RESUMO
Osteosarcoma is the most common primary malignant bone tumor in children and adolescents. However, the underlying mechanism of osteosarcoma carcinogenesis and progression remains unknown. In the present study, we evaluated the expression profile of miRNAs in osteosarcoma tissues and the adjacent normal tissues. We found that the expression of miR-422a was down-regulated in osteosarcoma tissues and cell lines. In addition, we observed significantly elevated levels of repressive H3K9me3 and H3K27me3 and decreased active H3K4me3 on the promote region of miR-422a in osteosarcoma cells and clinical samples. Furthermore, up-regulation of miR-422a exhibited both in vitro and in vivo anti-tumor effects by inhibiting osteosarcoma cell growth and inducing apoptosis and cell cycle arrest. We also found that miR-422a targeted BCL2L2 and KRAS and negatively regulated their protein expression. Furthermore, restoration of miR-422a and knockdown of BCL2L2 and KRAS promoted apoptosis and induce cell cycle arrest in osteosarcoma cells. Taken together, the present study demonstrates that miR-422a may serve as a tumor suppressor in osteosarcoma via inhibiting BCL2L2 and KRAS translation both in vitro and in vivo Therefore, miR-422a could be developed as a novel therapeutic target in osteosarcoma.
Assuntos
Proteínas Reguladoras de Apoptose/biossíntese , Apoptose , Neoplasias Ósseas/metabolismo , Proliferação de Células , Genes Supressores de Tumor , MicroRNAs/metabolismo , Osteossarcoma/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/biossíntese , RNA Neoplásico/metabolismo , Proteínas Reguladoras de Apoptose/genética , Neoplasias Ósseas/genética , Neoplasias Ósseas/patologia , Linhagem Celular , Feminino , Humanos , Masculino , MicroRNAs/genética , Osteossarcoma/genética , Osteossarcoma/patologia , Biossíntese de Proteínas , Proteínas Proto-Oncogênicas p21(ras)/genética , RNA Neoplásico/genéticaRESUMO
We aim to investigate the effect of miR-106a-5p on the proliferation, migration, and invasion of osteosarcoma (OS) cells by targeting high-mobility group AT-hook 2 (HMGA2). Real-time fluorescent quantitative polymerase chain reaction (RT-qPCR) was used for detecting the expressions of miR-106-5p and HMGA2 in 137 OS and adjacent normal bone tissues. Immunohistochemistry was applied for the HMGA2 protein expression detection. Luciferase reporter gene assay was conducted for verifying whether miR-106-5p targeted HMGA2. MG63 and U2SO cells were respectively divided into five groups: Blank, miR-106a-5p, scramble, HMGA2-siRNA, and miR-106a-5p+HMGA2 groups. RT-qPCR and western blot were applied for detecting the expressions of miR-106a-5p and HMGA2 in five groups. Proliferation rate, cell cycle, invasion, and migration ability of OS cells were detected using methyl thiazolyl-tetrazolium, 5-ethynyl-2'-deoxyuridine (Edu) assay, flow cytometry, and Transwell. Compared with adjacent normal tissues, OS tissues presented with decreased miR-106a-5p expressions, elevated HMGA2 mRNA, and positive expressions (all p < 0.05). The sensitivity and specificity of miR-106a-5p were 97.8%, 93.43%, and HMGA2 mRNA were 97.8%, 99.27%, separately. miR-106a-5p and HMGA2 expressions were associated with tumor size, Enneking stage, distant metastasis, and lung metastasis. Expressions of HMGA2 in OS cells in miR-106a-5p and HMGA2 siRNA groups were both significantly decreased with the same downregulation level, and the proliferation rates in both groups were obviously slowed down after 48 h (both p < 0.001). Edu positive cells, S phase cells (majority of cells blocked at G0/G1 phase), migratory and invasive cells were obviously decreased (all p < 0.05). Downregulation of miR-106a-5p was found in OS tissues, and upregulation of miR-106a-5p can inhibit the proliferation, migration, and invasion by targeting HMGA2 in OS cells.
Assuntos
Neoplasias Ósseas/genética , Regulação Neoplásica da Expressão Gênica , Proteína HMGA2/genética , Neoplasias Pulmonares/genética , MicroRNAs/genética , Osteossarcoma/genética , Adolescente , Neoplasias Ósseas/metabolismo , Neoplasias Ósseas/patologia , Linhagem Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Criança , Pré-Escolar , Feminino , Proteína HMGA2/antagonistas & inibidores , Proteína HMGA2/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundário , Masculino , MicroRNAs/metabolismo , Invasividade Neoplásica , Osteoblastos/metabolismo , Osteoblastos/patologia , Osteossarcoma/metabolismo , Osteossarcoma/secundário , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Adulto JovemRESUMO
Diverse functions of microRNAs have been investigated in tumorigenesis in osteosarcoma (OS), involving the regulation of proliferation, invasion, migration, apoptosis and drug resistance. MiR-367 was found to be an oncogene and increased in OS. However, the function of miR-367 in drug resistance in OS cells is still unknown. In this study, we found that miR-367 was up-regulated in OS tissues and OS cell cultures. Meanwhile, treatment with adriamycin (ADR) induced apoptosis of OS cells with upregulation of miR-367. Notably, KLF4 was demonstrated to be a direct target of miR-367 by gene reporter assay, and miR-367 significantly blocked both mRNA and protein level of KLF4. In addition, overexpression of miR-367 markedly suppressed the increase of KLF4 induced by ADR in OS cells, as well as Bax and cleaved caspase-3, which were significantly reversed by anti-miR-367 transfection. Taken together, our data demonstrates that miR-367 and KLF4 play important roles in OS treatment and ADR resistance, suggesting that miR-367 is a potential biomarker of chemotherapy resistance in OS and also probably a novel therapeutic target against OS.
