RESUMO
BACKGROUND: Tumour stromal cells differ from its normal counterpart. We have shown that tumour endothelial cells (TECs) isolated from tumour tissues are also abnormal. Furthermore, we found that mRNAs of vascular endothelial growth factor-A (VEGF-A) and cyclooxygenase-2 (COX-2) were upregulated in TECs. Vascular endothelial growth factor-A and COX-2 are angiogenic factors and their mRNAs contain an AU-rich element (ARE). AU-rich element-containing mRNAs are reportedly stabilised by Hu antigen R (HuR), which is exported to the cytoplasm. METHODS: Normal endothelial cell (NEC) and two types of TECs were isolated. We evaluated the correlation of HuR and accumulation of VEGF-A and COX-2 mRNAs in TECs and effects of HuR on biological phenotypes of TECs. RESULTS: The HuR protein was accumulated in the cytoplasm of TECs, but not in NECs. Vascular endothelial growth factor-A and COX-2 mRNA levels decreased due to HuR knockdown and it was shown that these ARE-mRNA were bound to HuR in TECs. Furthermore, HuR knockdown inhibited cell survival, random motility, tube formation, and Akt phosphorylation in TECs. CONCLUSION: Hu antigen R is associated with the upregulation of VEGF-A and COX-2 mRNA in TECs, and has an important role in keeping an angiogenic switch on, through activating angiogenic phenotype in tumour endothelium.
Assuntos
Antígenos de Superfície/metabolismo , Antígenos de Superfície/farmacologia , Ciclo-Oxigenase 2/genética , Células Endoteliais/metabolismo , Neoplasias/irrigação sanguínea , Proteínas de Ligação a RNA/metabolismo , Proteínas de Ligação a RNA/farmacologia , Fator A de Crescimento do Endotélio Vascular/genética , Antígenos de Superfície/genética , Linhagem Celular Tumoral , Movimento Celular , Sobrevivência Celular , Ciclo-Oxigenase 2/metabolismo , Proteínas ELAV , Proteína Semelhante a ELAV 1 , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Melanoma/irrigação sanguínea , Neoplasias Bucais/irrigação sanguínea , Fosforilação , RNA Mensageiro , Proteínas de Ligação a RNA/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
HuR, a ubiquitously expressed member of the Hu protein family that binds and stabilizes an AU-rich element (ARE)-containing mRNAs, is known to shuttle between the nucleus and the cytoplasm via several export pathways. When normal cells were treated with heat shock, HuR was exported to the cytoplasm in a chromosome maintenance region 1 (CRM1)-dependent manner. However, in this study, we demonstrate that HuR is exported to the cytoplasm in oral cancer cells even if the cells were treated with the inhibitor of the CRM1-independent export pathway. Immunohistochemical and biochemical analyses showed that HuR existed in both the cytoplasm and the nucleus in oral cancer cells, such as HSC-3 and Ca9.22, but existed entirely inside the nucleus in normal cells. AU-rich element-mRNAs were also exported to the cytoplasm and stabilised in the oral cancer cells, which were inhibited by HuR knockdown. This export of HuR was not affected by at least 7 h of treatment of leptomycin B (LMB), which is an inhibitor of the CRM1-dependent export pathway. These findings suggest that HuR is exported to the cytoplasm in oral carcinoma cells in a different manner from that of normal cells, and is likely to occur through the perturbation of a normal export pathway.