MicroRNAs (miRNAs) are small non-coding RNAs that regulate the expression of different genes, including genes involved in cancer progression. A functional link between hypoxia, a key feature of the tumor microenvironment, and miRNA expression has been documented. We investigated whether and how miR-20b can regulate the expression of vascular endothelial growth factor (VEGF) in MCF-7 breast cancer cells under normoxic and hypoxia-mimicking conditions (CoCl(2) exposure). Using immunoblotting, ELISA, and quantitative real-time PCR, we demonstrated that miR-20b decreased VEGF protein levels at 4 and 24 h following CoCl(2) treatment, and VEGF mRNA at 4 h of treatment. In addition, miR-20b reduced VEGF protein expression in untreated cells. Next, we investigated the molecular mechanism by which pre-miR-20b can affect VEGF transcription, focusing on hypoxia inducible factor 1 (HIF-1) and signal transducer and activator of transcription 3 (STAT3), transcriptional inducers of VEGF and putative targets of miR-20b. Downregulation of VEGF mRNA by miR-20b under a 4 h of CoCl(2) treatment was associated with reduced levels of nuclear HIF-1 alpha subunit and STAT3. Chromatin immunoprecipitation (ChIP) assays revealed that HIF-1 alpha, but not STAT3, was recruited to the VEGF promoter following the 4 h of CoCl(2) treatment. This effect was inhibited by transfection of cells with pre-miR-20b. In addition, using siRNA knockdown, we demonstrated that the presence of STAT3 is necessary for CoCl(2)-mediated HIF-1 alpha nuclear accumulation and recruitment on VEGF promoter. In summary, this report demonstrates, for the first time, that the VEGF expression in breast cancer cells is mediated by HIF-1 and STAT3 in a miR-20b-dependent manner.

The angiopoietin-like 4 (ANGPTL4) protein belongs to a superfamily of secreted proteins structurally related to factors modulating angiogenesis known as angiopoietins. At first, ANGPTL4 has been identified as an adipokine exclusively involved in lipid metabolism, because of its prevalent expression in liver and adipose tissue. This protein regulates lipid metabolism by inhibiting lipoprotein lipase (LPL) activity and stimulating lipolysis of white adipose tissue (WAT), resulting in increased levels of plasma triglycerides (TG) and fatty acids. Subsequently, ANGPTL4 has been shown to be involved in several nonmetabolic and metabolic conditions, both physiological and pathological, including angiogenesis and vascular permeability, cell differentiation, tumorigenesis, glucose homoeostasis, lipid metabolism, energy homeostasis, wound healing, inflammation, and redox regulation. The transcriptional regulation of ANGPTL4 can be modulated by several transcription factors, including PPARα, PPARβ/δ, PPARγ, and HIF-1α, and nutritional and hormonal conditions. Several studies showed that high levels of ANGPTL4 are associated with poor prognosis in patients with various solid tumors, suggesting an important role in cancer onset and progression, metastasis, and anoikis resistance. Here, we have discussed the potential role of ANGPTL4 in mediating the cross talk between metabolic syndromes, such as diabetes and obesity, and cancer through regulation of its expression by PPARs.

BackgroundRecent findings indicate that exosomes released from cancer cells contain microRNAs (miRNAs) that may be delivered to cells of tumor microenvironment.ResultsTo elucidate whether miRNAs secreted from chronic myelogenous leukemia cells (CML) are shuttled into endothelial cells thus affecting their phenotype, we first analysed miRNAs content in LAMA84 exosomes. Among the 124 miRNAs identified in LAMA84 exosomes, we focused our attention on miR-126 which was found to be over-overexpressed in exosomes compared with producing parental cells. Transfection of LAMA84 with Cy3-labelled miR-126 and co-culture of leukemia cells with endothelial cells (EC) confirmed that miR-126 is shuttled into HUVECs. The treatment of HUVECs with LAMA84 exosomes for 24 hours reduced CXCL12 and VCAM1 expression, both at the mRNA and protein level, and negatively modulated LAMA84 motility and cells adhesion. Transfection in HUVECs of miR-126 inhibitor reversed the decrease of CXCL12 and restored the motility and adhesion of LAMA84 cells while the over-expression of miR-126, showed opposite effects.ConclusionOur results show that the miR-126 shuttled by exosomes is biologically active in the target cells, and support the hypothesis that exosomal miRNAs have an important role in tumor-endothelial crosstalk occurring in the bone marrow microenvironment, potentially affecting disease progression.

miRNAs have the potential for better understanding of tumor biology, but could also provide clinical advancement in management and therapy of various malignancies. The possibility of miRNA detection in peripheral blood would allow an eager expansion of their application in various clinical settings for cancer. The applicability of miRNA expression profiles still needs to be defined.