contributing factor causing pathological joint formation and instability that leads to secondary osteoarthritis in KBD patients. [Copyright &y& Elsevier]

Summary: Objective: Oxidative stress occurs when the metabolic balance of a cell is disrupted through exposure to excess pro-oxidant. Whilst it is known that unregulated production or exposure to exogenous sources of pro-oxidants induces chondrocyte cell death and degrades matrix components in vitro, relatively little is known of the effects of pro-oxidants on articular cartilage in situ. The objective of this study was to determine if a single exposure to the pro-oxidant hydrogen peroxide (H2O2) induces a degenerative phenotype. Methods: Articular cartilage explants were obtained from skeletally mature bovine steers and exposed to a single dose of hydrogen peroxide (0.1–1.0mM) and cultured for up to 21 days. Cell death, and sulfated glycosaminoglycan loss into the medium and gene expression were quantitatively determined. Adoption of an abnormal chondrocyte phenotype was analyzed through the expression of 3B3(−), nitrotyrosine and procollagen type IIA epitopes in cartilage explants. Results: Cell death occurred primarily at the surface zone of cartilage in a dose-dependent manner in H2O2 treated explants, and supplementation of standard serum-free medium with insulin–selenium–transferrin significantly reduced cell death (>fourfold). Nitric oxide synthase-2 gene expression and proteoglycan loss increased in oxidant treated explants in a concentration-dependent manner. Antibody labeling to 3B3(−), procollagen type IIA and nitrotyrosine was present in all treated explants but absent in untreated explants. Conclusions: This study demonstrates that a single exposure to high levels of pro-oxidant causes the expression of genes and antibody epitopes that are associated with early degenerative changes observed in experimental osteoarthritis. [Copyright &y& Elsevier]