Fernando Marco

Chitosan scaffolds for osteochondral tissue regeneration

A variety of biomaterials have been introduced as potential substrates for cartilage repair. One such candidate is chitosan, which shares some characteristics with glycosaminoglycan and hyaluronic acid present in articular cartilage. Depending on chitosan source and preparation procedure, variations into its properties can be attained. Thus, the aim of this article is to study and select the most adequate chitosan properties for in vivo osteochondral tissue regeneration. In this work, chitosan molecular weight, deacetylation degree, and calcium content are tested as material variable properties. According to these properties, porous scaffolds were prepared, implanted in rabbit knee osteochondral defects, and evaluated 3 months after surgery. Results show in vitro a considerable influence of the material molecular weight on the scaffold structure. In vivo, different tissue responses were observed depending on the implanted chitosan properties. Some samples showed no material degradation, multiple adverse tissue responses, and no bone/cartilage tissue formation. Other samples showed no adverse responses and bone and cartilage tissue regeneration. The chitosan with intact mineral content (17.9 wt %), lowest molecular weight (11.49 KDa), and lowest deacetylation degree (83%) shows a well structured subchondral bone and noticeable cartilaginous tissue regeneration, being it the best one of those tested for osteochondral defect regeneration.

Early lymphocyte activation in the synovial microenvironment in patients with osteoarthritis: comparison with rheumatoid arthritis patients and healthy controls

Osteoarthritis (OA) is largely considered to be a non-inflammatory disease, although there is compelling evidence that subclinical inflammation is a common event, even in the absence of acute inflammatory flares. In this study we analyze, by means of CD5 and CD69 expression, the infiltration and early activation of CD5+cells, mostly lymphocytes, in both synovial membrane and synovial fluid from advanced OA patients and compare them with samples from patients with rheumatoid arthritis and healthy controls. The number of infiltrating CD5+ cells in both synovial membrane and synovial fluid from patients with advanced OA was significantly reduced as compared with rheumatoid arthritis patients. However, synovial membrane and synovial fluid CD5+ cells on OA exhibited a phenotype with evidence of recent activation comparable to that observed in RA.

Bilateral Smith fracture of the radius caused by airbag deployment.

A passenger-side occupant of a car involved in a collision suffered bilateral Smiths fracture after extending both arms for protection and being hit by the deploying airbag.

Large-scale gene expression in bone marrow mesenchymal stem cells: a putative role for COL10A1 in osteoarthritis

Objectives To elucidate disease-specific molecular changes occurring in osteoarthritis (OA) by analysing the differential gene expression profiles of bone marrow mesenchymal stem cells (BM-MSCs) from patients with OA compared with those without OA. Methods Expression profiles of BM-MSCs from eight paired patients with OA and patients with hip fracture without signs of OA were compared by DNA microarray expression analysis and significant differences were evaluated by computational Gene Set Enrichment Analysis. To validate the involvement of COL10A1 as part of the most downregulated gene set in OA, three tagging single nucleotide polymorphisms were genotyped in 191 patients with OA and 283 controls. COL10A1 expression was also assessed by quantitative RT-PCR in additional subjects. Results Expression levels in 9% (1967) of the overall transcripts were significantly different (p<0.05) between MSCs from patients with OA and controls (532 genes reached twofold differences: 240 were upregulated and 292 were downregulated). Cell development and differentiation were the functional categories accounting for most genes with altered expression. Interestingly, several genes related to the Wnt/-catenin pathway and collagen genes were downregulated in MSCs from patients with OA. The collagen gene set was clearly downregulated in OA. Furthermore, the expression of COL10A1 was significantly reduced in patients with OA. A genetic association between the COL10A1 rs11965969 polymorphism and OA was also found. Conclusion COL10A1 downregulation seems to have a role in the establishment of a defective and/or unstable subchondral cartilage matrix in OA disease. It is proposed that OA may be linked to the intrinsic defective regenerative potential of BM-MSCs resulting from its reduced expression of fate commitment-related genes.

Differential proteome of bone marrow mesenchymal stem cells from osteoarthritis patients

OBJECTIVE Adult mesenchymal stem cells (MSCs) are multipotent cells whose primary reservoir is bone marrow (BM). Following situations of extensive tissue damage, MSCs are mobilized and migrate to the site of injury. Osteoarthritis (OA) is a condition that involves extensive cartilage and bone damage. To gain insight into the pathogenesis of OA, we have analyzed the differential BM-MSCs proteome of OA patients. METHODS MSCs protein extracts were prepared from BM aspirates from six patients with OA and from six hip fracture subjects without OA, and analyzed by Two-dimensional gels, using the differential in-gel electrophoresis approach. Differentially expressed proteins were identified by mass spectrometry. In addition, the chemotactic responses of OA and control MSCs were assessed. RESULTS The majority of proteins that changed at least 1.5-fold (P<0.05) belonged to the following three categories: metabolic enzymes (14 proteins, 36%), cytoskeleton/motility (12 proteins, 32%), and transporters (three proteins, 8%). In OA MSCs, a high percentage of metabolic enzymes (n=8, 57%) were up-regulated and most of the proteins related to cytoskeleton/motility (n=9, 75%) were down-regulated. There was a significant increase in the migration response of OA MSCs to platelet-derived growth factor-BB (chemotaxis index CI: 5.13+/-1.19 vs 3.35+/-0.42, P=0.043). CONCLUSIONS In this study, we have described the differential proteome of BM-MSCs from OA patients together with an increased chemotactic response of these cells in the context of OA. These results could indicate an activation of OA BM-MSCs in response to chemotactic signals sent by the altered subchondral bone in an attempt to heal damaged tissue.

Altered Expression of Wnt Signaling Pathway Components in Osteogenesis of Mesenchymal Stem Cells in Osteoarthritis Patients

Introduction Osteoarthritis (OA) is characterized by altered homeostasis of joint cartilage and bone, whose functional properties rely on chondrocytes and osteoblasts, belonging to mesenchymal stem cells (MSCs). WNT signaling acts as a hub integrating and crosstalking with other signaling pathways leading to the regulation of MSC functions. The aim of this study was to evaluate the existence of a differential signaling between Healthy and OA-MSCs during osteogenesis. Methods MSCs of seven OA patients and six healthy controls were isolated, characterised and expanded. During in vitro osteogenesis, cells were recovered at days 1, 10 and 21. RNA and protein content was obtained. Expression of WNT pathway genes was evaluated using RT-qPCR. Functional studies were also performed to study the MSC osteogenic commitment and functional and post-traslational status of β-catenin and several receptor tyrosine kinases. Results Several genes were downregulated in OA-MSCs during osteogenesis in vitro. These included soluble Wnts, inhibitors, receptors, co-receptors, several kinases and transcription factors. Basal levels of β-catenin were higher in OA-MSCs, but calcium deposition and expression of osteogenic genes was similar between Healthy and OA-MSCs. Interestingly an increased phosphorylation of p44/42 MAPK (ERK1/2) signaling node was present in OA-MSCs. Conclusion Our results point to the existence in OA-MSCs of alterations in expression of Wnt pathway components during in vitro osteogenesis that are partially compensated by post-translational mechanisms modulating the function of other pathways. We also point the relevance of other signaling pathways in OA pathophysiology suggesting their role in the maintenance of joint homeostasis through modulation of MSC osteogenic potential.

Unreamed intramedullary locking nailing for open tibial fractures

Summary. We reviewed the results of the treatment of 24 cases of open tibial fractures using unreamed intramedullary locking nailing. The fractures were classified, following the Gustilo system as grade I-7, grade II-7 and grade III-10. The average time to achieve bony union was 22 weeks with a 26% incidence of pseudoarthrosis. There were no cases of deep infection. Five cases healed with shortening of over 1 cm, but we did not observe angular deformity in any of the patients. In 2 fractures with associated articular lesions, joint motion was limited at final follow up. The nail broke in 2 cases and the screws in 5. The surgical procedure is well tolerated by patients, allows good management of soft tissue lesions and rehabilitation with low rate of infection and malunion. The main disadvantages have been the relative high incidence of nonunion and breakage of metal.Résumé. Les auteurs rapportent les résultats du traitement de vingt-quatre fractures ouvertes de la jambe stabilisées avec un enclouage verrouillé sans alésage. On a classifié les fractures en suivant le systeme de Gustilo: 7 grade I, 7 grade II et 10 grade III. La moyenne de consolidation fut de 22 semaines avec un pourcentage de pseudarthroses de 26%. Il n’y a pas eu d’infection profonde. Cinq cas ont consolidé avec un raccourcissement de plus d’un cm mais on n’a pas observé d’angulation. On a noté une limitation de la mobilité articulaire chez deux patients qui avaient des lésions articulaires. On a eu deux ruptures du clou et cinq de vis. Nous concluons que cette technique chirurgicale est bien tolerée par la patient, elle permet un bon maniement des tissus nous, une réhabilitation précoce et elle a un pourcentage bas d’infection ou mauvaise consolidation. Le principal désavantage a été la relative incidence de pseudarthroses et de ruptures du matériel par stabilité insuffisante.

Midterm Results of Optetrak Posterior-Stabilized Total Knee System After 7 to 12 Years in a University Hospital

A clinical study has been carried out on 434 posterior-stabilized knee prostheses (Optetrak; Exactech, Gainesville, Fla) implanted between 1995 and 2000 in a university general hospital by 23 surgeons. At a mean follow-up of 8.8 years, 297 knees in 249 patients were available for review. Average patient age at surgery was 71.8 years. Average body mass index was 28.8 kg/m(2). Mean flexion range was 108° .The average knee score (Hospital for Special Surgery) increased from 48 points preoperatively to 86 points (60-97 points) at the final review. Of the patients, 81% had an excellent or good result, 14.9% had a fair result, and 4.1% had a poor result. Using the Kaplan-Meier method, we obtained a 91.3% predicted implant survival at 12 years including septic and aseptic revision in the best-case scenario.

The ubiquitin–proteasome pathway and viral infections in articular cartilage of patients with osteoarthritis

Many viruses can evolve different strategies to exploit the ubiquitin–proteasome pathway (UPP) for their own benefit. Some data have recently established connections between UPP and osteoarthritis (OA). The aim of this study was to determine the possible involvement of viral infections linked with the UPP in the physiopathology of OA. Samples of human cartilage were obtained from 12 patients with clinical and radiological features of OA and from 12 normal controls. DNA was extracted from cultured chondrocytes from these patients, and quantitative real-time PCR was performed to analyse the DNA/RNA prevalence and viral loads of HSV, EBV, HCMV, enterovirus, and HTLV-1. The prevalence of total viral DNA/RNA among patients with OA was 16.7% (mean viral load of 7.86 copies/μg DNA), EBV being responsible for the two positive samples, while the prevalence in controls was 0%. We did not detect any positive samples for HSV, CMV, enterovirus, and HTLV-1 among patients with OA and controls. This first approach to the study of the prevalence of viruses linked to the UPP in articular cartilage of end-stage OA patients provides evidences supporting the risk of EBV transmission or reactivation in a subset of patients with disorders requiring tissue regeneration.

175 THE UBIQUITIN-PROTEASOME PATHWAY AND VIRAL INFECTIONS IN ARTICULAR CARTILAGE OF PATIENTS WITH OSTEOARTHRITIS

of c-jun N terminal kinase (JNK), p38 MAP kinase and extracellular-signal regulated kinase (ERK)-1/2. GlcN inhibited JNK and p38 phosphorylation and consequently c-jun and to a lower degree junD DNA binding activity. Moreover, we found also down-regulation of c-jun mRNA transcription level. Conclusions: These results demonstrated for the first time, in human chondrocytes, that GlcN inhibits cytokine-stimulated MMP production, by affecting MAP kinase phosphorylation and consequently AP-1 transcription factor.