We were able to identify a tight subgroup of patients with OA, characterised by an increased inflammatory response that could be regulated by epigenetics. The identification and isolation of this subgroup may be critical for the development of effective treatment and disease prevention.

Objective. Some studies indicate that the mitochondrion is implicated in osteoarthritis (OA). To test the hypothesis that mitochondrial DNA (mtDNA) haplogroups contribute to the prevalence and severity of knee OA, we analyzed the European mtDNA haplogroups in a Spanish population of patients with knee OA and healthy control subjects.Methods. We combined the single-base extension assay with the polymerase chain reaction-restriction fragment length polymorphism technique to obtain the different single-nucleotide polymorphisms that characterize the European haplogroups in 457 patients with knee OA and 262 radiologic controls. Knee OA radio-

ObjectivesThe genetic aetiology of osteoarthritis has not yet been elucidated. To enable a well-powered genome-wide association study (GWAS) for osteoarthritis, the authors have formed the arcOGEN Consortium, a UK-wide collaborative effort aiming to scan genome-wide over 7500 osteoarthritis cases in a two-stage genome-wide association scan. Here the authors report the findings of the stage 1 interim analysis.MethodsThe authors have performed a genome-wide association scan for knee and hip osteoarthritis in 3177 cases and 4894 population-based controls from the UK. Replication of promising signals was carried out in silico in five further scans (44 449 individuals), and de novo in 14 534 independent samples, all of European descent.ResultsNone of the association signals the authors identified reach genome-wide levels of statistical significance, therefore stressing the need for corroboration in sample sets of a larger size. Application of analytical approaches to examine the allelic architecture of disease to the stage 1 genome-wide association scan data suggests that osteoarthritis is a highly polygenic disease with multiple risk variants conferring small effects.ConclusionsIdentifying loci conferring susceptibility to osteoarthritis will require large-scale sample sizes and well-defined phenotypes to minimise heterogeneity.

Objective To evaluate the influence of the mitochondrial DNA (mtDNA) haplogroups in the risk of incident knee osteoarthritis (OA) and to explain the functional consequences of this association to identify potential diagnostic biomarkers and therapeutic targets. Methods Two prospective cohorts contributed participants. The osteoarthritis initiative (OAI) included 2579 subjects of the incidence subcohort, and the cohort hip and cohort knee (CHECK) included 635, both with 8-year follow-up. The analysis included the association of mtDNA haplogroups with the rate of incident knee OA in subjects from both cohorts followed by a subsequent meta-analysis. Transmitochondrial cybrids harbouring haplogroup J or H were constructed to detect differences between them in relation to physiological features including specific mitochondrial metabolic parameters, reactive oxygen species production, oxidative stress and apoptosis. Results Compared with H, the haplogroup J associates with decreased risk of incident knee OA in subjects from OAI (HR=0.680; 95% CI 0.470 to 0.968; p<0.05) and CHECK (HR=0.728; 95% CI 0.469 to 0.998; p<0.05). The subsequent meta-analysis including 3214 cases showed that the haplogroup J associates with a lower risk of incident knee OA (HR=0.702; 95% CI 0.541 to 0.912; p=0.008). J cybrids show a lower free radical production, higher cell survival under oxidative stress conditions, lower grade of apoptosis as well as lower expression of the mitochondrially related pro-apoptotic gene BCL2 binding component 3 (BBC3). In addition, J cybrids also show a lower mitochondrial respiration and glycolysis leading to decreased ATP production. Conclusions The physiological effects of the haplogroup J are beneficial to have a lower rate of incident knee OA over time. Potential drugs to treat OA could focus on emulating the mitochondrial behaviour of this haplogroup.

Mitochondria and mitochondrial DNA (mtDNA) variation are now recognized as important factors in the development of osteoarthritis (OA). Mitochondria are the energy powerhouses of the cell, and also regulate different processes involved in the pathogenesis of OA including inflammation, apoptosis, calcium metabolism and the generation of reactive oxygen species (ROS) and reactive nitrogen species (RNS). Mitochondria contain their own genetic material, mtDNA, which evolved through the sequential accumulation of mtDNA variants to enable humans to adapt to different climates. The ROS and reactive metabolic intermediates that are by-products of mitochondrial metabolism are regulated in part by mtDNA and are among the signals that transmit information between mitochondria and the nucleus. These signals can alter nuclear gene expression and, when disrupted, affect a number of cellular processes and metabolic pathways, leading to disease. mtDNA variation influences OA-associated phenotypes, including those related to metabolism, inflammation and even ageing, as well as nuclear epigenetic regulation. This influence also enables the use of specific mtDNA haplogroups as complementary diagnostic and prognostic biomarkers of OA.

BackgroundMitochondrion has an important role in the osteoarthritis (OA) pathology. We have previously demonstrated that the alteration of the mitochondrial respiratory chain (MRC) contributes to the inflammatory response of the chondrocyte. However its implication in the process of cartilage destruction is not well understood yet. In this study we have investigated the relationship between the MRC dysfunction and the regulation of metalloproteases (MMPs) in human normal chondrocytes in culture.MethodsHuman normal chondrocytes were isolated from human knees obtained form autopsies of donors without previous history of rheumatic disease. Rotenone, 3-Nitropropionic acid (NPA), Antimycin A (AA), Sodium azide and Oligomycin were used to inhibit the activity of the mitochondrial complexes I, II, III, IV and V respectively. The mRNA expression of MMPs -1, -3 and -13 was studied by real time PCR. The intracellular presence of MMP proteins was evaluated by western blot. The liberation of these proteins to the extracellular media was evaluated by ELISA. The presence of proteoglycans in tissue was performed with tolouidin blue and safranin/fast green. Immunohistochemistry was used for evaluating MMPs on tissue.ResultsFirstly, cells were treated with the inhibitors of the MRC for 24 hours and mRNA expression was evaluated. An up regulation of MMP-1 and -3 mRNA levels was observed after the treatment with Oligomycin 5 and 100 μg/ml (inhibitor of the complex V) for 24 hours. MMP-13 mRNA expression was reduced after the incubation with AA 20 and 60 μg/ml (inhibitor of complex III) and Oligomycin. Results were validated at protein level observing an increase in the intracellular levels of MMP-1 and -3 after Oligomycin 25 μg/ml stimulation [(15.20±8.46 and 4.59±1.83 vs. basal=1, respectively (n=4; *P<0.05)]. However, AA and Oligomycin reduced the intracellular levels of the MMP-13 protein (0.70±0.16 and 0.3±0.24, respectively vs. basal=1). In order to know whether the MRC dysfunction had an effect on the liberation of MMPs, their levels were evaluated in the supernatants. After 36 hours of stimulation, values were: MMP-1=18.06±10.35 with Oligomycin 25 μg/ml vs. basal=1, and MMP-3=8.49±4.32 with Oligomycin 5 μg/ml vs. basal=1 (n=5; *P<0.05). MMP-13 levels in the supernatants were reduced after AA 60 μg/ml treatment (0.50±0.13 vs. basal=1) and Oligomycin 25 μg/ml (0.41±0.14 vs. basal=1); (n=5; *P<0.05). The treatment of explants with Oligomycin, showed an increase in the positivity of MMP-1 and -3. Explants stimulated with AA or Oligomycin revealed a decrease in MMP-13 expression. Proteoglycan staining demonstrated a reduction of proteoglycan levels in the tissues treated with Oligomycin.ConclusionsThese results reveal that MRC dysfunction modulates the MMPs expression in human normal chondrocytes demonstrating its role in the regulation of the cartilage destruction.

BackgroundOxidative stress due to the overproduction of nitric oxide (NO) and other oxygen reactive species (ROS), play a main role in the initiation and progression of the OA disease and leads to the degeneration of mitochondria. Therefore, the goal of this work is to describe the difference in telomere length of peripheral blood leukocytes (PBLs) and Nitric Oxide (NO) production between mitochondrial DNA (mtDNA) haplogroup J and non-J carriers, as indirect approaches of oxidative stress.MethodsThe telomere length of PBL was analyzed in DNA samples from 166 healthy controls (114 J and 52 non-J) and 79 OA patients (41 J and 38 non-J) by means of a validated qPCR method. The NO production was assessed in 7 carriers of the haplogroup J and 27 non-J carriers, by means of the colorimetric reaction of the Griess reagent in supernatants of cultured chondrocytes. Inducible nitric oxide synthase (iNOS) mRNA from these samples was analyzed by qPCR. Appropiated statistical analyses were performedResultsCarriers of the haplogroup J showed a significantly longer telomere length of PBLs than non-J carriers, regardless of age, gender and diagnosis (p = 0.025). Cultured chondrocytes carrying the mtDNA haplogroup J also showed a lower NO production than non-J carriers (p = 0.043). No significant correlations between age and telomore length of PBLs were detected neither for carriers of the haplogroup J nor for non-J carriers. A strong positive correlation between NO production and iNOS expression was also observed (correlation coefficient = 0.791, p < 0.001).ConclusionThe protective effect of the mtDNA haplogroup J in the OA disease arise from a lower oxidative stress in carriers of this haplogroup, since this haplogroup is related to lower NO production and hence longer telomere length of PBLs too.

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.