RESUMO
Naturally fluorescent proteins have been widely used in biological research. In this study, we found that the simple and effective way to obtain enhanced green fluorescent protein (EGFP) nude mice is to cross transgenic EGFP C57BL/6J mice with nude (nu/nu) mice. EGFP expression is identified by tail genotyping. Establishment of the orthotopic EGFP nude mouse model used surgical orthotopic implantation. The morphology and human glioma cell markers, such as glial fibrillary acidic protein (GFAP) and S-100, remain unchanged in this mouse model. The tumor blood vessels obtained from the orthotopic model show brilliant EGFP fluorescence as observed by fluorescence microscopy. These findings suggested that this is an ideal mouse model with which to study interaction among host, tumor, and tumor microenvironment; the findings also suggested that the host (EGFP nude mouse) was involved in tumor angiogenesis.
Assuntos
Modelos Animais de Doenças , Proteínas de Fluorescência Verde/biossíntese , Neoplasias Experimentais , Animais , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Feminino , Glioma/patologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Nus , Camundongos Transgênicos , Neovascularização Patológica/patologia , Reação em Cadeia da PolimeraseRESUMO
UHRF2 is a member of the ubiquitin plant homeo domain RING finger family, which has been proven to be frequently up-regulated in colorectal cancer cells and play a role as an oncogene in breast cancer cells. However, the role of UHRF2 in glioma cells remains unclear. In this study, we performed real-time quantitative PCR on 32 pathologically confirmed glioma samples (grade I, 4 cases; grade II, 11 cases; grade III, 10 cases; and grade IV, 7 cases; according to the 2007 WHO classification system) and four glioma cell lines (A172, U251, U373, and U87). The expression of UHRF2 mRNA was significantly lower in the grade III and grade IV groups compared with the noncancerous brain tissue group, whereas its expression was high in A172, U251, and U373 glioma cell lines. An in vitro assay was performed to investigate the functions of UHRF2. Using a lentivirus-based RNA interference (RNAi) approach, we down-regulated UHRF2 expression in the U251 glioma cell line. This down- regulation led to the inhibition of cell proliferation, an increase in cell apoptosis, and a change of cell cycle distribution, in which S stage cells decreased and G2/M stage cells increased. Our results suggest that UHRF2 may be closely related to tumorigenesis and the development of gliomas.
Assuntos
Neoplasias Encefálicas/genética , Glioma/genética , RNA Mensageiro/genética , RNA Interferente Pequeno/genética , Ubiquitina-Proteína Ligases/genética , Adulto , Apoptose , Encéfalo/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Ciclo Celular , Proliferação de Células , Feminino , Citometria de Fluxo , Seguimentos , Regulação Neoplásica da Expressão Gênica , Glioma/metabolismo , Glioma/patologia , Humanos , Técnicas In Vitro , Masculino , Gradação de Tumores , Prognóstico , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Risco , Células Tumorais Cultivadas , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Ubiquitina-Proteína Ligases/metabolismoRESUMO
BACKGROUND: The gliomas represent the most common primary malignant brain tumors; however, little is known about the molecular pathogenesis of these tumors. Recent research reveals that the oncogenesis and development of gliomas have a close relation to the overexpression of several oncogenes and the inactivation of tumor suppressor genes. Whether the RING finger protein, RNF138, a newly discovered protein, plays a role in glioma oncogenesis is unknown. The present study investigates the expression levels of RNF138 mRNA in glioma samples and noncancerous brain samples and its function in the human glioma cell line U251. METHODS: RT-PCR was used to ascertain the expression of RNF138 mRNA in the glioma cell lines U251, SHG44, U87, A172, and U373. The RNF138 mRNA expression levels of 35 pathological confirmed glioma samples (Grade I - 4 cases, Grade II - 13 cases, Grade III - 11 cases, and Grade IV - 7 cases) and five noncancerous brain tissue samples were analyzed by real-time quantitative PCR. By RNA interference (RNAi) with the lentivirus vector system, the expression of RNF138 was inhibited in the human astrocytomas-glioblastoma multiforme cell line U251. The effects of RNF138-knockdown on cell proliferation were assessed by Cellomics, and cell cycle and cell apoptosis were assessed by FACS. RESULTS: The RNF138 mRNA is expressed in the five glioma cell lines, and its expression level is significantly higher in glioma tissue than in noncancerous brain tissue. By down-regulation of RNF138 expression, U251 cell proliferation was inhibited and cell apoptosis increased. At the same time, S stage cells lessened and G2 stage cells increased. CONCLUSION: The RNF138 gene is highly expressed in glioma tissue and glioma cell lines. It plays an important role in glioma cell proliferation, apoptosis, and cell cycle.