1,25(OH)2D3 inhibits osteogenic differentiation through activating ß­catenin signaling via downregulating bone morphogenetic protein 2.

The present study explored whether bone morphogenetic proteins (BMPs) and Wnt/ß­catenin signaling pathways were involved in the 1,25(OH)2D3­induced inhibition of osteogenic differentiation in bone marrow­derived mesenchymal stem cells (BMSCs). To evaluate the osteogenic differentiation of BMSCs, the expression levels of ossification markers, including BMP2, Runt­related transcription factor 2 (Runx2), Msh homeobox 2 (Msx2), osteopontin (OPN) and osteocalcin (OCN), and the activity of alkaline phosphatase (ALP), as well as the calcified area observed by Alizarin red­S staining, were investigated. Chromatin immunoprecipitation (ChIP) assay was used to detect the effect of 1,25(OH)2D3 on the DNA methylation and histone modification of BMP2, while an immunoprecipitation (IP) assay was performed to assess the crosstalk between Smad1 and disheveled­1 (Dvl­1) proteins. It was observed that 1,25(OH)2D3 significantly decreased the expression levels of BMP2, Runx2, Msx2, OPN and OCN, and reduced ALP activity and the calcified area in BMSCs, whereas these effects were rescued by BMP2 overexpression. ChIP assay revealed that BMSCs treated with 1,25(OH)2D3 exhibited a significant increase in H3K9me2 level and a decrease in the acetylation of histone H3 at the same BMP2 promoter region. In addition, 1,25(OH)2D3 treatment promoted the nuclear accumulation of ß­catenin by downregulating BMP2. Furthermore, the ß­catenin signaling inhibitor XAV­939 weakened the inhibitory effect of 1,25(OH)2D3 on osteogenic differentiation. Additionally, knockdown of ß­catenin rescued the attenuation in Dvl­1 and Smad1 interaction caused by 1,25(OH)2D3. Overexpression of Smad1 also reversed the inhibitory effect of 1,25(OH)2D3 on osteogenic differentiation. Taken together, the current study demonstrated that 1,25(OH)2D3 inhibited the differentiation of BMSCs into osteoblast­like cells by inactivating BMP2 and activating Wnt/ß­catenin signaling.

Small molecule TSC01682 inhibits osteosarcoma cell growth by specifically disrupting the CUL4B-DDB1 interaction and decreasing the ubiquitination of CRL4B E3 ligase substrates.

The direct interaction between Cullin 4B (CUL4B) and DNA damage-binding protein 1 (DDB1) is required for the assembly of Cullin4B-RING E3 ligase complex (CRL4B), which are involved in the tumorigenesis of osteosarcoma through ubiquitinating and degrading multiple tumor suppressors and cell cycle regulators. Thus, targeting CUL4B-DDB1 interaction to prevent the assembly of CRL4B may be a potent approach to inhibit osteosarcoma cell growth. In the present study, we identified six naturally-sourced small molecules that can specifically disrupt the CUL4B-DDB1 interaction using an in vitro high-throughput screening (HTS) system in yeast. We focused our investigation on revealing the molecular effects of TSC01682, the most active compound capable of inhibiting osteosarcoma cell growth. Biochemically, TSC01682 significantly repressed the CUL4B-DDB1 interaction in both yeast cells and osteosarcoma cells. Moreover, TSC01682 treatment in osteosarcoma cells also caused a decrease of other CRL4B components including CUL4-associated factor 11 (DCAF11) and DCAF13, but an increase of two CRL4B substrates including cyclin-dependent kinase inhibitor 1A (CDKN1A, also known as p21) and phosphatase and tensin homolog deleted on chromosome 10 (PTEN) through inhibiting their ubiquitination. Consistent with these molecular changes, TSC01682 treatment significantly inhibited cell proliferation, colony formation, invasion, and in vivo tumor growth. Collectively, our results suggest that TSC01682 is a potent compound capable of disrupting the CUL4B-DDB1 interaction, and it may be developed as a chemotherapeutic drug for osteosarcoma treatment.

Long non-coding RNA HOXA-AS2 promotes migration and invasion by acting as a ceRNA of miR-520c-3p in osteosarcoma cells.

Osteosarcoma (OS) is the commonest primary malignant tumour originating from bone. Previous studies demonstrated that long non-coding RNAs (lncRNAs) could participate in both oncogenic and tumor suppressing pathways in various cancer, including OS. The HOXA cluster antisense RNA2 (HOXA-AS2) plays an important role in carcinogenesis, however, the underlying role of HOXA-AS2 in OS progression remains unknown. The aim of the present study was to evaluate the expression and function of HOXA-AS2 in OS. The qRT-PCR analysis was to investigate the expression pattern of HOXA-AS2 in OS tissues. Then, the effects of HOXA-AS2 on cell proliferation, cell cycle, apoptosis, migration, and invasion were assessed in OS in vitro. Furthermore, bioinformatics online programs predicted and luciferase reporter assay were used to validate the association of HOXA-AS2 and miR-520c-3p in OS cells. We observed that HOXA-AS2 was up-regulated in OS tissues. In vitro experiments revealed that HOXA-AS2 knockdown significantly inhibited OS cells proliferation by promoting apoptosis and causing G1 arrest, whereas HOXA-AS2 overexpression promoted cell proliferation. Further functional assays indicated that HOXA-AS2 significantly promoted OS cell migration and invasion by promoting epithelial-mesenchymal transition (EMT). Bioinformatics online programs predicted that HOXA-AS2 sponge miR-520c-3p at 3'-UTR with complementary binding sites, which was validated using luciferase reporter assay. HOXA-AS2 could negatively regulate the expression of miR-520c-3p in OS cells. In conclusion, our study suggests that HOXA-AS2 acts as a functional oncogene in OS.

Increased risk of vertebral fracture in patients with rheumatoid arthritis: A meta-analysis.

The relationship between rheumatoid arthritis and risk of vertebral fracture has been reported by several observational studies. However, there is no higher-level evidence study, such as meta-analysis, that has investigated the relationship, and its mechanisms are not yet fully clear. This meta-analysis aimed to provide a summary of an observational study of the relationship between rheumatoid arthritis and the risk of vertebral fractures.Relevant studies were identified by searching PubMed and EMBASE databases (up to August 1, 2016). We included published observational studies (cohort or case-control design) evaluating the relationship between rheumatoid arthritis and the risk of vertebral fractures. Two reviewers searched and abstracted the data independently. Relative risks (RRs) with 95% confidence intervals (CIs) were used throughout the whole analysis.Seven observational studies (2 cohort studies, 2 nested case-control studies, and 3 case-control studies) were included in this meta-analysis. The results showed that the pooled RR of vertebral fracture for individuals with rheumatoid arthritis was 2.34 (95% CI 2.05-2.63, I = 35.4%, P for heterogeneity = 0.16). Further subgroup analysis by sex showed that the pooled RRs for both women and men, and only women were 2.14 (95% CI 1.47-2.8, I = 48.5%, P for heterogeneity = 0.12) and 2.39 (95% CI 2.07-2.70, I = 34%, P for heterogeneity = 0.22), respectively. Subgroup analysis by study design showed that the pooled RRs for cohort studies, nested case-control studies, and case-control studies were 2.31 (95% CI 1.95-2.67, I = 4.8%, P for heterogeneity = 0.31), 1.89 (95% CI 1.01-2.77, I = 72.1%, P for heterogeneity = 0.06), and 2.62 (95% CI 2.04-3.91, I = 26.1%, P for heterogeneity = 0.26), respectively.Based on our meta-analysis, rheumatoid arthritis should be regarded as an independent risk factor of vertebral fracture. Further studies are needed to institute prevention and treatment strategies.

FBXW7 acts as an independent prognostic marker and inhibits tumor growth in human osteosarcoma.

F-box and WD repeat domain-containing 7 (FBXW7) is a potent tumor suppressor in human cancers including breast cancer, colorectal cancer, gastric cancer and hepatocellular carcinoma. In this study, we found that the expressions of FBXW7 protein and mRNA levels in osteosarcoma (OS) cases were significantly lower than those in normal bone tissues. Clinical analysis indicated that FBXW7 was expressed at lower levels in OS patients with advanced clinical stage, high T classification and poor histological differentiation. Furthermore, we demonstrated that high expression of FBXW7 was correlated with a better 5-year survival of OS patients. Multivariate Cox regression analysis indicated that FBXW7 was an independent prognostic marker in OS. Our in vitro studies showed that FBXW7 overexpression inhibited cell cycle transition and cell proliferation, and promoted apoptosis in both U2OS and MG-63 cells. In a nude mouse xenograft model, FBXW7 overexpression slowed down tumor growth by inducing apoptosis and growth arrest. Mechanistically, FBXW7 inversely regulated oncoprotein c-Myc and cyclin E levels in both U2OS and MG-63 cells. Together these findings suggest that FBXW7 may serve as a prognostic biomarker and inhibit tumor progression by inducing apoptosis and growth arrest in OS.

Functional annotation of rheumatoid arthritis and osteoarthritis associated genes by integrative genome-wide gene expression profiling analysis.

BACKGROUND: Rheumatoid arthritis (RA) and osteoarthritis (OA) are two major types of joint diseases that share multiple common symptoms. However, their pathological mechanism remains largely unknown. The aim of our study is to identify RA and OA related-genes and gain an insight into the underlying genetic basis of these diseases. METHODS: We collected 11 whole genome-wide expression profiling datasets from RA and OA cohorts and performed a meta-analysis to comprehensively investigate their expression signatures. This method can avoid some pitfalls of single dataset analyses. RESULTS AND CONCLUSION: We found that several biological pathways (i.e., the immunity, inflammation and apoptosis related pathways) are commonly involved in the development of both RA and OA. Whereas several other pathways (i.e., vasopressin-related pathway, regulation of autophagy, endocytosis, calcium transport and endoplasmic reticulum stress related pathways) present significant difference between RA and OA. This study provides novel insights into the molecular mechanisms underlying this disease, thereby aiding the diagnosis and treatment of the disease.

Correlation of synovial fluid HMGB-1 levels with radiographic severity of knee osteoarthritis.

PURPOSE: This study measured high-mobility group box 1 (HMGB-1) levels in serum and synovial fluid (SF) in patients with primary knee osteoarthritis (OA) and correlated these levels with radiographic disease severity. METHODS: Seventy-eight OA patients and 30 controls were enrolled in this study. All OA patients were scored according to the Kellgren-Lawrence (KL) grading system. HMGB-1 levels were measured by enzyme-linked immunosorbent assay (ELISA). RESULTS: SF HMGB-1 levels were significantly higher in knee OA patients, compared with controls (P < 0.01). Moreover, SF HMGB-1 levels were positively associated with KL scores (P < 0.01). Multinomial logistic regression demonstrated that the SF HMGB-1 level was an independent factor for radiographic severity of OA (P=0.002); however, serum HMGB-1 levels did not differ significantly between OA patients and controls and did not correlate with KL scores (P > 0.05). CONCLUSION: These results demonstrate that HMGB-1 levels in SF of knee OA patients are independently associated with radiographic disease severity.