RESUMO
MicroRNAs (miRNAs, miR) are of critical importance in growth and metastasis of cancer cells; however, the underlying functions of miRNAs in osteosarcoma (OS) remain largely unknown. This study was aimed to elucidate the role of miR-221 in regulating the biological behavior of OS cells. The proliferation ability was examined by cell counting kit-8 (CCK-8) and cell cycle assay. The abilities of cell migration, invasion, and apoptosis were monitored by transwell assay and flow cytometry, respectively. The effect of miR-221 on cyclin-dependent kinase inhibitor 1B (CDKN1B) expression was evaluated by luciferase assays, real-time polymerase chain reaction, and Western blot analysis. We found that miR-221 was elevated in OS cell lines compared with the normal osteoblastic cell line. Transfection of the miR-221 inhibitor into MG63 and U-2OS cell lines obviously suppressed cell proliferation, migration, and invasion, which is accompanied with cell cycle arrest in G0/G1 phase. Furthermore, luciferase reporter assays indicated that CDKN1B is directly targeted by miR-221 in OS cells. Knockdown of CDKN1B inhibited the effects of miR-221 inhibitor, along with decreased Bax and caspase-3 and increased cyclin E, cyclin D1, Bcl-2, Snail, and Twist1 expression. The results suggested that miR-221 might act as a potentially useful target for treatment of OS.
Assuntos
Apoptose , Neoplasias Ósseas/metabolismo , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , MicroRNAs/metabolismo , Osteossarcoma/metabolismo , RNA Neoplásico/metabolismo , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Movimento Celular , Inibidor de Quinase Dependente de Ciclina p27/genética , Humanos , MicroRNAs/genética , Invasividade Neoplásica , Osteossarcoma/genética , Osteossarcoma/patologia , RNA Neoplásico/genéticaRESUMO
Long noncoding RNA ILF3-AS1 (ILF3-AS1) has been reported to be abnormally expressed in several tumors. However, its expression pattern and function in osteosarcoma have not been investigated. In this study, we showed that ILF3-AS1 expression was significantly up-regulated in both osteosarcoma tissues and cell lines. We first reported that ILF3-AS1 upregulation was induced by nuclear transcription factor SP1. Clinical assays revealed that higher expression of ILF3-AS1 was associated with advanced clinical stage, distant metastasis and shorter overall survival. in multivariate analysis, ILF3-AS1 expression level was found to be an independent prognostic factor for osteosarcoma patients. Functional investigations showed that knockdown of ILF3-AS1 suppressed the proliferation, migration and invasion of osteosarcoma cells, and promoted apoptosis. Bioinformatic software predicted that miR-212 both targeted the 3'-UTR of ILF3-AS1 and SOX5, which was confirmed using luciferase reporter assay, RT-PCR and Western blot. Taken together, ILF3-AS1 displayed its tumor-promotive roles in the progression of osteosarcoma through miR-212/SOX5 axis. Our findings help to elucidate the tumorigenesis of osteosarcoma, and future study will provide a novel therapeutic target for osteosarcoma.
Assuntos
Neoplasias Ósseas/genética , MicroRNAs/genética , Osteossarcoma/genética , RNA Longo não Codificante/genética , Fatores de Transcrição SOXD/genética , Fator de Transcrição Sp1/genética , Neoplasias Ósseas/diagnóstico , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Progressão da Doença , Regulação Neoplásica da Expressão Gênica , Humanos , Osteossarcoma/diagnóstico , Osteossarcoma/patologia , Prognóstico , Regulação para CimaRESUMO
BACKGROUND: Despite the major advances in the treatment, the overall survival of osteosarcoma remains poor. MicroRNAs (miRNAs) are involved in tumorigenesis and progression though modulating their target genes. In the present study, the roles of miR-1285-3p in osteosarcoma was investigated. METHODS: Microarray profiling was applied to distinguish the up and down regulated microRNAs in osteosarcoma. Quantitative real-time PCR (qRT-PCR) assay was performed to detect the expression of miR-1285-3p and YAP1 expression. MTT and transwell assays were carried out to determine the cells proliferation and invasion respectively. Moreover, dual luciferase reporter assay was performed to evaluate the binding efficiency between miR-1285-3p and the 3'UTR of YAP1. RESULTS: MiR-1285-3p was down regulated in osteosarcoma tissues and cell lines and the reduction of miR-1285-3p expression predicted a poor overall survival of osteosarcoma patients. Ectopic expression of miR-1285-3p inhibited osteosarcoma cell proliferation, colony formation and invasion. In addition, YAP1 was further demonstrated as a direct target of miR-1285-3p. Moreover, overexpression of YAP1 reversed the inhibitory effects of miR-1285-3p on osteosarcoma cells proliferation and invasion. CONCLUSIONS: MiR-1285-3p which was low expressed in osteosarcoma inhibited the proliferation and invasion of osteosarcoma cells via direct targeting YAP1. These results suggested that miR-1285-3p might be a potential therapeutic targets and biomarker in osteosarcoma.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Ósseas/diagnóstico , Neoplasias Ósseas/metabolismo , MicroRNAs/metabolismo , Osteossarcoma/diagnóstico , Osteossarcoma/metabolismo , Fosfoproteínas/genética , Regiões 3' não Traduzidas , Biomarcadores Tumorais/genética , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Regulação para Baixo/genética , Feminino , Expressão Gênica , Humanos , Masculino , MicroRNAs/genética , Invasividade Neoplásica , Osteossarcoma/genética , Osteossarcoma/patologia , Prognóstico , Ligação Proteica , Taxa de Sobrevida , Fatores de Transcrição , Proteínas de Sinalização YAP , Adulto JovemRESUMO
OBJECTIVE: To explore the inhibitory effect of bevacizumab, a vascular endothelial growth factor antibody, on angiogenesis in human osteosarcoma of nude mice. METHODS: Twenty-one nude mice were inoculated with red fluorescent protein (RFP)-labeled human osteosarcoma cell line 143B-RFP, that is, clones that expressed RFP in the cytoplasm, and randomly assigned to one of three groups: G1 (Control group, injected with saline solution); G2 (intraperitoneal bevacizumab 2 mg/kg twice per week) and G3 (intraperitoneal bevacizumab 5 mg/kg, twice per week). The tumor-bearing mice were examined in a fluorescence light box that was illuminated periodically. The primary tumors were measured by fluorescence imaging weekly and their volumes calculated. RESULTS: The mean tumor volumes were significantly smaller in the G3 (186.4 ± 100.8 mm(3) ) than the control group (587.0 ± 406.8 mm(3) ) (P < 0.05) on Day 31, and again significantly smaller in the G3 (677.3 ± 461.9 mm(3) ) than the control group (3162.6 ± 1529.2 mm(3) ) on Day 38 (P < 0.01). The average tumor volume in the G2 group was 493.5 ± 425.4 mm(3) on Day 31 and 1870.1 ± 1524.8 mm(3) on Day 38. The effect on tumor volume was greater in the G3 than the G2 group. Three mice in the G2 group, four in the G3 group and four in the control group developed lung metastases that were confirmed by pathological examination; these differences were not statistically significant (P < 0.05). CONCLUSIONS: Bevacizumab exhibits strong antiangiogenesis activity in experimental osteosarcoma in a nude mouse model but does not influence the incidence of lung metastasis. Our findings may have considerable potential for the treatment of osteosarcoma.