Aims

The effects of remnant preservation on the anterior cruciate ligament (ACL) and its relationship with the tendon graft remain unclear. We hypothesized that the co-culture of remnant cells and bone marrow stromal cells (BMSCs) decreases apoptosis and enhances the activity of the hamstring tendons and tenocytes, thus aiding ACL reconstruction.

Aims

Osteoarthritis (OA) is prevalent among the elderly and incurable. Intra-articular parathyroid hormone (PTH) ameliorated OA in papain-induced and anterior cruciate ligament transection-induced OA models; therefore, we hypothesized that PTH improved OA in a preclinical age-related OA model.

Aims

Proliferation, migration, and differentiation of anterior cruciate ligament (ACL) remnant and surrounding cells are fundamental processes for ACL reconstruction; however, the interaction between ACL remnant and surrounding cells is unclear. We hypothesized that ACL remnant cells preserve the capability to regulate the surrounding cells’ activity, collagen gene expression, and tenogenic differentiation. Moreover, extracorporeal shock wave (ESW) would not only promote activity of ACL remnant cells, but also enhance their paracrine regulation of surrounding cells.

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: 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.