Aims

Osteoarthritis (OA) is the most common chronic pathema of human joints. The pathogenesis is complex, involving physiological and mechanical factors. In previous studies, we found that ferroptosis is intimately related to OA, while the role of Sat1 in chondrocyte ferroptosis and OA, as well as the underlying mechanism, remains unclear.

Aims

This study aimed to evaluate the effectiveness of the induced membrane technique for treating infected bone defects, and to explore the factors that might affect patient outcomes.

Objectives

Osteoporosis has become an increasing concern for older people as it may potentially lead to osteoporotic fractures. This study is designed to assess the efficacy and safety of ten therapies for post-menopausal women using network meta-analysis.

We investigated the safety and efficacy of treating osteoporotic vertebral compression fractures with an intravertebral cleft by balloon kyphoplasty. Our study included 27 patients who were treated in this way. The mean follow-up was 38.2 months (24 to 54). The anterior and middle heights of the vertebral body and the kyphotic angle were measured on standing lateral radiographs before surgery, one day after surgery, and at final follow-up. Leakage of cement was determined by CT scans. A visual analogue scale and the Oswestry disability index were chosen to evaluate pain and functional activity. Statistically significant improvements were found between the pre- and post-operative assessments (p < 0.05) but not between the post-operative and final follow-up assessments (p > 0.05). Asymptomatic leakage of cement into the paravertebral vein occurred in one patient, as did leakage into the intervertebral disc in another patient.

We suggest that balloon kyphoplasty is a safe and effective minimally invasive procedure for the treatment of osteoporotic vertebral compression fractures with an intravertebral cleft.

In a rabbit model we investigated the efficacy of a silk fibroin/hydroxyapatite (SF/HA) composite on the repair of a segmental bone defect. Four types of porous SF/HA composites (SF/HA-1, SF/HA-2, SF/HA-3, SF/HA-4) with different material ratios, pore sizes, porosity and additives were implanted subcutaneously into Sprague-Dawley rats to observe biodegradation. SF/HA-3, which had characteristics more suitable for a bone substitite based on strength and resorption was selected as a scaffold and co-cultured with rabbit bone-marrow stromal cells (BMSCs). A segmental bone defect was created in the rabbit radius. The animals were randomised into group 1 (SF/HA-3 combined with BMSCs implanted into the bone defect), group 2 (SF/HA implanted alone) and group 3 (nothing implanted). They were killed at four, eight and 12 weeks for visual, radiological and histological study.

The bone defects had complete union for group 1 and partial union in group 2, 12 weeks after operation. There was no formation of new bone in group 3. We conclude that SF/HA-3 combined with BMSCs supports bone healing and offers potential as a bone-graft substitute.

Introduction: Ethanol is one of risk factors associated with osteonecrosis, it has been demonstrated that ethanol induces adipogenesis, decreases osteogenesis in bone marrow stroma cells and produces intracellular lipid deposits, resulting in the death of osteocytes.

Materials and Methods: In this approach, we isolated human bone marrow stroma cells and triggered for different differentiations.

Results: These cells could be induced for osteogenesis, adipogenesis, and chondrogenesis. We also evaluated cell surface markers of isolated human bone marrow stromal cells that were found to express CD29, CD49d, CD62 CD90, CD105/SH2, SH3, CD133, and CD166, but not CD31, CD34, CD45, or CD56.

Discussion: We demonstrated that ethanol decreases the expression of osteogenic genes, but increases adipogenic genes expressions. Moreover, we found that ethanol decreases the beta-catenin-dependent canonical Wnt signaling pathway related gene expressions, including Wnt 3a and LRP5 genes. Interestingly, ethanol also diminishes the intra-nuclear translocation of β-catenin in human bone marrow stromal cells. Therefore, these results indicate that ethanol might decrease osteogenic gene expressions through Wnt signaling pathway.

Introduction: The main purpose of this study was to analyze the accuracy of conventional versus navigated open wedge corrective osteotomies of the proximal tibia. Furthermore, the intraoperative radiation dosage and the time of the operative procedure of both groups were compared.

Methods: 20 legs of 11 fresh cadaver (9 male, 2 female, age 35–71 years) were randomly assigned to conventional open wedge high tibial osteotomy (HTO) (n=10) or navigated open wedge HTO (n=10). Two legs had to be excluded because of pre-existing knee injuries. The aim of all corrective operations was to align the mechanical axis to pass through 80% of the tibial plateau (80% Fujisawa line), regardless of the preexisting alignment. The intraoperative mechanical axis was evaluated either by the cable technique for conventional HTO, or by a navigation module for navigated HTO (Medivision, Oberdorf/Switzerland). An angle fixed implant with interlocking screws (Tomofix, Mathys, Bettlach/Switzerland) was used to minimize postoperative loss of correction. Postoperatively, CT-scans were performed and the Fujisawaline and MPTA measured with a computer software for deformity analysis (Med-iCAD) The main outcome parameter was the accuracy of the correction, which was measured by the Fujisawa line. Secondary outcome parameters were the intraoperative radiation measured by the dose area product and the time of the operative procedure. For statistical analysis the standard deviation (S.D.) was calculated and the paired t-test applied.

Results: After conventional HTO, the mechanical axis was intersecting the Fujisawa line at 72.1% of the tibial plateau (range 60.4–82.4%, S.D. 7.2%). In contrast, after navigated HTO the tibia plateau was passed through 79.7% (range 75.5–85.8%, S.D. 3.3%). Thus, the accuracy of the correction was significantly higher after navigated HTO (p=0.020). In addition, the standard deviation of the corrections was significantly lower after navigated HTO (p=0.012). The medial proximal tibia angle (MPTA) increased 7.9° (range: 4.7–12.1°) after conventional HTO and 9.1° (range: 4.6–12.6°) after navigated HTO. The average dose area products of the conventional HTO (49.5 cGy/cm2, range 36.0–81.2 cGy/cm2) and navigated HTO (42.8 cGy/cm2, range 28.3–58.1 cGy/cm2) were comparable (p=0.231). However, navigated HTO elongated the operation time significantly (navigated HTO: 82 min, range 55–98 min; conventional HTO: 59 min, range 47–73 min) (p< 0.001).

Conclusion: Continuous three-dimensional imaging of the axis and of intraoperative tools with the a navigation module significantly improves the accuracy of open wedge osteotomies of the proximal tibia. Prospective clinical studies will show whether the results of this cadaver study can be transferred to the regular clinical use.

Introduction: Alcohol can induce osteoporosis and osteonecrosis. Studies have demonstrated that alcohol contributed to abnormal lipid metabolism in cells in bone marrow but the mechanisms have not been defined. The purpose of this study was to evaluate the effect of alcohol on the differentiation of pluripotential cells cloned from bone marrow.

Materials and Methods: The cells were maintained in culture and treated with either increasing concentrations of ethanol (0.09, 0.15, and 0.21 mol/L) or without alcohol to serve as controls. Morphologic features of the cells were monitored using a phase-contrast microscope. Alkaline phosphatase activity was determined using a colorimetric assay. Gene expression of adipogenesis [422 (aP2), PPAR y] and osteogenesis (osteocalcin) was evaluated using the Northern blot technique and reverse transcription-polymerase chain reaction (RT-PCR). ANOVA was used for statistical analysis.

Results: The cells treated with ethanol started to accumulate triglyceride vesicles at Day 7; the number of adipocytes and the percentage of the area that contained the cells with fat vesicles increased significantly; and the level of alkaline phosphatase activity diminished with longer durations of exposure and with higher concentrations of ethanol. Analysis of gene expression showed diminished expression of osteocalcin without a significant increase in the expression of the fat cell specific gene, 422 (aP2), and PPAR y, in cells treated with ethanol. This suggested that adipogenesis may occur at a point downstream in the fatty acid metabolism pathway.

Discussion: Alcohol induces bone marrow fatty changes in patients and in animal models contributing to osteoporosis and osteonecrosis. This study demonstrated that alcohol treatment decreased osteogenesis while enhancing adipogenesis by bone marrow stromal cells, which may be one of the mechanisms leading to osteoporosis and osteonecrosis. Inhibition of adipogenesis may lead to the prevention of the disease.

Clinical relevance: This is a novel finding that alcohol induces adipogenesis in a cloned bone marrow stromal cell. The results explain the clinical observation that there is increased adipogenesis in alcohol-induced osteoporosis and osteonecrosis.

Introduction: Treatment of osteonecrosis continues to be a challenging problem in orthopedic practice. Arthroplasty is generally successful but long-term results are inferior especially in young adults. Alternative treatments such as core decompression and trap-door procedures provide only temporary benefits and need much improvement. The replacement of necrotic bone to promote osteogenesis and angiogenesis and healing subchondral bone are future approaches. Autogenous cancellous bone is the preferred graft material but its supply is limited. Allografts are useful but not as desirable as autografts. Substitutes for bone grafts have been actively researched but few are available currently. In this study, we have attempted to use genetically engineered bone marrow stem cells in order to enhance the healing of a bone defect in a mouse model.

Materials and Methods: A bone marrow stem cell was cloned from Balb/c mice and transfected with LacZ and neomycin resistance genes. The cells were cultured for 7 to 10 days and both the osteoblastic and angiogenic properties of the cells were examined using Northern blots to detect osteocalcin and VEGF gene expression. The cells were also analyzed for alkaline phosphatase activity to demonstrate the osteoblastic phenotype of the cells. A suspension containing 2 x 10⁷ cells/ml phosphate buffered solution was prepared for cell transplantation. A total of forty-eight, 8-week old Balb/c mice were used in this study. A 1.2 mm defect was created bilaterally with an electric drill in the femurs of 24 mice to mimic the core decompression and trap-door procedures. 2 x 10⁶ cells were transplanted into each defect of the right femur while the left femur served as a control trap-door defect which was injected with PBS but without cells. An equal number of cells were injected either at subcutaneous sites, in the hindquarter muscles, or into the renal capsule (8 mice in each site) to evaluate ossification at ectopic sites. Animals were sacrificed at 2, 4, 6 and 8 weeks. Defect repair was evaluated radiographically and the contribution to osteogenesis by transplanted cells was studied histomorphometrically using tissue sections stained with X-gal as well as biochemically on DNA extracts using primers for the neomycin resistance gene.

Results: Radiopaque tissue appeared two weeks after the cells were transplanted into bone defects, muscle, subcutaneous sites, and the renal capsule. Histological analysis demonstrated that these tissues consist of newly formed bone from transplanted cells that stained positively with X-gal and contained neo DNA. The repair tissue did not contain cartilaginous areas indicating that ossification surrounding the D1-BAG cells was not through the endochondral process. At four weeks, 4 of 6 femora showed a defect that was filled with new bone. At 6 weeks, all of the defects (6 of 6) contained fully restored bone. However, in the control side that was injected with PBS (no cells) only 2 of 6 at 4 weeks, 3 of 6 at 6 weeks, and 5 of 6 at 8 weeks showed complete repair. All histological sections of bone defects (n = 24) were examined histomorphometrically using a computerized image analysis system. Transplantation of marrow stem cells into bone defects produced more bone at an earlier time point than controls and, the process of enhanced ossification continued throughout the healing process.

Discussion: The cloned bone marrow stem cell can directly form bone after transplantation into bone defects and into ectopic sites, indicating that the in vitro expanded bone marrow stem cells can serve as a grafting material to enhance healing of bone defects and the treatment of osteonecrosis. In addition, this study demonstrates that genetic labelling is a useful tool in studies of cell differentiation in vivo and that bone marrow stem cells may be useful as a carrier of genetically-engineered factors in the treatment of skeletal diseases.

Introduction: Studies have shown steroidal and non-steroidal anti-inflammatory drugs (NSAIDs) suppress bone remodeling. Previous results have indicated that NSAIDs suppress proliferation and induce cell death in cultured osteoblasts and pluripotent stem cells (D1-cells), suggesting these effects might be one of the mechanisms contributing to their inhibitory effects on bone remodeling in vivo. On the other hand, our previous results indicated that dexamethasone treatment shifts the characteristics of osteogenesis into adipogenesis in D1-cells. However, the influences of NSAID on adipogenesis in pluripotent stem cells have rarely been investigated. In this study, we tested the adipogenesis of D1-cells upon long-term treatment of NSAIDs. NSAID influence on the osteocalcin expressions of D1-cells was also examined.

Materials and Methods: The effects of treatments with indomethacin, ketorolac, diclofenac and piroxicam (10⁻⁵ and 10⁻⁴ M) for 2, 4 6 or 8 days were evaluated. Lipid droplets in cultures were detected by oil red staining. Adipsin and osteocalcin mRNA expressions were examined by RT-PCR.

Results: In this study, 10⁻⁴M of NSAID treatment for 4–8 days induced adipogenesis in D1-cells, while shorter duration and lower concentration did not. Mild adipogenesis also occurred in cultures treated with 10⁻⁵M of indomethacin for 6 or 8 days, revealing the strongest effect among the 4 NSAIDs. Piroxicam revealed less effects on adipogenesis in D1-cells. However, despite 2-days of treatment with 10⁻⁵M indomethacin, NSAIDs did not affect the expression of osteocalcin either at 10⁻⁵–10⁻⁴M or during 2–8 days of treatments.

Conclusion: These results suggest that high dose and long term administration of NSAIDs may induce adipogenesis in pluripotent stem cells.

Introduction: Our previous study found that glucocorticoids shifted the properties of osteogenesis to adipogenesis in murine marrow stem cells. These effects may be one of the important mechanisms in the pathogenesis of osteonecrosis. Statins prevented these steroid effects. In this study, we investigated the effects of dexamethasone and lovastatin on the expressions of bone morphogenetic protein-2 (BMP2) in the bone marrow stroma cells cultured from osteonecrotic patients.

Materials and Methods: Bone marrow fluid aspiration from iliac crest was performed in osteonecrosis (ON) and non-ON patients after surgical treatment for their hip disorder. The mean age of the patients was 59 years in the ON group and 63 years in the non-ON group. Nucleated stroma cells were isolated from bone marrow fluid by percol separation. The third passage cultures were used for experiments. Drug treatments for cultures included dexamethasone (10⁻⁷M), lovastatin (10⁻⁶ M), and dexamethasone plus lovastatin for 4 days. BMP-2 mRNA expression was evaluated by RT-PCR. Different responses to drugs between the ON group and the non-ON group were compared.

Results: Bone marrow stroma cells of ON patients were found to be more susceptible to the suppressive effect of dexamethasone on BMP2 expression.

Discussion: Lovastatin stimulated the osteogenesis and reversed the steroid suppressive effect in bone marrow stroma cells in non-ON cases. However, this reverse effect was found to be mild in ON cases.

Introduction: Core binding factor 1 (Cbfa1) is one of the most important transcription factors that direct the osteogenesis of mesenchymal stem cells and osteoblastic functions. It is likely that the factors controlling Cbfa1 expression would trigger the early steps of osteoblast differentiation.

Materials and Methods: By using reporter gene assay for 4.5 kb Cbfa1 promoter, it was found that the first 320 bp of Cbfa1 promoter are active in D1 cells. Within this region, electromobility shift assays delineated a 6 bp of CACATG bound specifically by the proteins from D1 cell nuclear extract. Antibody super-shift and DNA-coupling magnetic bead pull-down assay indicated that the protein bound to this sequence is USF2. Site-specific mutagenesis revealed that this sequences contributed mainly to the activity of 320 bp Cbfa1 promoter.

Discussion: In conclusion, USF2 is the major regulator for the expression of Cbfa1 gene.

Introduction: Nonsteroidal anti-inflammatory drugs (NSAIDs) have been reported to suppress bone repair and remodeling in vivo. Our previous studies showed that NSAIDs inhibited osteoblast proliferation and induced cell death in fetal rat osteoblast cultures. However, the NSAIDs effects on the functions of human osteoblasts remain unclear. Newly developed selective cyclo-oxygenase 2 (COX-2) inhibitors, celecoxib and refecoxib, have been reported to have lower risk of gastrointestinal complications than traditional nonsteroidal anti-inflammatory drugs. A recent report showed that refecoxib decreased bone ingrowth in an animal study. However, the effects of COX-2 selective inhibitors on human osteoblasts have rarely been investigated. In this study, the effects of steroid, non-selective, and selective COX-2 inhibitors on proliferation, cell cycle kinetics, and cytotoxicity in cultured human osteoblasts were examined.

Materials and Methods: Indomethacin,ketorolac,piroxicam, and diclofenac (10⁻⁵ and 10⁻⁴M); dexamethasone (10⁻⁷ and 10⁻⁶M); Celecoxib and DFU, an analogue of rofecoxib, (10⁻⁷–10⁻⁴M) were tested for 24 or 48 hr in human osteoblast cultures.

Results: In this study, we found that a 24 hour treatment of COX-2 selective inhibitors, celecoxib and DFU, significantly inhibited proliferation, arrested cell cycle, and had cytotoxicity in cultured human osteoblasts. However, the inhibitory effect on proliferation could be reversed if these agents were withdrawn for 24 hours. Indomethacin, ketorolac, diclofenac, and piroxicam also significantly inhibited proliferation and arrested cell cycle at the G₀/G₁ phase, but had no cytotoxic effects on human osteoblasts.

Discussion: These results suggest that the COX-2 selective and non-selective NSAIDs may affect osteoblastic functions through different mechanisms.