Steroid-induced osteonecrosis of the femoral head (SONFH) is a common debilitating orthopedic disease. The bone marrow mesenchymal stem cells (BMSCs) are a type of mesenchymal stem cells which play crucial roles in bone repair. The adipogenic/osteogenic differentiation disorder of BMSCs has been widely perceived contributing to SONFH. However, the regulatory mechanism of BMSCs differentiation disorder still remains unclear. Circular RNA (circRNA), a kind of stable ncRNA, plays important roles in regulating gene expression via various ways. To date, there are no studies to uncover the circRNA expression profile and screen out the key circRNAs playing crucial roles in adipogenic/osteogenic differentiation disorder of SONFH-BMSCs. In present study, we detected the circRNA expression profiles in SONFH-BMSCs for the first time. A total of 820 circRNAs were differentially expressed in SONFH-BMSCs, including 460 up- and 360 down-regulated circRNAs. Bioinformatics analysis indicates circRNA CDR1as, one up-regulated circRNA, may play crucial role in adipogenic/osteogenic differentiation disorder of SONFH-BMSCs via CDR1as-miR-7-5p-WNT5B axis. Knocking-down CDR1as resulted in increasing of osteogenic differentiation and decreasing of adipogenic differentiation of BMSCs, while over-expressing CDR1as resulted in decreasing of osteogenic differentiation and increasing of adipogenic differentiation of BMSCs. The miR-7-5p binding sites of CDR1as and WNT5B were verified by luciferase reporter gene assay. Our study may provide new insights into the molecular mechanisms of osteogenic/adipogenic differentiation disorder of SONFH-BMSCs and new biomarkers for the diagnosis and treatment of SONFH.

Long noncoding RNAs (lncRNAs) are critical gene expression regulators and are involved in several bone diseases. To explore the potential roles of lncRNAs in osteonecrosis of the femoral head (ONFH), we investigated for the first time the lncRNA expression profile of bone marrow mesenchymal stem cells (BMSCs) from patients with steroid-induced ONFH (SONFH) with microarray and bioinformatics analysis. A total of 1878 lncRNAs and 838 mRNAs were significantly up-regulated while 1842 lncRNAs and 1937 mRNAs were statistically down-regulated in the SONFH group compared with control group. The results validated by qRT-PCR were consistent with the microarray profiling data, especially involved in upregulation and downregulation of critical lncRNAs as well as mRNAs expression related to adipogenic and osteogenic differentiation. Pathway analyses revealed 40 signaling pathways with significant differences, especially the signaling pathways to regulate stem cell pluripotency. The CNC and ceRNA network indicated that lncRNA RP1-193H18.2, MALAT1 and HOTAIR were associated with abnormal osteogenic and adipogenic differentiation of BMSCs in the patients with SONFH. Our results suggest the lncRNA expression profiles were closely associated with the abnormal adipogenic and osteogenic transdifferentiation of BMSCs during the development of SONFH and explore a new insight into the molecular mechanisms of SONFH.

Osteosarcoma (OS) is a primary malignant bone tumor with a high fatality rate. Many circRNAs have been proved to play important roles in the pathogenesis of some diseases. However, the occurrence of circRNAs in OS remains little known. The circular RNA (circRNA) expression file GSE96964 dataset, which included seven osteosarcoma cell lines and one control sample (osteoblast cell line), was downloaded from the Gene Expression Omnibus (GEO) database to explore the potential function of circRNAs in osteosarcoma by competing endogenous RNA (ceRNA) analysis. Three gene expression profiles of OS were downloaded from GEO database and then used for the pathway enrichment analysis, Venn analysis and protein-protein interaction (PPI) network analysis. Real-time qPCR validation and RNA interference were conducted to verify our prediction. Differentially expressed circRNAs between OS and control, including 8 up-regulated and 102 down-regulated circRNAs, were generated and ceRNA analysis for 5 most up-regulated or 5 most down-regulated circRNAs in OS were then performed. The pathway enrichment analysis of gene expression profiles indicated differentially expressed genes (DEGs) of three gene profiles significantly enriched in cell cycle pathway, cell adhesion molecules (CAMs) pathway, oxidative phosphorylation pathway, cytokine-cytokine receptor interaction pathway, p53 signaling pathway and proteoglycans in cancer pathway, which were critical important pathways in the pathogenesis of OS. The Venn analysis showed that 2 (one is a pseudogene) up-regulated and 39 down-regulated DEGs were co-expressed in all three gene profiles. Then PPI networks of 41 co-expressed DEGs (up- and down-regulated DEGs) were constructed to predict their functions using the GeneMANIA. The expression levels of these related RNAs also matched our predictions really well. Ultimately, we found cell adhesion molecule 1 (CADM1) gene was not only a co-expression mRNA of the three mRNA expression profiles of OS, but also are predicted to be regulated by hsa_circ_0032462, hsa_circ_0028173, hsa_circ_0005909 by functioning as miRNAs ‘Sponge’ in human osteosarcoma. These over-expressed circRNAs may result in the over expression of CADM1 which promote the development of OS. We envision this discovery of these important moleculars, incuding hsa_circ_0032462, hsa_circ_0028173, hsa_circ_0005909 and CADM1 may lead to further development of new concepts, thus allowing for more opportunities in diagnosis and therapy of OS.

Background: It has been well known that the degeneration of hip articular cartilage with osteonecrosis of the femoral head (ONFH) increases the instability of hip and accelerates the development process of ONFH. A better understanding of the expression of chondrogenesis-related and arthritis-related genes of cartilage along with the progression of ONFH seems to be essential for further insight into the molecular mechanisms of ONFH pathogenesis. Methods: We analyzed the differentially expressed gene profile (GSE74089) of human hip articular cartilage with ONFH. The functions and pathway enrichments of differentially expressed genes (DEGs) were analyzed via GO and KEGG analysis. The expression of six selected critical chondrogenesis-related and four arthritis-related genes in eight human hip articular cartilage with femoral neck fracture (FNF) and 26 human hip articular cartilage with different stages ONFH (6 cases of Ficat stage II, 10 cases of Ficat stage III and 10 cases of Ficat stage IV) were detected. Results: A total of 2,174 DEGs, including 1,482 up-regulated and 692 down-regulated ones, were obtained in the ONFH cartilage specimens compared to the control group. The GO and KEGG enrichment analysis indicated that the function of these DEGs mainly enriched in extracellular matrix, angiogenesis, antigen processing and presentation. The results showed a significant stepwise up-expression of chondrogenesis-related genes, including MMP13, ASPN, COL1A1, OGN, COL2A1 and BMP2, along with the progression of ONFH. The arthritis-related genes IL1β, IL6 and TNFα were only found up-expressed in Ficat IV stage which indicated that the arthritis-related molecular changes were not significant in the progression of ONFH before Ficat III stage. However, the arthritis-related gene PTGS2 was significant stepwise up-expression along with the progression of ONFH which makes it to be a sensitive arthritis-related biomarker of ONFH. Conclusion: Expression changes of six chondrogenesis-related and four arthritis-related genes were found in hip articular cartilage specimens with different ONFH Ficat stages. These findings are expected to a get a further insight into the molecular mechanisms of ONFH progression.

Multiple gene polymorphisms have been demonstrated to correlate with the susceptibility to osteonecrosis of the femoral head (ONFH). However, as a complex disease induced by multiple genes, the development of ONFH has rarely been reported to involve in gene interaction. In this study, we first explored the association of 10 variants interactions in receptor activator of nuclear factor-kappa B (RANK), RANK ligand (RANKL), osteoprotegerin (OPG), tumor necrosis factor receptor-associated factor 6 (TRAF6), and nuclear factor of activated T cells cytoplasmic 1 (NFATC1) genes with the development and clinical phenotypes of ONFH in a 377 ONFH case–control study with using Mass ARRAY^® platform. Our results showed that not only a total of 6 interactional variants in the paired 10 variants interactions were significantly associated with the development of ONFH (OPG rs2073617 and NFATC1 rs754093, p < 0.019; OPG rs2073618 and NFATC1 rs754093, p < 0.008; OPG rs2073617 and RANKL rs1054016, p < 0.039, respectively) but also a total of 4 paired interactional variants were found to involve significantly in the increased risk of bilateral hip lesions in ONFH (OPG rs2073617 and TRAF6 rs5030411, p = 0.044; RANK rs884205 and TRAF6 rs5030411, p = 0.045, respectively). Moreover, the results from generalized multifactor dimensionality reduction also showed that the five best models were identified and associated significantly with ONFH risk, p = 0.001, 0.01, 0.01, 0.01, and 0.01, respectively. Our results first suggest that the variants in RANK/RANKL/OPG pathway genes affected the development of ONFH in gene interaction manner through the interaction of the paired variants and multiple variants.

Osteoarthritis (OA) is one of the most common diseases worldwide, but the pathogenic genes and pathways are largely unclear. The aim of this study was to screen and verify hub genes involved in OA and explore potential molecular mechanisms. The expression profiles of GSE12021 and GSE55235 were downloaded from the Gene Expression Omnibus (GEO) database, which contained 39 samples, including 20 osteoarthritis synovial membranes and 19 matched normal synovial membranes. The raw data were integrated to obtain differentially expressed genes (DEGs) and were deeply analyzed by bioinformatics methods. The Gene Ontology (GO) and pathway enrichment of DEGs were performed by DAVID and Kyoto Encyclopedia of Genes and Genomes (KEGG) online analyses, respectively. The protein-protein interaction (PPI) networks of the DEGs were constructed based on data from the STRING database. The top 10 hub genes VEGFA, IL6, JUN, IL1β, MYC, IL4, PTGS2, ATF3, EGR1, and DUSP1 were identified from the PPI network. Module analysis revealed that OA was associated with significant pathways including TNF signaling pathway, cytokine-cytokine receptor interaction, and osteoclast differentiation. The qRT-PCR result showed that the expression level of IL6, VEGFA, JUN, IL-1β, and ATF3 was significantly increased in OA samples (p < 0.05), and these candidate genes could be used as potential diagnostic biomarkers and therapeutic targets of OA.