NT5E Genetic Mutation Is a Rare But Important Cause of Intermittent Claudication and Chronic Limb-Threatening Ischemia.

BACKGROUND: NT5Egenetic mutations are known to result in calcification of joints and arteries (CALJA), and worldwide, 14 patients from 7 families have been reported.Methods and Results:A total of 5 patients from 2 independent families with CALJA were found in Japan. Of them, 3 complained of intermittent claudication (IC), and 1 suffered from bilateral chronic limb-threatening ischemia (CLTI). Whole-exome sequencing analysis revealed an identical mutation pattern (c.G3C on the exon 1 start codon) that was unique compared withNT5Emutations reported in other countries. CONCLUSIONS: Vascular specialists need to recognize CALJA as a rare cause of ischemic IC and CLTI.

Synovial fluid matrix metalloproteinase-2 and -9 activities in dogs suffering from joint disorders.

The activity of matrix metalloproteinase (MMP)-2 and MMP-9 in synovial fluids (SF) sampled from dogs with joint disorders was investigated by gelatin zymography and densitometry. Pro-MMP-2 showed similar activity levels in dogs with idiopathic polyarthritis (IPA; n=17) or canine rheumatoid arthritis (cRA; n=4), and healthy controls (n=10). However, dogs with cranial cruciate ligament rupture (CCLR; n=5) presented significantly higher pro-MMP-2 activity than IPA and healthy dogs. Meanwhile, dogs with IPA exhibited significantly higher activity of pro- and active MMP-9 than other groups. Activity levels in pro- and active MMP-9 in cRA and CCLR dogs were not significantly different from those in healthy controls. Different patterns of MMP-2 and MMP-9 activity may reflect the differences in the underlying pathological processes.

Decreased performance in IDUA knockout mouse mimic limitations of joint function and locomotion in patients with Hurler syndrome.

BACKGROUND: Mucopolysaccharidosis type I (MPS I) is caused by the deficiency of alpha-L-iduronidase (IDUA), which is involved in the degradation of glycosaminoglycans (GAGs), such as heparan sulfate and dermatan sulfate in the lysosome. It has been reported that joint symptoms are almost universal in MPS I patients, and even in the case of attenuated disease, they are the first symptom that brings a child to medical attention. However, functional tests and biological markers have not been published for the evaluation of the limitations in joint and locomotion in animal model-mimicking MPS. METHODS: We generated IDUA knockout (KO) mice to observe whether they present impairment of joint function. KO mice were characterized phenotypically and tested dual-energy X-ray absorptiometry analysis (DEXA), open-field, rotarod, and grip strength. RESULTS: The IDUA KO mice, generated by disruption between exon 6 and exon 9, exhibited clinical and laboratory findings, such as high urinary GAGs excretion, GAGs accumulation in various tissues, and significantly increased bone mineral density (BMD) in both female and male mice in the DEXA of the femur and whole bone. Remarkably, we observed a decrease in grasp function, decreased performance in the rotarod test, and hypo-activity in the open-field test, which mimic the limitations of joint mobility and decreased motor performance in the 6-min walk test in patients with MPS I. CONCLUSIONS: We generated a new IDUA KO mouse, tested open field, rotarod and grip strength and demonstrated decrease in grip strength, decreased performance and hypo-activity, which may be useful for investigating therapeutic approaches, and studying the pathogenesis of joint and locomotion symptoms in MPS I.

ADAMTS-4 activity in synovial fluid as a biomarker of inflammation and effusion.

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.

Equine myeloperoxidase: a novel biomarker in synovial fluid for the diagnosis of infection.

REASONS FOR PERFORMING STUDY: Equine joint infection is a life-threatening disorder, and confirmation of the diagnosis can be difficult. Synovial fluid biomarkers may assist the discrimination between infectious and noninfectious joint disease. OBJECTIVES: This study investigates whether the immunological detection of total and enzymatically active myeloperoxidase (MPO) assists the diagnosis of joint infection in horses. METHODS: The following 4 sample groups were included: healthy; osteochondritis dissecans (OCD); traumatic synovitis; and culture-confirmed infected joints. Synovial fluid was analysed for total MPO by a horse-specific sandwich enzyme-linked immunosorbent assay (ELISA) and for active MPO using the specific immunological extraction followed by enzymatic detection (SIEFED) technique. Western blot analysis was performed to confirm the antibody specificity. RESULTS: Synovial fluid from infected joints contained significantly more total and active MPO than samples from healthy joints, joints affected by OCD and joints with traumatic synovitis. Cut-off values were set at 5000 and 350 ng/ml for total and active MPO, respectively, with fair sensitivity, specificity, positive and negative predictive values and likelihood ratios for infection. Correlation coefficients were reported between the total as well as the active MPO levels and the routine synovial fluid parameters, i.e. the white blood cell count, the neutrophil count and the total protein level. No correlation was observed between MPO and either the age of the horse or the joint affected. Western blotting confirmed the antibody specificity for equine MPO. CONCLUSIONS AND POTENTIAL RELEVANCE: Synovial fluid MPO was identified as a very promising biomarker to augment the discrimination of infectious vs. noninfectious joint disease in horses. Both ELISA and SIEFED techniques can be used for its specific and rapid detection. The analysis of synovial fluid MPO can be used as a complementary test to aid in the discrimination between infectious and noninfectious joint disease, especially when the white blood cell counts and the total protein level are inconclusive.

Chondrodysplasia and abnormal joint development associated with mutations in IMPAD1, encoding the Golgi-resident nucleotide phosphatase, gPAPP.

We used whole-exome sequencing to study three individuals with a distinct condition characterized by short stature, chondrodysplasia with brachydactyly, congenital joint dislocations, cleft palate, and facial dysmorphism. Affected individuals carried homozygous missense mutations in IMPAD1, the gene coding for gPAPP, a Golgi-resident nucleotide phosphatase that hydrolyzes phosphoadenosine phosphate (PAP), the byproduct of sulfotransferase reactions, to AMP. The mutations affected residues in or adjacent to the phosphatase active site and are predicted to impair enzyme activity. A fourth unrelated patient was subsequently found to be homozygous for a premature termination codon in IMPAD1. Impad1 inactivation in mice has previously been shown to produce chondrodysplasia with abnormal joint formation and impaired proteoglycan sulfation. The human chondrodysplasia associated with gPAPP deficiency joins a growing number of skeletoarticular conditions associated with defective synthesis of sulfated proteoglycans, highlighting the importance of proteoglycans in the development of skeletal elements and joints.

Development of an in vitro model to investigate joint ochronosis in alkaptonuria.

OBJECTIVES: Alkaptonuria (AKU) is a genetic disorder caused by lack of the enzyme responsible for breaking down homogentisic acid (HGA), an intermediate in tyrosine metabolism. HGA is deposited as a polymer, termed ochronotic pigment, in collagenous tissues. Pigmentation is progressive over many years, leading to CTDs including severe arthropathies. To investigate the mechanism of pigmentation and to determine how it leads to arthropathy, we aimed to develop an in vitro model of ochronosis. METHODS: Osteosarcoma cell lines MG63, SaOS-2 and TE85 were cultured in medium containing HGA from 0.1 µM to 1 mM. Cultures were examined by light microscopy and transmission electron microscopy, and Schmorl's stain was used to detect pigment deposits in vitro, following the observation that this stain identifies ochronotic pigment in AKU tissues. The effects of HGA on cell growth and collagen synthesis were also determined. RESULTS: There was a dose-related deposition of pigment in cells and associated matrix from 33 µM to 0.33 mM HGA. Pigmentation in vitro was much more rapid than in vivo, indicating that protective mechanisms exist in tissues in situ. Pigment deposition was dependent on the presence of cells and was observed at HGA concentrations that were not toxic. There was an inhibition of cell growth and a stimulation of type I collagen synthesis up to 0.33 mM HGA, but severe cell toxicity at 1 mM HGA. CONCLUSION: We have developed an in vitro model of ochronosis that should contribute to understanding joint destruction in AKU and to the aetiology of OA.

Apoptosis of ligamentous cells of the cranial cruciate ligament from stable stifle joints of dogs with partial cranial cruciate ligament rupture.

OBJECTIVE: To describe the presence and amount of apoptotic ligamentous cells in different areas of partially ruptured canine cranial cruciate ligaments (prCCLs) and to compare these findings with apoptosis of ligamentous cells in totally ruptured cranial cruciate ligaments (trCCLs). ANIMALS: 20 dogs with prCCLs and 14 dogs with trCCLs. PROCEDURES: Dogs with prCCLs or trCCLs were admitted to the veterinary hospital for stifle joint treatment. Biopsy specimens of the intact area of prCCLs (group A) and the ruptured area of prCCLs (group B) as well as specimens from trCCLs (group C) were harvested during arthroscopy. Caspase-3 and poly (ADP-ribose) polymerase (PARP) detection were used to detect apoptotic ligamentous cells by immunohistochemistry. RESULTS: No difference was found in the degree of synovitis or osteophytosis between prCCLs and trCCLs. No difference was found in degenerative changes in ligaments between groups A and B. A substantial amount of apoptotic cells could be found in > 90% of all stained slides. A correlation (r(s) = 0.71) was found between the number of caspase-3-and PARP-positive cells. No significant difference was found in the amount of apoptotic cells among the 3 groups. No significant correlation could be detected between the degree of synovitis and apoptotic cells or osteophyte production and apoptotic cells. CONCLUSIONS AND CLINICAL RELEVANCE: The lack of difference between the 3 groups indicates that apoptosis could be a factor in the internal disease process leading to CCL rupture and is not primarily a consequence of the acute rupture of the ligament.

Joint diseases and matrix metalloproteinases: a role for MMP-13.

The role of matrix metalloproteinases in disease has been investigated over the last two decades. A focus on this family of proteases is particularly emphasized in two major arthritides in humans, osteoarthritis and rheumatoid arthritis. Early work described the presence of multiple MMP family members in the joint of the disease state and recent advances in the development of new knockout mice and disease models have allowed investigators to directly test the role of the MMP proteases in arthritis. MMP-13 is expressed by chondrocytes and synovial cells in human OA and RA and is thought to play a critical role in cartilage destruction. The recent development of an MMP-13 knockout mouse has documented the important role for this enzyme in cartilage formation and further studies under disease conditions promise to reveal the function of this enzyme in disease pathology. This review describes a body of research that supports the development of novel selective MMP-13 inhibitors with the hope of developing these compounds in clinical trials for the treatment of arthritis.

Determination of MMP-2 and -9 activities in synovial fluid of horses with osteoarthritic and arthritic joint diseases using gelatin zymography and immunocapture activity assays.

REASON FOR PERFORMING STUDY: Matrix metalloproteinases (MMPs)-2 and -9 activities have been found elevated in synovial fluid from various joint diseases in man. However, in the horse few data are available. OBJECTIVES: To explore the clinical significance of MMP-2 and -9 activities in synovial fluid of horses with different forms of joint diseases. METHODS: Gelatin zymography and MMP-2 and -9 immunocapture activity assays were applied on synovial fluids from control joints and joints with aseptic joint disease (AJD) and septic arthritis (SA). Additionally, MMP-2 and -9 activities were measured in samples from SA to monitor the disease process. RESULTS: Zymographic analysis revealed that samples from AJD and SA contained significantly increased latent MMP-2 activity compared to controls. Samples from SA showed significantly increased monomeric latent MMP-9 activity compared with all other affected joints and controls. Trace amounts of MMP-9 activity, due to the active and dimer form, were detected in samples from SA; however, these bands were absent in samples from AJD and controls. Using immunocapture activity assays, MMP-2 and -9 activities were found to be significantly elevated in joints from SA compared to controls and AJD samples. MMP-2 activity in samples from AJD was significantly increased compared to controls. Both MMP activities decreased in the joints from SA in the course of successful therapy. CONCLUSIONS: Data from zymographic analysis confirmed that MMP-2 and -9 were elevated in equine joint diseases. Immunocapture activity assays have been shown to be suitable for the quantitative determination of MMP-2 and -9 activities in synovial fluid of horses. Both MMP-2 and -9 activities seem to be useful to indicate SA, and MMP-2 activity might be a suitable marker for AJD. POTENTIAL RELEVANCE: These findings encourage the potential use of MMP-2 and -9 as additional aids to clinical investigation. Further work is required to validate the clinical significance of MMP activities in the progress of different joint diseases in horses.

Measurement of equine myeloperoxidase (MPO) activity in synovial fluid by a modified MPO assay and evaluation of joint diseases - an initial case study.

The aim of this study was to develop a specific myeloperoxidase (MPO) activity assay in the synovial fluid of horses and investigate whether MPO activity is increased in different forms of joint diseases. Synovial fluid samples were taken from affected joints from horses with osteoarthritis, chronic non-septic arthritis and septic arthritis, and from healthy control horses. MPO activity was measured using a specific modified o-dianisidine-assay containing 4-aminobenzoic acid hydrazide as a potent and specific inhibitor of the MPO. This assay is characterized by high reproducibility. The results reveal only a slight elevation of MPO activity in the synovial fluid of horses with osteoarthritis and chronic non-septic arthritis. However, in the cases of septic arthritis a significant increase in MPO activity was found when compared to the controls. In conclusion the first field study suggests that synovial fluid MPO may be used as a marker for septic arthritis in horses.

Activity of N-acetyl-beta-hexosaminidase and its isoenzymes in serum and synovial fluid from patients with different arthropathies.

OBJECTIVE: To evaluate the activity of N-acetyl-Beta-hexosaminidase (HEX) and its isoenzymes in the serum and synovial fluid of healthy volunteers and patients with an injury to the anterior cruciate ligament and/or meniscus (ACL) osteoarthritis (OA), juvenile idiopathic arthritis (JIA) and rheumatoid arthritis (RA). METHODS: The activity of HEX and its isoenzymes was determined according to Zwierz et al. method. Protein content was determined by the biuret method. RESULTS: The specific activity of HEX and its isoenzymes in the serum of patients with JIA showed a tendency to increase in comparison to the reference group. The specific activity of total HEX in the serum of RA patients was significantly increased in comparison to control. Our results show, that specific activity of HEX in synovial fluid, in the reference group 4.2 +/- 0.21 microkat/kg protein (0.25 unit/mg protein), is similar to activity in normal temporomandibular joint fluid (0.3 unit/mg protein). Therefore, we included this group in our research. In patients with OA and ACL injuries, HEX and its isoenzymes showed a tendency to increase in the specific activity in synovial fluid. The specific activity of HEX and its isoenzymes in the synovial fluid of patients with RA and JIA was significantly elevated in comparison to the control and the remaining groups. CONCLUSION: In the synovial fluid of patients with JIA and RA, the specific activity of HEX and its isoenzymes significantly increased in comparison to control and patients with diseases of a non-inflammatory etiology (OA and ACL). In the synovial fluid of control and diseased groups, HEX constituted a higher percent of total proteins than in serum.

Localization of cathepsin K and tartrate-resistant acid phosphatase in synovium and cranial cruciate ligament in dogs with cruciate disease.

OBJECTIVE: To localize cathepsin K and tartrate-resistant acid phosphatase (TRAP) in synovium and cranial cruciate ligament (CCL) of dogs with cruciate disease. ANIMALS: Dogs (n=15) with cruciate disease and ruptured CCL, and 12 dogs with intact CCL. METHODS: Synovium and CCL were examined histologically and cells containing cathepsin K or TRAP were identified immunohistochemically and histochemically, respectively. RESULTS: Increased cellular localization of cathepsin K and TRAP was detected in synovium and ruptured CCL in dogs with cruciate disease, when compared with tissues from dogs with intact CCL. Inflammation of synovium with TRAP+ macrophage-like cells was seen in 73% of dogs with CCL disease, but was not seen in dogs with intact CCL. The presence of cathepsin K and TRAP protein in synovium and CCL tissues was significantly correlated in dogs with CCL rupture. CONCLUSION: Inflammation of the epiligament of ruptured CCL with cathepsin K+ and TRAP+ macrophage-like cells forms part of a similar, more generalized chronic inflammatory change within the periarticular tissues of the stifle of a large proportion of dogs with CCL rupture. CLINICAL RELEVANCE: Production of matrix-degrading enzymes by the synovium may induce progressive pathologic rupture of the CCL. Therefore, these collagenolytic pathways may offer a novel target for medical therapy of joint inflammation in canine patients with cruciate disease.

Degenerative knee joint disease in mice lacking 3'-phosphoadenosine 5'-phosphosulfate synthetase 2 (Papss2) activity: a putative model of human PAPSS2 deficiency-associated arthrosis.

OBJECTIVE: Murine brachymorphism (bm) results from an autosomal recessive mutation of the Papss2 gene that encodes 3'-phosphoadenosine 5'-phosphosulfate synthetase 2, one of the principal enzymes required for the sulfation of extracellular matrix molecules in cartilage and other tissues. A spondyloepimetaphyseal dysplasia has been identified in Pakistani kindred having a mutation of PAPSS2. In addition to skeletal malformations that include short stature evident at birth due to limb shortening, brachydactyly, and kyphoscoliosis, affected individuals demonstrate premature onset degenerative joint disease. We investigated whether loss of Papss2 activity would similarly lead to degenerative joint disease in mice. METHODS: Mice carrying the bm mutation on a C57BL/6 background were obtained from the Jackson Laboratory. Limbs were analyzed by micro-computed tomography (microCT) and histology. RESULTS: At 12 months of age both male and female bm mice exhibited severe degenerative knee joint disease, with cartilage damage being primarily evident in the patello-femoral and medial compartments. Control 12-14-month-old C57BL/6 mice, in contrast, only occasionally demonstrated minimal cartilage damage. muCT imaging of bm limbs revealed shortened diaphyses associated with flared metaphyses in the proximal elements of both fore and hind limbs. Additionally, the bm hind limbs demonstrated extensive structural alterations, characterized by distortion of the patello-femoral groove, and prominent bowing of both tibia and fibula. CONCLUSIONS: The bm mutant, which develops severe articular cartilage lesions of the knee joint by approximately 12 months of age, represents a novel example of murine degenerative joint disease, possibly representing a model of human PAPSS2 deficiency-associated arthrosis.

The influence of repeated arthrocentesis and exercise on matrix metalloproteinase and tumour necrosis factor alpha activities in normal equine joints.

REASONS FOR PERFORMING STUDY: Matrix metalloproteinases (MMPs) and tumour necrosis factor alpha (TNF-alpha) may be useful as biomarkers of joint disease or inflammation. However, activity of both MMPs and TNF-alpha in synovial fluid (SF) may be influenced by nonpathological factors such as arthrocentesis or exercise. OBJECTIVE: To investigate the influence of repeated arthrocentesis and exercise on MMP and TNF-alpha activities in SF from normal equine joints. METHODS: SF was collected from the left metacarpophalangeal, radiocarpal and tarsocrural joints of 16 horses. Eight of these horses were subsequently subjected to an exercise programme on a treadmill and 8 were box-rested as controls. Arthrocentesis was repeated 14, 145, 17 and 24 days after the start of the exercise programme. General MMP and TNF-alpha activities were determined in SF. RESULTS: Repeated arthrocentesis caused a gradual increase but the exercise regimen no significant increase in MMP activity. There was a significant increase in TNF-alpha activity in SF collected from horses 2 h after cessation of the exercise programme. CONCLUSIONS AND POTENTIAL RELEVANCE: When using MMPs as biomarkers for joint disease, at least 14 days should elapse after previous arthrocentesis before subsequent SF collection. Moderate exercise does not increase MMP activity in SF from normal joints and it may be possible to ignore this as a source of error in evaluating MMP activity in diseased joints.

Alkaptonuria, ochronosis, and ochronotic arthropathy.

OBJECTIVES: To describe the clinical presentation and course of a relatively large group of Italian adult patients screened for mutation of the homogentisate dioxygenase gene causing alkaptonuria (AKU) and ochronosis, and to review typical and atypical facets of this condition. METHODS: We reviewed the medical records of 9 patients affected by ochronotic arthropathy who were observed in our institutions between 1979 and 2001. All patients were diagnosed as having AKU through a rapid urine test with alkali. Mutation screening was performed by single-strand conformation analysis of all homogentisate dioxygenase exons, followed by sequencing of altered conformers. RESULTS: Our 9 cases had similar clinical features and they reflected those described in the literature: a progressive degenerative arthropathy mainly affecting axial and weight-bearing joints associated with extraarticular manifestation. Musculoskeletal symptoms began in most of our patients around the age of 30 years with back pain and stiffness: involvement of the large peripheral joints usually occurred several years after spinal changes. Ochronotic peripheral arthropathy generally was degenerative, but joint inflammation was observed in some cases; this could be attributed to an inflammatory reaction of the ochronotic shard in the synovial membrane. CONCLUSIONS: Ochronosis is a model of arthropathy with known etiologic factors. Over time, AKU, the genetically determined metabolic defect, leads to the accumulation of pigment and the development of this crippling condition. Most of the clinical findings may be explained by inhibition of collagen crosslinks, but some require additional interpretation. For example, inflammatory features of the ochronotic joint only occur in a minority of cases, and may be attributable to ochronotic shards. Further studies are needed to establish the genotype-phenotype correlation to identify mutations that are predictive of severe disease. For this purpose, the Italian Study Group on Alkaptonuria (www.dfc.unifi.it/aku) is enrolling affected patients in an on-line database to characterize the molecular defects and their relationship to clinical data.

Zymographic analysis of latent and activated forms of matrix metalloproteinase-2 and -9 in synovial fluid: correlation to polymorphonuclear leukocyte infiltration and in response to infection.

BACKGROUND: Matrix metalloproteinase-2 and-9 (MMP-2, MMP-9), and gelatinase A and B participate in the degradation of the extracellular matrix proteins in a variety of inflammatory connective tissue diseases including arthritis. METHODS: Synovial fluid was collected by aseptic aspiration from patients with rheumatoid arthritis (RA), osteoarthritis (OA), gout, infected joint, septic arthritis, and systemic lupus erythematosus (SLE). Synovial fluid was subjected to cell count with polymorphonuclear leukocyte (PMN) differential, Gram staining and culture as necessary. MMP-2 and -9 were characterized by substrate gel electrophoresis (gelatin zymography) to resolve latent and activated 'partially proteolyzed' forms. RESULTS: Gelatin zymography revealed that MMP-9 (92, 130, 225 kDa) in synovial fluid was associated with extent of white blood cell infiltration specifically PMNs. In contrast, fibroblast MMP-2 (72 kDa) was present in all synovial fluids irrespective of PMN count. No MMP-9 was detected in the osteoarthritic specimen with low PMN count. Higher PMN count was associated with the presence of activated MMPs, especially in specimens that were confirmed culture positive. Activated synovial fluid MMPs persisted despite resolution of infection. DISCUSSION: Latent and activated MMP-2 and MMP-9 in synovial fluids fluctuate in proportion to PMN infiltration and specifically in response to infection. The presence of activated MMPs post-therapy would suggest that use of specific MMP inhibitors be indicated to eliminate activated MMPs that apparently persist post-infection.

Synovial fluid gelatinase concentrations and matrix metalloproteinase and cytokine expression in naturally occurring joint disease in horses.

OBJECTIVES: To determine concentrations of matrix metalloproteinase (MMP)-2 and -9 in synovial fluid; and mRNA expression of MMP-1, -13, and -3; interleukin[IL]-1alpha and beta; and tumor necrosis factor (TNF)-alpha in synovial membrane and articular cartilage from horses with naturally occurring joint disease. SAMPLE POPULATION: Synovial fluid (n = 76), synovial membrane (59), and articular cartilage (45) from 5 clinically normal horses and 55 horses with joint disease categorized as traumatic (acute [AT] or chronic [CT]), osteochondritis dissecans (OCD), or septic (S). PROCEDURE: Synovial fluid gelatinase concentrations were analyzed, using zymography. Synovial membrane and articular cartilage mRNA expression for MMP-1, -3, and -13, IL-1alpha and beta, TNF-alpha, type-II collagen, and aggrecan were analyzed, using quantitative reverse transcriptase-polymerase chain reaction. RESULTS: Synovial fluid pro-MMP-2 concentration was significantly higher in diseased joints than normal joints. Septic joints had significantly higher concentrations of pro and active MMP-9. Stromelysin-1 was expressed in > or = 80% of synovial membrane and articular cartilage samples and was strongly influenced by age. Collagenases were rarely expressed, with MMP-13 expressed only in diseased joints. Interleukin-1beta expression was significantly higher in all OCD samples and was influenced by age. Tumor necrosis factor-alpha expression was significantly higher in cartilage from joints with AT and OCD. There was no correlation between MMP or cytokines and type-II collagen or aggrecan expression. CONCLUSIONS AND CLINICAL RELEVANCE: Matrix metalloproteinase-2 and -3 are abundant in naturally occurring joint disease and normal joints. Interleukin-1beta and TNF-alpha may be important in the pathogenesis of OCD. Age affects MMP and IL-1beta concentrations.

Diabetic complications are associated with liver enzyme activities in people with type 1 diabetes.

This study was performed to clarify if diabetic complications are associated with liver enzyme activities in type 1 diabetic outpatients. Elevated activities of serum aminotransferases are a common sign of liver disease and are observed more frequently among people with diabetes than in the general population. Many studies have shown an association between specific diabetic complications and disturbances in various tissues, such as diabetic nephropathy and cardiovascular diseases, but only limited data are available on the possible association between diabetic complications and liver function. We studied 28 patients with type 1 diabetes. Mean age was 43.4+/-9.5 (S.D.), and duration of diabetes 25.2+/-9.7. Limited joint mobility (LJM) was assessed by the Rosenbloom's method. Background and proliferative retinopathy, and peripheral symmetrical polyneuropathy were also assessed. Activities of alanine amino transferase (ALT), gamma-glutamyl transferase (GGT) and alkaline phosphatase (ALP) in serum were determined. The metabolic control of the diabetes was evaluated by the glycosylated haemoglobin A(1c) (HbA(1c)) level and lipid values were also measured. ALT activity was associated with LJM (P<0.01) and with neuropathy (P<0.01). Association between GGT activity and LJM (P<0.01) and neuropathy (P<0.01) were also found. GGT activity was also associated with the severity of retinopathy (P<0.01). None of these associations was explained by confounding effects of diabetes duration, age, body mass index (BMI), HbA(1c) or alcohol consumption. In conclusion, diabetic complications such as LJM, retinopathy and neuropathy are associated with liver enzyme activities independent of alcohol consumption, BMI and metabolic control of diabetes.

Serum creatine kinase in patients with rheumatic diseases.

Serum creatine kinase (CK) activity is reduced in some conditions, including rheumatic diseases, but the aetiology and significance remain to be clarified. The aim of this study was to investigate relationships between serum CK activity and other muscle enzymes, muscle mass, renal function, steroid use and disease activity in patients with rheumatic diseases. Serum CK activity was measured in sera from 498 patients with rheumatic diseases: rheumatoid arthritis (RA, n = 145), systemic lupus erythematosus (SLE, n = 31), spondyloarthropathies (SpA, n = 35), polyarthralgia/arthritis (Poly, n = 74), miscellaneous group (MI, n = 46), and in non-inflammatory arthropathies (NIA, n = 167) as controls. Serum CK level was significantly reduced in RA (45.4 +/- 1.9 IU/l), SLE (46.4 +/- 4.2 IU/l), SpA (64.7 +/- 5.6 IU/l) and MI (63.4 +/- 4.8 IU/l), but not in poly (70.2 +/- 3.1 IU/l), compared to controls (78.9 +/- 2.4 IU/l) (P < 0.05). CK values correlated with aspartate aminotransferase (AST), erythrocyte sedimentation rate (ESR), body mass index (BMI) and platelets (Plat) in RA; ESR and haemoglobin (Hb) in SLE, AST, ESR and Hb in SpA; lactate dehydrogenase (LDH), AST, ESR and Hb in Poly; LDH, AST, ESR, Hb and Ccr in MI; and LDH, AST and ESR in controls. In all patients with rheumatic diseases CK level was significantly correlated with LDH, AST, alanine aminotransferase (ALT), ESR, C-reactive protein (CRP) and BMI and prednisolone dose, but not with Ccr, age and disease duration. In conclusion, our data support the possibility that reduced CK activity is inversely correlated with inflammatory activity and correlated with other muscle enzymes, muscle mass and steroid use, but not with renal function, age and disease duration in rheumatic diseases.