Shan Hu Huang

Tumor suppressive microRNA-424 inhibits osteosarcoma cell migration and invasion via targeting fatty acid synthase

Numerous studies have recently suggested that miRNAs contribute to the development of various types of human cancer as well as to their invasive and metastatic capacities. The aim of this study was to investigate the functional significance of miR-424 and to identify its possible target genes in osteosarcoma (OS) cells. Previously, inhibition of fatty acid synthase (FASN) has been shown to suppress OS cell proliferation, invasion and migration. The prediction was made using the microRNA.org and TargetScan.human6.0.database. The results showed that FASN is a promising target gene of miR-424. FASN may be a direct target of miR-424 as shown by the luciferase reporter assays. Furthermore, miR-424 expression was increased in osteosarcoma cells by transfection with has-miR-424. FASN mRNA and protein expression levels were measured by RT-PCR and western blot analysis. Cell migration and invasion was measured using Transwell migration and Transwell invasion assays. Expression levels of FASN mRNA and protein were greatly decreased in U2OS cells transfected with has-miR-424. The migration and invasion of cells was significantly decreased by the upregulation of miR-424. These findings suggested that miR-424 plays a key role in inhibiting OS cell migration and invasion through targeting FASN.

Inhibition of fatty acid synthase suppresses osteosarcoma cell invasion and migration via downregulation of the PI3K/Akt signaling pathway in vitro

In the present study, the effect of fatty acid synthase (FASN) inhibition on cell invasion and migration in vitro was investigated. A recombinant plasmid containing a microRNA targeting the FASN gene was used to inhibit FASN expression in U2‑OS cells. Cell migration and invasion were investigated using wound healing and Transwell invasion assays. We found that cell invasion and migration were suppressed by inhibiting FASN. In addition, the effect of inhibition of FASN on phosphorylation of Akt was investigated by detecting the expression levels of pAkt using western blot analysis. Furthermore, protein expression levels of nuclear factor‑κB (NF‑κB; p65) and matrix metalloproteinase (MMP)‑2 and ‑9 were also measured by western blot analysis. Results demonstrated that expression levels of pAkt, NF‑κB (p65) and MMP‑2 and ‑9 proteins were reduced significantly by inhibiting FASN. Therefore, we confirmed that inhibition of FASN by RNA interference suppresses osteosarcoma cell metastasis via downregulation of the phosphoinositide 3‑kinase/Akt/NF‑κB signaling pathway in vitro.

Inhibition of fatty acid synthase supresses osteosarcoma cell invasion and migration.

BACKGROUND Fatty acid synthase (FASN) is overexpressed in a variety of human cancers, and may be involved in cancer metastasis. Hence, the strategies targeted on FASN may have therapeutic potential for treating cancer metastasis. OBJECTIVES The aim of this study is to investigate the correlation of FASN expression with metastasis in human osteosarcoma. MATERIALS AND METHODS Human osteosarcoma cell lines U2-OS and osteosarcoma biopsy specimens were employed in this study. The expression of FASN protein in osteosarcoma specimens was detected by IHC (immunohistochemistry) and the relationship with metastasis was analyzed. We performed the cerulenin, an inhibitor of FASN, to inhibit FASN expression in U2-OS cells. Western blot and RT-PCR were performed to investigate the expression of FASN in U2-OS cells. Cells mobility was detected by wound healing and Transwell assays. RESULTS Results showed that the FASN expression level in the cases with pulmonary metastases was significantly higher than in those without metastasis. In vitro, the invasion and migration of U2-OS cells were suppressed by inhibiting FASN. Our findings suggested that FASN may be involved in osteosarcoma metastasis.

Demethylation-mediated miR-129-5p up-regulation inhibits malignant phenotype of osteogenic osteosarcoma by targeting Homo sapiens valosin-containing protein (VCP)

Previous studies demonstrated that increased Homo sapiens valosin-containing protein (VCP) may be involved in osteosarcoma (OS) metastasis. However, the underlying mechanism of VCP over-expression in OS remains unknown. In the present study, we found a significantly negative correlation between miR-129-5p and VCP protein expression in OS tissues with pulmonary metastasis (Spearmans rho, rs = −0.948). Bioinformatical prediction, Luciferase reporter assay, Western blot, and RT-PCR assays performed on OS cells indicated that VCP is a target of miR-129-5p. In addition, three CPG islands in the region of miR-129-5p promoter were detected by bioinformatical prediction, and significantly higher expression of miR-129-5p and lower methylation level of miR-129-2 gene in OS cells treated with 5-Aza-2′-deoxycytidine (a potent DNA demethylating agent) than in those untreated cells were observed. Furthermore, lower migratory and invasive ability was found in cells with elevated miR-129-5p than in those with decreased miR-129-5p. These findings indicated that increased miR-129-5p may be mediated by demethylation and inhibit OS cell migration and invasion by targeting VCP in OS, and targeting miR-129-5p/VCP signaling pathway may serve as a therapeutic strategy for OS management, although further studies will be necessary.

Inhibiting valosin-containing protein suppresses osteosarcoma cell metastasis via AKT/nuclear factor of kappa B signaling pathway in vitro

BACKGROUND AND AIM The strategies of targeting valosin-containing protein (VCP) may have therapeutic potential for treating cancer metastasis. In this study, we aim to investigate the correlation of VCP protein expression in osteosarcoma (OS) tissues with pulmonary metastasis and its possible molecular mechanism. MATERIALS AND METHODS Expression of VCP in 60 OS specimens was detected by immunohistochemistry (IHC) and the relationship with metastasis was analyzed. An artificial micro ribonucleic acid, targeting VCP, was performed to silence the expression of VCP in U2-OS cells. Cell mobility was detected by wound healing and Transwell assays. Western blot and real-time polymerase chain reaction were performed to investigate the expression of VCP in U2-OS cells. Furthermore, the protein of pAKT (phosphorylated serine/threonine protein kinase) and nuclear factor of kappa B protein 65 were measured by western blot to evaluate the effect of silencing VCP on AKT/nuclear factor of kappa B (NF-kB) signaling pathway. RESULTS The results showed that the expression level of VCP protein in cases with pulmonary metastases was significantly higher than that in those without metastasis (P = 0.004). The invasion and migration of U2-OS cells were suppressed by silencing VCP. Furthermore, silencing VCP could down-regulate the phosphorylation of AKT and nuclear transfer of NF-kB. CONCLUSIONS Our findings suggested that inhibition of VCP could suppress OS cells invasion and migration through down-regulating AKT/NF-kB signaling pathway.

Inhibition of Aurora-B suppresses osteosarcoma cell migration and invasion.

Previous studies have suggested that Aurora-B may be involved in cancer metastasis. However, its role has been poorly evaluated in osteosarcoma (OS). The aim of this study was to investigate the correlation between Aurora-B expression and metastasis in human OS. The human OS cell line, U2-OS, and OS biopsy specimens were used in the study. The expression of Aurora-B protein was examined using immunohistochemistry and western blotting in OS tissues and U2-OS cells, respectively. AZD1152-hydroxyquinazoline-pyrazol-anilide, an inhibitor of Aurora-B, was used to inhibit Aurora-B expression in U2-OS cells. The effect of Aurora-B inhibition on U2-OS cell proliferation, invasion and migration was assessed using MTT, colony formation, wound healing and Transwell assays. The results showed that positive expression of the Aurora-B protein was observed in the nucleus, and that Aurora-B expression levels in the cases with pulmonary metastases were significantly higher than in those without metastasis. In vitro, the proliferation, invasion and migration of U2-OS cells were suppressed by the inhibition of Aurora-B. These results suggest that Aurora-B may be involved in OS metastasis, and may be a promising target in the treatment of OS metastasis.

RNA interference-mediated knockdown of Aurora-B alters the metastatic behavior of A549 cells via modulation of the phosphoinositide 3-kinase/Akt signaling pathway

Accumulating evidence has revealed that an elevated expression level of Aurora-B is associated with metastasis in various types of malignant tumor. However, it is currently unclear whether this molecule is involved in non-small lung cancer (NSCLC) metastasis, and the molecular mechanisms associated with Aurora-B and metastasis remain unknown. In the present study, in order to investigate whether Aurora-B is involved in the development and metastasis of NSCLC, the Aurora-B protein expression in NSCLC tissues was detected by immunohistochemistry and its association with metastasis was analyzed. The results revealed that the expression levels of the Aurora-B protein in tissues obtained from NSCLC patients with lymph node metastasis were significantly higher than those without metastatic disease. Furthermore, the effect of Aurora-B inhibition on A549 cell migration and invasion, as well as the activity of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway was evaluated. Aurora-B was inhibited in the A549 cells using short hairpin RNA, and the cell migration and invasion rates were investigated using wound healing and Transwell invasion assays. In addition, the expression of the main proteins in the PI3K/Akt/nuclear factor-κB (NF-κB) signaling pathway, and matrix metalloproteinase (MMP)-2 and -9 were measured by western blot analysis. The results demonstrated that cell migration and invasion were decreased as a result of silencing Aurora-B. Furthermore, the activity of the PI3K/Akt/NF-κB signaling pathway and the expression of MMP-2 and -9 protein were suppressed by silencing Aurora-B. The results of the present study indicate that the knockdown of Aurora-B suppresses A549 cell invasion and migration via the inhibition of the PI3K/Akt signaling pathway in vitro and thus, targeting Aurora-B may present a potential treatment strategy for NSCLC.

Let-7i inhibits the malignant phenotype of osteosarcoma cells by targeting Aurora-B.

Our previous study indicated that Aurora-B is involved in osteosarcoma (OS) cell invasion and metastasis; however, the mechanism underlying Aurora-B overexpression in OS remains unknown. In the present study, significantly downregulated let-7i expression in OS tissues and OS cells was observed compared with that in normal adjacent tumorous tissues and human osteoblast cell lines. Bioinformatic predictions have revealed a conserved binding site in a microRNA locus on Aurora‑B, suggesting the potential of let‑7i targeting the Aurora‑B gene. To validate this, a luciferase reporter assay was performed on OS cells. The results indicated that Aurora‑B is a likely to be a direct target negatively regulated by let‑7i. The expression of let‑7i in OS cells was restored by infection with let‑7i mimics. Results revealed that Aurora‑B mRNA and protein expression levels were significantly decreased. Furthermore, the proliferation, migration and invasion abilities of OS cells were significantly suppressed by infection with let‑7i mimics. Notably, the inhibitory effect of silencing Aurora‑B by LV‑shAurora‑B on cell proliferation, migratory and invasive ability was significantly lower than that by let‑7i mimics, which indicated that let‑7i inhibits cell malignant phenotypes partially by targeting Aurora‑B in OS cells. All data suggested that let‑7i may be a novel potential target for OS treatment.

Focal fibrocartilaginous dysplasia in the thoracic vertebra: A case report

Focal fibrocartilaginous dysplasia (FFCD) is a rare, paraneoplastic disease that often presents in children and teenagers. Previous studies have reported cases of lesions in the proximal tibia and distal femur, as well as lesions in the upper extremities. The present study describes a case of FFCD on the transverse process and the rib. The imaging findings were found to correspond with the typical observations of FFCD and a biopsy from the nidus revealed pathological results similar to those of previous reports. Thus, the present study demonstrated that FFCD affects tubular bones as well as flat bones. Further studies are required to investigate the underlying mechanism and treatment of FFCD.