RESUMO
MicroRNAs are a group of endogenously expressed, single-stranded, 18-24 nt RNAs that regulate diverse cellular pathways. Although documented evidence indicates that some microRNAs can function as oncogenes or tumor-suppressors, the role of miR-214 in regulating human cervical cancer cells remains unexplored. We determined the expression level of miR-214 and found it is downregulated in cervical cancer compared with normal tissue. Overexpression of miR-214 in HeLa cells, a human cervical cancer cell line, significantly inhibited cell proliferation according to the MTT and colony forming assays. HeLa cells that stably overexpress miR-214 downregulate the expression of MEK3 and JNK1 at both mRNA and protein levels. Further investigation revealed that miR-214 regulates the expression of MEK3 and JNK1 by targeting the 3'UTRs of these genes. Collectively, these results suggest that miR-214 negatively regulates HeLa cell proliferation by targeting the noncoding regions of MEK3 and JNK1 mRNAs.
Assuntos
MicroRNAs/genética , Neoplasias do Colo do Útero/genética , Adulto , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo , Feminino , Células HeLa , Humanos , MAP Quinase Quinase 3/biossíntese , MicroRNAs/fisiologia , Pessoa de Meia-Idade , Proteína Quinase 8 Ativada por Mitógeno/biossíntese , RNA Mensageiro/metabolismoRESUMO
RNA interference (RNAi) has been shown to be an effective method for inhibiting the expression of a given gene in human cells by targeting with short duplex RNA (short-interfering RNA or siRNA). However, more and more studies suggest that non-specific effects can be induced by siRNAs, such as off-target inhibition, activation of interferon response, and saturation of cellular silencing machinery. It has been known that more than 90% of human tumors exhibit telomerase activity. Consequently, telomerase is believed to be a broad-spectrum molecular marker of malignancies. In the present study we attempt to develop a tumor-specific RNAi system using the human telomerase reverse transcriptase promoter. This system may provide a basis for RNAi therapy.
Assuntos
Regiões Promotoras Genéticas , RNA Interferente Pequeno/genética , Telomerase/genética , Sequência de Bases , Linhagem Celular , Linhagem Celular Tumoral , Primers do DNA , Genes Reporter , Humanos , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
BACKGROUND: Clinical anti-vascular epithelial growth factor (VEGF) therapy trials faced a major challenge due to upregulated expression of other pro-angiogenic factors, such as fibroblast growth factor-2 (FGF-2). RC28, a novel recombinant dual decoy receptor IgG1 Fc-fusion protein, can block VEGFA and FGF-2 simultaneously. It is designed for the treatment of neovascular age-related macular degeneration and other pathological ocular neovascularization. The present study investigated the prevention efficacy of RC28 on choroidal neovascularization (CNV) in a monkey model and compared to the other mono VEGF antagonists; biodistribution and pharmacokinetics performance were also investigated. METHODS: ELISA and endothelial cell proliferation, migration, and tubule formation assay evaluated the bioactivity of RC28 in vitro, and an initial comparison was made among the mono target antagonists, Bevacizumab (Avastin), Ranibizumab (Lucentis), Aflibercept (EYLEA), Conbercept (KH902), and Ranibizumab (Lucentis). Laser-induced CNV in monkeys, and both VEGF and FGF-2 serum levels were detected in animals before and after the CNV model were induced. RC28 prevention efficacy was compared to other VEGF antagonists on CNV with respect to the incidence of CNV and several ophthalmic examinations. Ocular and systemic levels of RC28 were analyzed by 89Zr-labeled RC28 after single intravitreal administration for the biodistribution and pharmacokinetic profiles. RESULTS: RC28 is a unique fusion protein with high affinity to both VEGF and FGF-2, and beneficial to in vitro and in vivo bioactivity. The in vivo pharmacological studies demonstrated that the incidence of CNV formation was largely reduced in RC28 treatment groups with a low dosage as compared to other VEGF antagonist control groups. Furthermore, traces of RC28 were detected as dispersing from eyeballs to the liver after 20â¯days, and a prolonged half-time pharmacokinetic profile was exhibited.