Superoxide dismutase activity is significantly lower in end-stage osteoarthritic cartilage than non-osteoarthritic cartilage.

Recent studies have shown that superoxide dismutase 1 (SOD1), SOD2, and SOD3 are significantly decreased in human osteoarthritic cartilage. SOD activity is a marker that can be used to comprehensively evaluate the enzymatic capacities of SOD1, SOD2, and SOD3; however, the trend of SOD activity in end-stage osteoarthritic tissues remains unknown. In the present study, we found that SOD activity in end-stage osteoarthritic synovium of the knee was significantly lower than that in control synovium without the influence of age. The SOD activity was significantly lower in the end-stage knee osteoarthritic cartilage than in the control, but a weak negative correlation was observed between aging and SOD activity. However, SOD activity in end-stage hip osteoarthritic cartilage was significantly lower than that in control cartilage without the influence of aging. The relationship between osteoarthritis and SOD activity was stronger than the relationship between aging and SOD activity. These results indicate that direct regulation of SOD activity in joint tissues may lead to suppression of osteoarthritis progression.

Functional Characterization of the Osteoarthritis Susceptibility Mapping to CHST11-A Bioinformatics and Molecular Study.

The single nucleotide polymorphism (SNP) rs835487 is associated with hip osteoarthritis (OA) at the genome-wide significance level and is located within CHST11, which codes for carbohydrate sulfotransferase 11. This enzyme post-translationally modifies proteoglycan prior to its deposition in the cartilage extracellular matrix. Using bioinformatics and experimental analyses, our aims were to characterise the rs835487 association signal and to identify the causal functional variant/s. Database searches revealed that rs835487 resides within a linkage disequilibrium (LD) block of only 2.7 kb and is in LD (r2 ≥ 0.8) with six other SNPs. These are all located within intron 2 of CHST11, in a region that has predicted enhancer activity and which shows a high degree of conservation in primates. Luciferase reporter assays revealed that of the seven SNPs, rs835487 and rs835488, which have a pairwise r2 of 0.962, are the top functional candidates; the haplotype composed of the OA-risk conferring G allele of rs835487 and the corresponding T allele of rs835488 (the G-T haplotype) demonstrated significantly different enhancer activity relative to the haplotype composed of the non-risk A allele of rs835487 and the corresponding C allele of rs835488 (the A-C haplotype) (p < 0.001). Electrophoretic mobility shift assays and supershifts identified several transcription factors that bind more strongly to the risk-conferring G and T alleles of the two SNPs, including SP1, SP3, YY1 and SUB1. CHST11 was found to be upregulated in OA versus non-OA cartilage (p < 0.001) and was expressed dynamically during chondrogenesis. Its expression in adult cartilage did not however correlate with rs835487 genotype. Our data demonstrate that the OA susceptibility is mediated by differential protein binding to the alleles of rs835487 and rs835488, which are located within an enhancer whose target may be CHST11 during chondrogenesis or an alternative gene.

Lysosomal enzymes in serum and synovial fluid in patients with osteoarthritis.

OBJECTIVE: To assess the activity of arylsulfatase (AS), acid phosphatase (ACP), cathepsin D (CAT-D) and alpha-1 antitrypsin (AAT) in blood serum and synovial fluid (SF) of patients with hip or knee osteoarthritis (OA). METHODS: The study included 43 subjects with OA (35 hip OA, 8 knee OA), submitted total joint replacement. The control group consisted of 58 subjects with no past history of musculoskeletal disorders. RESULTS: The OA blood serum samples showed a significantly higher level of lysosomal enzymes activity than in the control group (AS by 17.8%, AAT by 42.4%); only the CAT-D activity decreased by 50%). AS, ACP and CAT-D activities were about two-fold higher in SF when compared with blood of OA patients. The differences between the genders were visible in the SF: Total protein concentration, activity of ACP (both higher in OA men) and activity of CAT-D (higher in OA women). Between the involved hip and knee, there were no significant differences in all estimated parameters in the blood serum of the OA group. In regard to the SF, only ACP activity was significantly higher in patients with a hip involved. CONCLUSIONS: The osteoarthritic SF enzymatic profile differs from that in normal joints. The OA in joints is not reflected in the systemic response. Our preliminary results suggest further studies on role of lysosomal enzymes (ACP and AS) as biomarkers for the detection of osteoarthritis.

Expression of gelatinases (MMP-2, MMP-9) in human articular cartilage.

Osteoarthritis (OA) is a chronic degenerative joint disorder characterized by destruction of the articular cartilage, subchondral bone alterations and synovitis. Matrix metalloproteinases (MMPs) are expressed in joint tissues of patients with osteoarthritis (OA). The objective of this study was to define the steady state levels of two different MMPs to provide more insight into the role of MMPs in cartilage destruction in OA. We investigated the expression of gelatinases through immunohistochemistry Our results show that high levels of MMP-2 and MMP-9 are present in OA and suggest that once these MMPs are fully activated they may contribute to the cartilage destruction in OA.

Subchondral cyst development and MMP-1 expression during progression of osteoarthritis: an immunohistochemical study.

BACKGROUND: Subchondral bone cyst (SBC) formation is often identified in patients with osteoarthritis. Furthermore, several studies have shown that expression of matrix metalloproteinases (MMPs) is elevated in patients with OA. OBJECTIVES: The aim of our study is to correlate the presence of SBCs and MMP-1 expression with the osteochondral alterations during OA progression. METHODS: We studied the cartilage and subchondral bone of 15 patients who had undergone total knee or hip replacement due to primary OA. As controls, we used the femoral heads of three patients without macroscopic OA changes. We evaluated three specimens per patient. RESULTS: Specimens were divided in four groups based on the Mankin histological severity score. Using immunohistochemistry, we noted SBCs at the site of greatest disease severity. Specifically, these were present more frequently in group III (Mankin score: 6-7) and IV (Mankin: ≥ 8), compared with group I (Mankin: 1-3) and II (Mankin: 4-5). Mild OA stages (Mankin: 1-6) were characterized by degeneration and thinning of the cartilage, followed by increased osteoblast and osteoclast activity of the subjacent bone and the subsequent appearance of SBCs. Simultaneously, we observed expression of MMP-1 in groups I and II in the cartilage and III and IV in both the cartilage and the subchondral bone. Moreover, osteoblast-like cells in the lining of the SBCs showed an increased expression of MMP-1 in stages III and IV. CONCLUSION: Our study provides immunohistological evidence that SBCs accumulate in advanced OA and contain activated cells, which express MMP-1, suggesting that they may thus participate in the osteochondral changes of OA. LEVEL OF EVIDENCE: Level III; prospective comparative study.

[Features of biochemical changes in blood serum according to the form of progression of idiopathic and dysplastic coxarthrosis].

Carried out biochemical studies of blood serum (collagenase activity, glycosamineglicans and hydroxyproline fractions) of 72 patients with hip idiopathic osteoarthrosis and 30 patients with dysplastic osteoarthrosis of the iv-th stage in.accordance with J. H. Kellgren and J. S. Lavrence depending on the form of pathologic process progression. It has been proved that both with idiopathic and dysplastic coxartrosis metabolism of basic protein of osteochonrous tissue was broken both in catabolic and in synthetic phase of this process. The most deep changes 1 of biochemical values (collagenase, free and proteinbinded hydroxyprolines, the content of glycosamines) have been observed with rapid form of course progression of idiopathic and dysplastic coxarthrosis. Definite appropriateness promotesin.patients better understanding of coxarthrosis pathogenese, development of diagnostic and medical measures for patients with this severe orthopedic pathology.

Increased type II deiodinase protein in OA-affected cartilage and allelic imbalance of OA risk polymorphism rs225014 at DIO2 in human OA joint tissues.

OBJECTIVE: Genetic variation at the type II deiodinase (D2) gene (DIO2) was previously identified as osteoarthritis (OA) risk factor. To investigate mechanisms possibly underlying this association, we assessed D2 protein in healthy and OA-affected cartilage and investigated allelic balance of the OA risk polymorphism rs225014 at DIO2 in human OA joints. METHODS: Immunohistochemical staining of healthy and OA-affected cartilage was performed for D2. We then assessed allelic balance of DIO2 mRNA within OA-affected cartilage both at and away from the lesion, ligaments and subchondral bone. Allelic balance was measured by the amount of alleles 'C' and 'T' of the intragenic OA risk polymorphism rs225014 in heterozygous carriers. RESULTS: A markedly higher amount of D2 positive cells and staining intensity was observed in OA cartilage. A significant, 1.3-fold higher presence was observed for the OA-associated rs225014 'C' allele relative to the 'T' allele of DIO2, which was significant in 28 of 31 donors. CONCLUSION: In OA cartilage, D2 protein presence is increased. The allelic imbalance of the DIO2 mRNA transcript, with the OA risk allele 'C' of rs225014 more abundant than the wild-type 'T' allele in heterozygote carriers provides a possible mechanism by which genetic variation at DIO2 confers OA risk.

Mitochondrial DNA (mtDNA) haplogroups and serum levels of anti-oxidant enzymes in patients with osteoarthritis.

BACKGROUND: Oxidative stress play a main role in the initiation and progression of the OA disease and leads to the degeneration of mitochondria. To prevent this, the chondrocytes possess a well-coordinated enzymatic antioxidant system. Besides, the mitochondrial DNA (mtDNA) haplogroups are associated with the OA disease. Thus, the main goal of this work is to assess the incidence of the mtDNA haplogroups on serum levels of two of the main antioxidant enzymes, Manganese Superoxide Dismutase (Mn-SOD or SOD2) and catalase, and to test the suitability of these two proteins for potential OA-related biomarkers. METHODS: We analyzed the serum levels of SOD2 and catalase in 73 OA patients and 77 healthy controls carrying the haplogroups J, U and H, by ELISA assay. Knee and hip radiographs were classified according to Kellgren and Lawrence (K/L) scoring from Grade 0 to Grade IV. Appropriate statistical analyses were performed to test the effects of clinical variables, including gender, body mass index (BMI), age, smoking status, diagnosis, haplogroups and radiologic K/L grade on serum levels of these enzymes. RESULTS: Serum levels of SOD2 appeared statistically increased in OA patients when compared with healthy controls (p < 0.001). Even in those OA patients with higher OA severity (K/L grade IV), the serum levels of this antioxidant enzyme appeared more significantly increased than in OA patients with lower K/L grade (p < 0.001). The mtDNA haplogroups showed an influence on serum levels of catalase (p = 0.054), being carriers of the mtDNA haplogroup J those who showed higher serum levels than non-J carriers (p = 0.057). CONCLUSIONS: The increased levels of SOD2 in OA patients indicate an increased oxidative stress OA-related, therefore this antioxidant enzyme could be a suitable candidate biomarker for diagnosis of OA. Mitochondrial haplogroups significantly correlates with serum levels of catalase.

The role of reactive oxygen species scavenging enzymes in the development of septic loosening after total hip replacement.

In this study of 41 patients, we used proteomic, Western blot and immunohistochemical analyses to show that several reactive oxygen species scavenging enzymes are expressed differentially in patients with primary osteoarthritis and those with non-loosening and aseptic loosening after total hip replacement (THR). The patients were grouped as A (n = 16, primary THR), B (n = 10, fixed THR but requiring revision for polyethylene wear) and C (n = 15, requiring revision due to aseptic loosening) to verify the involvement of the identified targets in aseptic loosening. When compared with Groups A and B, Group C patients exhibited significant up-regulation of transthyretin and superoxide dismutase 3, but down-regulation of glutathione peroxidase 2 in their hip synovial fluids. Also, higher levels of superoxide dismutase 2 and peroxiredoxin 2, but not superoxide dismutase 1, catalase and glutathione perioxidase 1, were consistently detected in the hip capsules of Group C patients. We propose that dysregulated reactive oxygen species-related enzymes may play an important role in the pathogenesis and progression of aseptic loosening after THR.

Mitochondrial DNA haplogroups and serum levels of proteolytic enzymes in patients with osteoarthritis.

OBJECTIVE: To analyse the influence of mitochondrial DNA haplogroups, as well as the radiographic grade, on serum levels of proteolytic enzymes in patients with osteoarthritis (OA). METHODS: Serum levels of metalloproteinase-1 (MMP-1), MMP-3, MMP-13, myeloperoxidase and cathepsin K were analysed in 73 patients with OA and 77 healthy controls carrying the haplogroups J, U and H, by ELISA. Knee and hip radiographs were classified according to Kellgren and Lawrence (K/L) scoring from grade 0 to grade IV. Non-parametric and multiple regression analyses were performed to test the effects of clinical variables, including gender, age, smoking status, diagnosis, haplogroups and radiological K/L grade on serum levels of these enzymes. RESULTS: A significant influence of the haplogroups on the serum levels of MMP-3 and MMP-13 was detected (p=0.027 and p=0.035, respectively). Patients with OA with haplogroup H showed higher serum levels of MMP-3 than healthy controls. Serum levels of MMP-13 were significantly higher in patients with OA (p<0.001), and carriers of the haplogroup J showed lower levels than H carriers. Besides, levels of MMP-13 were proportionally higher in radiological groups B (K/L grade II and III) and C (K/L grade IV) than in group A (K/L grade 0 and I) (p=0.005). CONCLUSIONS: This study shows that haplogroups have a significant influence on serum levels of MMP-3 and MMP-13. The influence of the haplogroups on serum levels of MMP-3 is clearly dependent on the diagnosis, whereas the influence of the haplogroups on serum levels of MMP-13 is independent of diagnosis.

The (-765 G→C) promoter variant of the COX-2/PTGS2 gene is associated with a lower risk for end-stage hip and knee osteoarthritis.

BACKGROUND: Functional polymorphisms in genes of proinflammatory signalling cascades may contribute to the genetic risk of osteoarthritis (OA). OBJECTIVE: To examine a possible association between end-stage OA of the hip and knee joint and a known single nucleotide polymorphism (SNP) of the COX-2 gene promoter. METHODS: The SNP -765 G→C (rs20417) of the COX-2 gene promoter was genotyped by pyrosequencing in 531 (320 women/211 men) patients with OA from the Ulm Osteoarthritis Study and 400 (200 women/200 men) regional controls from the south-west of Germany. RESULTS: In the whole study population the C allele was associated with a lower risk (per allele OR 0.57; 95% CI 0.43 to 0.75, p<0.0001) and the G allele with a higher risk for end-stage OA. Analysis of subgroups confirmed this result for primary, bilateral, hip and knee OA. CONCLUSION: The promoter polymorphism rs20417 of the COX-2 gene contributes to the genetic risk for end-stage hip and knee OA.

Superoxide dismutase downregulation in osteoarthritis progression and end-stage disease.

BACKGROUND: Oxidative stress is proposed as an important factor in osteoarthritis (OA). OBJECTIVE: To investigate the expression of the three superoxide dismutase (SOD) antioxidant enzymes in OA. METHODS: SOD expression was determined by real-time PCR and immunohistochemistry using human femoral head cartilage. SOD2 expression in Dunkin-Hartley guinea pig knee articular cartilage was determined by immunohistochemistry. The DNA methylation status of the SOD2 promoter was determined using bisulphite sequencing. RNA interference was used to determine the consequence of SOD2 depletion on the levels of reactive oxygen species (ROS) using MitoSOX and collagenases, matrix metalloproteinase 1 (MMP-1) and MMP-13, gene expression. RESULTS: All three SOD were abundantly expressed in human cartilage but were markedly downregulated in end-stage OA cartilage, especially SOD2. In the Dunkin-Hartley guinea pig spontaneous OA model, SOD2 expression was decreased in the medial tibial condyle cartilage before, and after, the development of OA-like lesions. The SOD2 promoter had significant DNA methylation alterations in OA cartilage. Depletion of SOD2 in chondrocytes increased ROS but decreased collagenase expression. CONCLUSION: This is the first comprehensive expression profile of all SOD genes in cartilage and, importantly, using an animal model, it has been shown that a reduction in SOD2 is associated with the earliest stages of OA. A decrease in SOD2 was found to be associated with an increase in ROS but a reduction of collagenase gene expression, demonstrating the complexities of ROS function.

Matriptase is a novel initiator of cartilage matrix degradation in osteoarthritis.

OBJECTIVE: Increasing evidence implicates serine proteinases in pathologic tissue turnover. The aim of this study was to assess the role of the transmembrane serine proteinase matriptase in cartilage destruction in osteoarthritis (OA). METHODS: Serine proteinase gene expression in femoral head cartilage obtained from either patients with hip OA or patients with fracture to the neck of the femur (NOF) was assessed using a low-density array. The effect of matriptase on collagen breakdown was determined in cartilage degradation models, while the effect on matrix metalloproteinase (MMP) expression was analyzed by real-time polymerase chain reaction. ProMMP processing was determined using sodium dodecyl sulfate-polyacrylamide gel electrophoresis/N-terminal sequencing, while its ability to activate proteinase-activated receptor 2 (PAR-2) was determined using a synovial perfusion assay in mice. RESULTS: Matriptase gene expression was significantly elevated in OA cartilage compared with NOF cartilage, and matriptase was immunolocalized to OA chondrocytes. We showed that matriptase activated proMMP-1 and processed proMMP-3 to its fully active form. Exogenous matriptase significantly enhanced cytokine-stimulated cartilage collagenolysis, while matriptase alone caused significant collagenolysis from OA cartilage, which was metalloproteinase-dependent. Matriptase also induced MMP-1, MMP-3, and MMP-13 gene expression. Synovial perfusion data confirmed that matriptase activates PAR-2, and we demonstrated that matriptase-dependent enhancement of collagenolysis from OA cartilage is blocked by PAR-2 inhibition. CONCLUSION: Elevated matriptase expression in OA and the ability of matriptase to activate selective proMMPs as well as induce collagenase expression make this serine proteinase a key initiator and inducer of cartilage destruction in OA. We propose that the indirect effects of matriptase are mediated by PAR-2, and a more detailed understanding of these mechanisms may highlight important new therapeutic targets for OA treatment.

Oxidative equilibrium in the prophylaxis of degenerative joint changes: an analysis of pre- and postoperative activity of antioxidant enzymes in patients with hip and knee osteoarthritis.

BACKGROUND: Osteoarthritis is often accompanied by disturbance of oxidative equilibrium. The aim of the study was to analyze antioxidant defense system function in patients with hip and knee osteoarthritis by assessing superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) activity in erythrocytes. MATERIAL/METHODS: The study included 26 women and 18 men who underwent hip or knee arthroplasty. The pre- and postoperative activities of the antioxidant enzymes were evaluated. SOD activity was determined using the Misra/Fridovich method, CAT by the Beer/Seizer method, and the activity of GPx by the Paglia/Valentine method. The results were compared with those of a healthy control group. RESULTS: SOD activity of the patients was insignificantly lower than in the control group. However, after arthroplasty it increased in subgroups: in males by 13% (p<0.04), in females by 7% (p<0.05), in the group <69 years of age by ca. 14%) (p<0.01), and in the group with idiopathic OA by 12% (p<0.005). The activities of CAT and GPx in all the patient groups were significantly higher than in the controls. It increased even more on the 10th day after arthroplasty with the exception of the patients with rheumatoid osteoarthritis, in whom a decrease by 65.5% (p<0.008) were found, and in the patients > or =69 years old by 9.3% (NS). CONCLUSIONS: Knowledge of antioxidant enzyme activity might offer new targets for future therapeutic methods for the treatment of degenerative joint disease.

[Influence of copper (II) complex on the activity of selected oxidative enzymes]. / Wplyw kompleksu miedzi (LL) na aktywnosc wybranych enzymów antyoksydacyjnych.

UNLABELLED: Coxarthrosis results in physical limitations (hypokinesis) and a necessity to change one's life style to the one that does not require much movement. Physical inactivity leads rapidly to metabolic processes inhibition. The processes of protein, carbohydrate and lipid catabolism intensify. Abnormal metabolism of polyunsaturated fatty acids occurs, which consequently leads to the development of reactive oxygen species (ROS) and to the acceleration of lipid peroxidation. Therefore, it is well-founded to seek for a new group of compounds demonstrating interesting biochemical properties. Among such already known compounds are chromone, azole and flavone derivatives, known for their oxidative properties. The aim of the study was to assess the antioxidant enzymes: catalase (CAT) and glutathione peroxidase (GPX) in erythrocytes in 25 patients suffering from coxarthrosis in comparison with healthy persons. MATERIALS AND METHODS: The activity of catalase was determined in erythrocytes using the method of Beers and Sizer at 240 nm on the Beckman spectrophotometer. The activity of glutathione peroxidase was estimated using the method of Little and O'Brien at 412 nm on the Beckman spectrophotometer. The research was conducted with the permit of the Bioethics Committee No RNN/260/08/KB dated 20.05.2008. Blood for the tests was taken from periulnar vein to anticoagulant tubes (EDTA). The group consisted of 25 patients hospitalized in the Trauma and Orthopaedic Ward of the Hospital in Lódz. RESULTS: The results of the tests on erythrocytes in the patients with physical limitations reveal the increase of glutathione peroxidase and catalase activity by about 50% in all patients, who were administered the dichlorobis (N1-dichloromethyl-3,5-dimethylpyrazole, copper (II) compound in comparison to the control group, where the foregoing compound was not used. CONCLUSION: The research results significantly emphasise that adding of new chemical compounds to blood in investigation of oxidative properties has a considerable influence on the activity of the selected enzymes (catalase and glutathione peroxidase) in comparison to the control group. The research reveals an increase of the activity of all antioxidant enzymes, where newly synthesized chemical compounds were used. The results of antioxidative capacity tests confirmed that complex compounds with Cu (II) ions have a significant influence on the antioxidative status. The studies on the foregoing enzymes also indicate that overproduced free radicals participate in the whole course of the coxarthrosis.

[Influence of chromone compounds on the activity of selected antioxidant enzymes in patients with physical limitations]. / Wplyw zwiazków chromonu na aktywnosc wybranych enzymów antyoksydacyjnych u chorych z ograniczeniami ruchowymi.

UNLABELLED: Coxarthrosis used to be referred to as the "doctors' cross" pointing to the difficulties of the treatment. Degenerative processes in the bones usually concern large joints: knee joints in 25-40% of cases, similarly in case of hip joints. Because of the role the joints play, the arthrosis may result in physical limitations or disability. The aim of the study was to evaluate the activity of catalase and superoxide dismutase (CuZn-SOD) in erythrocytes in patients suffering from the coxarthrosis--as the studied group and in healthy people--as the control group. MATERIALS AND METHODS: Determining the activity of catalase and superoxide dismutase in the group of 17 patients hospitalized because of coxarthrosis. RESULTS: In the study about 90% increase of the activity of the enzymes was determined in the studied group, in which the chemical compound 3{(2-aminothiazolyl)methylene}4H-1-benzopyran-4-one was added to the blood, in comparison with the control group. CONCLUSION: The results of the test on catalase and superoxide dismutase clearly indicate that the free radicals participate in the whole course of coxarthrosis.

[Activity of superoxide dismutase (CuZn-SOD) in erythrocytes of patients after hips alloplasty]. / Aktywnosc dysmutazy ponadtlenkowej (CuZn-SOD) w krwinkach czerwonych chorych po alloplastyce stawu biodrowego.

UNLABELLED: Hip osteoarthritis leads, among others, to abnormally decreased physical activity (hypokinesia). Adverse effect of physical inactivity can cause inhibition of anabolic processes in favour of enhancement of protein, carbohydrate, and lipid catabolitic reactions, as well as inadequate metabolism of polyunsaturated fatty acids. These alterations can induce an increased lipid peroxide synthesis, overproduction of reactive oxygen species (ROS) and acceleration of lipid peroxidation processes. The aim of the study was to determine superoxide dismutase activity (CuZn-SOD) in red blood cells of patients suffering from hip osteoarthritis prior to and following total alloplasty as compared to healthy subjects, and also to evaluate effect of hypokinesia on oxidative stress. MATERIAL AND METHODS; CuZn-SOD activity in red blood cells was determined according to the Misra and Fridovich method in 36 patients with hip osteoarthritis hospitalized at the Traumatic-Orthopaedic Department of the Ministry of Internal Affairs and Administration Hospital in Lódz. RESULTS: In patients with decreased physical activity in ten days after alloplasty, enzyme activity increased (+24.9%), one month since the operation it decreased, but it higher as compared to result activity of CuZn-SOD prior to surgery (+16.8%). CONCLUSIONS: The results activity of superoxide dysmutase leads to ROS generation and their overgeneration in hip osteoarthritis and in first time of treatment.

Alendronate decreases TRACP 5b activity in osteoarthritic bone.

The activity of a tartrat-resistant acid phosphatase 5B (TRACP 5b), a marker of osteoclast function, was quantified in osteoarthritic bone specimens from patients treated with Alendronate. Prior to total hip replacement, 12 patients were randomized in a bisphosphonate and a control group. The bisphosphonate group received daily oral Alendronate for 50 days before operation. After operation, the femoral heads were harvested. Samples of the anterior femoral head (A1) and the intertrochanteric area (A2) were taken, analyzed with an immunoassay and stained for TRACP 5-positive-cells. The immunoassay revealed that TRACP-5b activity of the bisphosphonate group was significantly increased in A1 compared to A2, but not of the control group. Bisphosphonate treatment decreased enzyme activity compared to the controls: 0.41 U/mg vs. 0.31 U/mg in A1 and 0.26 U/mg vs. 0.18 U/mg in A2 (p<0.05). The histological examination shows significantly less TRACP-positive cells in bisphosphonate-treated bone sections, confirming the results. Our data suggest that bisphosphonates reduce TRACP 5b activity in the intertrochanteric area rather than in the anterior femoral head. Consequently, they are more effective in areas of well-supplied bone than in osteoarthritic bone tissue.

Increased level of cytokines and matrix metalloproteinases in osteoarthritic subchondral bone.

OBJECTIVE: The aim of this study was to investigate the expression of several cytokines, matrix metalloproteinases (MMPs), and tissue inhibitor of matrix metalloproteinases (TIMP)-1 in osteoarthritis (OA) and control sera and different joint tissues. METHODS: Serum, synovial fluid, cartilage, synovial and subchondral bone tissues were examined in OA and control subjects. The protein level of tumor necrosis factor (TNF)-alpha, interleukin (IL)-1alpha, IL-8, IL-10 and MMP-2, MMP-3, MMP-9, and TIMP-1 were measured by immunoanalysis. RESULTS: Serum levels of TNF-alpha, MMP-3 and -9 were significantly higher in OA patients than in controls. Conversely, serum IL-10 was decreased in OA patients. CRP was elevated when compared to healthy controls and decreased significantly 6 months after the surgery. In contrast to control samples, OA cartilage and synovium revealed significantly higher MMP-2, -3, -9 and IL-10. IL-1alpha was significantly higher in OA cartilage and IL-8 in OA synovium. Interestingly, MMP-3, -9, TIMP-1 and all tested cytokines were up-regulated in OA subchondral bone. DISCUSSION: This study demonstrates pro-inflammatory condition of OA pathology and supports the idea that vascularized subchondral region may increase the synthesis of cytokines and MMPs leading to degradation of adjacent cartilage.