RESUMO
The role of microRNAs (miRNAs) in glioma biology is increasingly recognized. To investigate the regulatory mechanisms governing the malignant signature of gliomas with different grades of malignancy, we analyzed miRNA expression profiles in human grade I-IV tumor samples and primary glioma cell cultures. Multiplex real-time PCR was used to profile miRNA expression in a set of World Health Organization (WHO) grade I (pilocytic astrocytoma), II (diffuse fibrillary astrocytoma), and IV (glioblastoma multiforme) astrocytic tumors and primary glioma cell cultures. Primary glioma cell cultures were used to evaluate the effect of transfection of specific miRNAs and miRNA inhibitors. miRNA microarray showed that a set of miRNAs was consistently upregulated in all glioma samples. miR-363 was upregulated in all tumor specimens and cell lines, and its expression correlated with tumor grading. The transfection of glioma cells with the specific inhibitor of miR-363 increased the expression level of tumor suppressor growth-associated protein 43 (GAP-43). Transfection of miR-363 induced cell survival, while inhibition of miR-363 significantly reduced glioma cell viability. Furthermore, miRNA-363 inhibition induced the downregulation of AKT, cyclin-D1, matrix metalloproteinase (MMP)-2, MMP-9, and Bcl-2 and upregulation of caspase 3. Together, these data suggest that the upregulation of miR-363 may play a role in malignant glioma signature.
Assuntos
Apoptose , Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/patologia , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Glioma/patologia , MicroRNAs/genética , Adulto , Idoso , Biomarcadores Tumorais/genética , Western Blotting , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/cirurgia , Seguimentos , Proteína GAP-43/genética , Proteína GAP-43/metabolismo , Glioma/genética , Glioma/metabolismo , Glioma/cirurgia , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Gradação de Tumores , Invasividade Neoplásica , Prognóstico , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Taxa de Sobrevida , Células Tumorais Cultivadas , Proteínas rap1 de Ligação ao GTP/genética , Proteínas rap1 de Ligação ao GTP/metabolismoRESUMO
Adrenocortical dysplasia (ACD) is a shelterin protein involved in the maintenance of telomere length and in cancer radioresistance. This study investigated the expression profile of ACD in human gliomas and its role in radioresistance of glioma cells. The expression of ACD was analyzed in 62 different grades of glioma tissues and correlated with prognosis. A radioresistant cell line was generated from U87MG cells. For mechanistic studies, ACD was inhibited by small interfering RNA-targeting ACD and the effect on cell radioresistance, telomerase activity, cyclinD1, caspase-3, hTERT, and BIRC1 was evaluated. Clonogenic assay was performed after irradiation, to investigate the effect of ACD silencing on radiation sensitivity. ACD expression appeared strongly upregulated in higher grade gliomas, and its expression was significantly correlated to grading and poor prognosis. In glioma cell lines, ACD expression pattern was similar to those observed in glioma tissues. In irradiated cells, ACD expression was increased in an ionizing radiation dose-dependent manner. A higher expression of ACD was observed in the radioresistant clones than parental cells. Silencing of ACD led to the enhanced radiation sensitivity, decreased telomerase activity and cyclin D1 expression, reduced expression of BIRC1, and finally to the upregulation of caspase-3. This study represents the first report, which demonstrated the expression pattern of ACD in gliomas and its prognostic value. Our results suggested that ACD is involved in glioblastoma radioresistance, likely through the modulation of telomerase activity, proliferation, and apoptosis. ACD might represent a potential molecular biomarker and a novel therapeutic target in glioblastoma.