Cartilage diseases have a significant impact on the patient's quality of life and are a heavy burden for the healthcare system. Better understanding, early detection and proper follow-up could improve quality of life and reduce healthcare related costs. Therefore, the aim of this study was to evaluate if difference between osteoarthritic (OA) and non-osteoarthritic (non-OA) knees can be detected quantitatively on cartilage and subchondral bone levels with advanced but clinical available imaging techniques. Two OA (mean age = 88.3 years) and three non-OA (mean age = 51.0 years) human cadaveric knees were scanned two times. A high-resolution peripheral quantitative computed tomography (HR-pQCT) scan (XtremeCT, Scanco Medical AG, Switzerland) was performed to quantify the bone microstructure. A contrast-enhanced clinical CT scan (GE Revolution Evo, GE Medical Systems AG, Switzerland) was acquired with the contrast agent Visipaque 320 (60 ml) to measure cartilage. Subregions dividing the condyle in four parts were identified semi-automatically and the images were segmented using adaptive thresholding. Microstructural parameters of subchondral bone and cartilage thickness were quantified. The overall cartilage thickness was reduced by 0.27 mm between the OA and non-OA knees and the subchondral bone quality decreased accordingly (reduction of 33.52 % in BV/TV in the layer from 3 to 8 mm below the cartilage) for the femoral medial condyle. The largest differences were observed at the medial part of the femoral medial condyle both for cartilage and for bone parameters, corresponding to clinical observations. Subchondral bone microstructural parameters and cartilage thickness were quantified using in vivo available imaging and apparent differences between the OA and non-OA knees were detected. Those results may improve OA follow-up and diagnosis and could lead to a better understanding of OA. However, further in vivo studies are needed to validate these methods in clinical practice

Osteoarthritis (OA) is a joint degenerative disease leading to chronic pain and disability, thus resulting in a major socioeconomic health burden. OA, which has long been believed to be a cartilage disease, is now considered a whole-joint disorder affecting various anatomical structures, including subchondral bone. Hyaluronic Acid (HA) is commonly used as intra-articular viscosupplementation therapy for its mechanical features and biological effects. Bisphosphonates (BPs) are antiresorptive agents inhibiting recruitment and maturation of osteoclast precursors and activity of mature osteoclasts in the bone. Pre-clinical evidences in the literature, show that intra-articular BPs could impact on OA progression, slowing down or reversing it. The combination of HA biological and mechanical role and Alendronate (ALD) antiresorptive effect could be an interesting strategy for OA treatment. This study describes the synthesis and characterization of FID-134, a new chemical derivative of HA conjugated with ALD by means of a covalent bond, cleavable in physiological condition. FID-134 was synthesized starting from 500 kDa HA: chemical structure and functionalization degree with ALD were investigated by NMR and ICP-OES. Kinetics of ALD release from FID-134 was determined in TRIS buffer at 37°C and compared to a simple mixture of HA+ALD. 20mg/mL formulations of FID-134 and HA+ALD were investigated for viscoelastic properties, in absence and presence of Ca. 2+. ions. The cytotoxicity of FID-134 and free ALD were tested on Saos-2 osteoblasts (ATCC HTB-85) and on primary bovine chondrocytes (PBC) at day 1, 3 and 7. The efficacy of FID-134 was assessed in an inflammatory arthritis in vitro model, where bovine cartilage biopsies were exposed to IL-1β/OSM (10ng/mL) for 3 weeks; at the same time, cartilage explants were treated with FID-134. Collagen release in the surnatants was quantified and compared to controls. FID-134 structure was confirmed by NMR and the 20% mol/mol functionalization degree was determined by ICP-OES. Only about 50% of total bound ALD was released from FID-134 within 7 days, resulting slower compared to HA+ALD mixture. In presence of Ca. 2+. ions, viscoelastic properties of FID-134 dramatically improved, while HA+ALD formulation remained unaffected. The cytotoxicity of ALD was evident at 100 μM on Saos-2 and PBC after 3 days, while no cytotoxicity was observed at 7 days with FID-134. In the cartilage explant model, a strong collagen release was detected in inflammatory conditions after 3 weeks; this tendency was reversed, and collagen release halved when FID-134 was added to the biopsies. The synthesized HA-ALD adduct, FID-134, opens the door for a new approach for OA treatment. The results suggest that FID-134 could be beneficial in cartilage degradation and in restoration of subchondral bone function. Finally, local administration and controlled BP release would likely overcome the drawbacks of ALD oral administration, such as unspecific features and long-term toxic side effects

The cartilage diseases such as osteoarthritis and chondral injuries are considered irreversible and the result of recent treatments remains not optimal. One of the reasons is due to the poor understanding of chondrocyte behaviours. To understand more about cartilage, we designed a series of novel experiments. First, a total joint of bovine metatarsophalanges was isolated as our novel model. We chose it because the configuration and the healing potential were similar to human, and many variables of large animal studies could be controlled in laboratory. The model not only provided a good ex vivo platform for cartilage researches but also connected in vitro cellular studies and in vivo animal studies. To mimic joint movement a special driving machine was designed. To characterise the novel model viabilities of chondrocytes and contents of sulphated glycosaminoglycan (GAGs) in extracellular matrixes were measured every seven days. The preliminary results revealed the viabilities of chondrocytes remained above 80% alive in the middle zone after four-weeks culture. The GAGs contents decreased after this culturing period. The experiments still carry on going to compare the static and dynamic models which joint movement could be a determinative factor to the viability of chondrocytes. Cellular treatment is the recent mainstream for cartilage diseases. If advanced knowledge in chondrocyte behaviours could be obtained from this model, development of optimal treatment will be possible in the future

Mesenchymal stem cells (MSCs) are self-renewing, multipotent cells that could potentially be used to repair injured cartilage in diseases. The objetive was to analyze different sources of human MSCs to find a suitable alternative source for the isolation of MSCs with high chondrogenic potential. Femoral bone marrow, adipose tissue from articular and subcutaneous locations (hip, knee, hand, ankle and elbow) were obtained from 35 patients who undewent different types of orthopedic surgery (21 women, mean age 69.83 ± 13.93 (range 38–91) years. Neoplasic and immunocompromised patients were refused. The Ethical Committee for Clinical Research of the Government of Aragón (CEICA) approved the study and all patients provided informed consent. Cells were conjugated wiith monoclonal antibodies. Cell fluorescence was evaluated by flow cytometry using a FACSCalibur flow cytometer and analysed using CellQuest software (Becton Dickinson). Chondrogenic differentiation of human MSCs from the various tissues at P1 and P3 was induced in a 30-day micropellet culture [Pittenger et al., 1999]. To evaluate the differentiation of cartilaginous pellet cultures, samples were fixed embedded in paraffin and cut into 5- υm-thick slices. The slices were treated with hematoxylin-eosin and safranin O (Sigma-Aldrich). Each sample was graded according to the Bern Histological Grading Scale [Grogan et al., 2006], which is a visual scale that incorporates three parameters indicative of cartilage quality: uniform and dark staining with safranin O, cell density or extent of matrix produced and cellular morphology (overall score 0–9). Stained sections were evaluated and graded by two different researchers under a BX41 dual viewer microscope or a Nikon TE2000-E inverted microscope with the NIS-Elements software. Statistics were calculated using bivariate analysis. Pearson's χ2 or Fisher's exact tests were used to compare the Bern Scores of various tissues. To evaluate the cell proliferation, surface marker expression and tissue type results, ANOVA or Kruskal-Wallis tests were used, depending on the data distribution. Results were considered to be significant when p was < 0.05. MSCs from all tissues analysed had a fibroblastic morphology, but their rates of proliferation varied. Subcutaneous fat derived MSCs proliferated faster than bone marrow. MSCs from Hoffa fat, hip and knee subcutaneous proliferated slower than MSCs from elbow, ankle and hand subcutaneous. Flow cytometry: most of cells lacked expression of CD31, CD34, CD36, CD117 (c-kit), CD133/1 and HLA-DR. At same time 95% of cells expressed CD13, CD44, CD59, CD73, CD90, CD105, CD151 y CD166. Fenotype showed no differences in cells from different anatomic places. Cells from hip and knee subcutaneous showed a worst differentiation to hyaline cartilage. Hoffa fat cells showed high capacity in transforming to hyaline cartilage. Cells from different anatomic places show different chondrogenic potential that has to be considered to choose the cells source

Aim

Osteoarthritis (OA) is caused by complex interactions between genetic and environmental factors. Epigenetic mechanisms control the expression of genes and are likely to regulate the OA transcriptome. We performed integrative genomic analyses to define methylation-gene expression relationships in osteoarthritic cartilage.