Dexamethasone (Dex)-induced osteoporosis has been described as the most severe side effect in long-term glucocorticoid therapy. The decreased bone mass and the increased marrow fat suggest that Dex possibly shifts the differentiation of bone marrow stromal cells (BMSCs) to favor adipocyte over osteoblast, but the underlying mechanisms are still unknown. In this paper, we established a Dex-induced osteoporotic mouse model, and found that BMSCs from Dex-treated mice are more likely to differentiate into adipocyte than those from control mice, even under the induction of bone morphogenetic protein-2 (BMP2). We also discovered both in vitro and in vivo that the expression level of adipocyte regulator CCAAT/enhancer-binding protein alpha (C/EBPalpha) is significantly upregulated in Dex-induced osteoporotic BMSCs during osteoblastogenesis by a mechanism that involves inhibited DNA hypermethylation of its promoter. Knockdown of C/EBPalpha in Dex-induced osteoporotic cells rescues their differentiation potential, suggesting that Dex shifts BMSC differentiation by inhibiting C/EBPalpha promoter methylation and upregulating its expression level. We further found that the Wnt/beta-catenin pathway is involved in Dex-induced osteoporosis and C/EBPalpha promoter methylation, and its activation by LiCl rescues the effect of Dex on C/EBPalpha promoter methylation and osteoblast/adipocyte balance. This study revealed the C/EBPalpha promoter methylation mechanism and evaluated the function of Wnt/beta-catenin pathway in Dex-induced osteoporosis, providing a useful therapeutic target for this type of osteoporosis.

In Canada, new models of orthopaedic care involving advanced practice physiotherapists (APP) are being implemented. In these new models, aimed at improving the efficiency of care for patients with musculoskeletal disorders, APPs diagnose, triage and conservatively treat patients. Formal validation of the efficiency and appropriateness of these emerging models is scarce. The purpose of this study is to assess the diagnostic agreement of an APP compared to orthopaedic surgeons as well as to assess treatment concordance, healthcare resource use, and patient satisfaction in this new model. 120 patients presenting for an initial consult for hip or knee complaints in an outpatient orthopaedic hospital clinic in Montreal, Canada, were independently assessed by an APP and by one of three participating orthopaedic surgeons. Each health care provider independently diagnosed the patients and provided triage recommendations (conservative or surgical management). Proportion of raw agreement and Cohen’s kappa were used to assess inter-rater agreement for diagnosis, triage, treatment recommendations and imaging tests ordered. Chi-Square tests were done in order to compare the type of conservative treatment recommendations made by the APP and the surgeons and Student t-tests to compare patient satisfaction between the two types of care. The majority of patients assessed were female (54%), mean age was 54.1 years and 91% consulted for a knee complaint. The raw agreement proportion for diagnosis was 88% and diagnostic inter-rater agreement was very high (κ=0.86; 95% CI: 0.80-0.93). The triage recommendations (conservative or surgical management) raw agreement proportion was found to be 88% and inter-rater agreement for triage recommendation was high (κ=0.77; 95% CI: 0.65-0.88). No differences were found between providers with respect to imaging tests ordered (p≥0.05). In terms of conservative treatment recommendations made, the APP gave significantly more education and prescribed more NSAIDs, joint injections, exercises and supervised physiotherapy (p<0.05). Patient satisfaction was significantly higher for APP care than for the surgeons care (p<0.05). The diagnoses and triage recommendations for patients with hip and knee disorders made by the APP were similar to the orthopaedic surgeons. These results provide evidence supporting the APP model for orthopaedic care.

Introduction: 4-Hydroxynonenal (HNE) is one of the most abundant and reactive aldehydes of lipid peroxidation products and exerts various effects on intracellular and extracellular signalling cascades. We have previously shown that HNE at low concentrations could be considered as an important mediator of catabolic and inflammatory processes in osteoarthritis (OA). In the present study, we focused on characterizing the signalling cascade induced by high HNE concentration involved in cell death in human OA chondrocytes. Methods: Markers of apoptosis were quantified with commercial kits. Protein levels were evaluated by Western blotting. Glutathione (GSH) and ATP levels were measured with commercial kits. Glucose uptake was assessed by 2-deoxy-D-[³H]-glucose. The role of GSH-S-transferase A4-4 (GSTA4-4) in controlling HNE-induced chondrocyte apoptosis was investigated by chondrocyte transfection with small interfering RNA (siRNA) or with the expression vector of GSTA4-4. Results: Our data showed that HNE at concentrations of up to 10 μM did not alter cell viability but was cytotoxic at concentrations of greater than or equal to 20 μM. HNE-induced chondrocyte death exhibited several classical hallmarks of apoptosis, including caspase activation, cytochrome c and apoptosis-induced factor release from mitochondria, poly (ADP-ribose) polymerase cleavage, Bcl-2 downregulation, Bax upregulation, and DNA fragmentation. Our study of signalling pathways revealed that HNE suppressed pro-survival Akt kinase activity but, in contrast, induced Fas/CD95 and p53 expression in chondrocytes. All of these effects were inhibited by an antioxidant, N-acetyl-cysteine. Analysis of cellular energy and redox status showed that HNE induced ATP, NADPH, and GSH depletion and inhibited glucose uptake and citric acid cycle activity. GSTA4-4 ablation by the siRNA method augmented HNE cytotoxicity, but, conversely, its overexpression efficiently protected chondrocytes from HNE-induced cell death. Conclusion: Our study provides novel insights into the potential mechanisms of cell death in OA cartilage and suggests the potential role of HNE in OA pathophysiology. GSTA4-4 expression is critically important for cellular defence against oxidative stress-induced cell death in OA cartilage, possibly by HNE elimination.

At present, gene transfection insufficient efficiency is a major drawback of non-viral gene therapy. The 2 main types of delivery systems deployed in gene therapy are based on viral or non-viral gene carriers. Several non-viral modalities can transfer foreign genetic material into the human body. To do so, polycation-based gene delivery methods must achieve sufficient efficiency in the transportation of therapeutic genes across various extracellular and intracellular barriers. These barriers include interactions with blood components, vascular endothelial cells and uptake by the reticuloendothelial system. Furthermore, the degradation of therapeutic DNA by serum nucleases is a potential obstacle for functional delivery to target cells. Cationic polymers constitute one of the most promising approaches to the use of viral vectors for gene therapy. A better understanding of the mechanisms by which DNA can escape from endosomes and traffic to enter the nucleus has triggered new strategies of synthesis and has revitalized research into new polycation-based systems. The objective of this review is to address the state of the art in gene therapy with synthetic and natural polycations and the latest advances to improve gene transfer efficiency in cells