<b>Materials and methods:</b> The levels of 3,4-dihydroxyphenylacetic acid, norepinephrine, serotonin, and 5-hydroxyindoleacetic acid in cancer tissue and in adjacent and non-oncological bone tissue were analysed by high-performance liquid chromatography, and the gene expression of catecholamine receptors and of dopamine β-hydroxylase, monoaminoxidase, ki67, and Runx2 in the osteosarcoma tissue, tissue adjacent to the tumour, non-oncological bone, and human brain tissue was analysed by RT-PCR.
Runx2 protein levels are reduced as expected in MC3T3-E1 cells arrested in late G(1) (by mimosine) or M phase (by nocodazole), but not in cell cycle arrested osteosarcoma cells.
Runx2, p53, and pRB status as diagnostic parameters for deregulation of osteoblast growth and differentiation in a new pre-chemotherapeutic osteosarcoma cell line (OS1).
A loss-of-function approach was used to investigate the effects of small hairpin RNA-mediated knockdown of YAP1 on the expression of RUNX2, CyclinD1, and matrix metalloproteinase-9 (MMP-9) as well as the proliferative activities and invasive potential in OS MG-63 cells (evaluated by MTT and Transwell assays, respectively).
Analysis of previously published OS aCGH data (GSE9654) and aCGH data from this study (GSE19180) identified significant deletion of WWOX in 30% (6/20) of OS samples, whilst significant increase in both RUNX2 and VEGFA gene copy numbers were detected in 55% (11/20) and 60% (12/20) of OS samples, respectively.
Antibodies raised to the unique C-terminal domain were shown to be reactive by immunoprecipitation and immunoblot assay, and were used in confocal immunofluorescence microscopy to reveal low level cytoplasmic staining in osteosarcoma and lymphoma cells that express high levels of Runx2 mRNA.
Based on this knowledge, we found more genes related to the pathogenesis of osteosarcoma and these genes could exert similar function as Runx2, a risk factor confirmed in osteosarcoma, this study may help better understand the genetic mechanism and provide new molecular markers and therapies for osteosarcoma.
Cell growth regulation of Runx2 is also observed in primary calvarial osteoblasts and other osteoblastic cells with relatively normal cell growth characteristics, but not in osteosarcoma cells (e.g.SAOS-2 and ROS17/2.8).
Finally, we revealed that silybin inhibited OS cell viability by altering the protein levels of β-catenin and Runt-related transcription factor 2 (RUNX2) as determined by western blot and immunocytochemistry (ICC).
Furthermore, the RUNX2 binding site, which encompasses positions ‑496 to ‑501 bp, was required to achieve maximal IPO8 promoter activity in Saos‑2 human osteosarcoma cells.
Here we link these two pathways by demonstrating, via gel shift and transient transfection analyses, that Cbfa1 binding to the proximal SOST promoter contributes to differential SOST expression in two osteosarcoma cell lines.
In conclusion, our data reveals that lncRNA SNHG20/miR-139/RUNX2 axis modulates the osteosarcoma tumorigenesis and apoptosis via mitochondrial apoptosis pathway, providing a novel insight for the pathophysiological process.
Intriguingly, runt-related transcription factor 2 (RUNX2) is expressed at higher level in numerous human cancers such as pancreatic cancer and osteosarcoma, indicating that, in addition to its pro-osteogenic role, RUNX2 has a pro-oncogenic potential.
NNT-As1 functions as a cancer-promoting lncRNA by downregulating miR-320a, thus increasing the protein expression level of beta-catenin, RUNX2 and IGF-1R as well as activation of Akt in osteosarcoma.
Nontumorigenic human fibroblasts (CCD-45SK and Hs67), peripheral blood lymphocytes, and several human tumor cell lines derived from cervix, colon, and breast carcinomas, epidermoid carcinoma, and osteosarcoma (HeLa, HT29, MCF7, Saos-2, and A431 cell lines) were studied.
Our analyses show that the RUNX2 interactome may be constitutively activated in osteosarcoma, and that the downstream intracellular pathways are strongly associated with the regulation of osteoblast differentiation and control of cell cycle and apoptosis in osteosarcoma.
Overexpression of RUNX2 protected JQ1-sensitive OS cells from the effect of JQ1, and siRNA-mediated inhibition of RUNX2 sensitized the same cells to JQ1.
Physiologic coupling of osteoblast differentiation to cell cycle withdrawal is mediated through runx2 and p27KIP1, and these processes are disrupted in osteosarcoma.