Dataset Information

Hypoxia-inducible factor-1alpha and vascular endothelial growth factor expression in circulating tumor cells of breast cancer patients.

ABSTRACT:

Introduction

The detection of peripheral blood circulating tumor cells (CTCs) and bone marrow disseminated tumor cells (DTCs) in breast cancer patients is associated with a high incidence of disease relapse and disease-related death. Since hypoxia-inducible factor-1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) play an important role in angiogenesis and tumor progression, the purpose of the current study was to investigate their expression in CTCs.

Methods

The expression of cytokeratins (CK), VEGF, vascular endothelial growth factor receptor-2 (VEGF2), HIF-1alpha and phosphorylated-focal adhesion kinase (pFAK) in CTCs from 34 patients with metastatic breast cancer who had detectable CK-19 mRNA-positive CTCs was assessed using double staining experiments and confocal laser scanning microscopy. Peripheral blood mononuclear cells (PBMCs) were stained with a monoclonal A45-B/B3 pancytokeratin antibody in combination with either VEGF or VEGFR2 or HIF-1alpha or pFAK antibodies, respectively.

Results

pFAK expression in circulating tumor cells was detected in 92% of patients whereas expression of VEGF, VEGF2 and HIF-1alpha was observed in 62%, 47% and 76% of patients, respectively. VEGF, VEGF2, HIF-1alpha and pFAK were expressed in 73%, 71%, 56% and 81%, respectively, of all the detected CTCs. Vascular endothelial growth mRNA was also detected by quantitative real-time RT-PCR in immunomagnetically-separated CTCs. Double and triple staining experiments in cytospins of immunomagnetically-isolated CTCs showed that VEGF co-expressed with HIF-1alpha and VEGF2.

Conclusions

The expression of pFAK, HIF-1alpha, VEGF and VEGF2 in CTCs of patients with metastatic breast cancer could explain the metastatic potential of these cells and may provide a therapeutic target for their elimination.

SUBMITTER: Kallergi G

PROVIDER: S-EPMC2815547 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Publications

Hypoxia-inducible factor-1alpha and vascular endothelial growth factor expression in circulating tumor cells of breast cancer patients.

Kallergi Galatea G Markomanolaki Harris H Giannoukaraki Vicky V Papadaki Maria A MA Strati Areti A Lianidou Evi S ES Georgoulias Vassilis V Mavroudis Dimitris D Agelaki Sofia S

Breast cancer research : BCR 20091117 6

<h4>Introduction</h4>The detection of peripheral blood circulating tumor cells (CTCs) and bone marrow disseminated tumor cells (DTCs) in breast cancer patients is associated with a high incidence of disease relapse and disease-related death. Since hypoxia-inducible factor-1alpha (HIF-1alpha) and vascular endothelial growth factor (VEGF) play an important role in angiogenesis and tumor progression, the purpose of the current study was to investigate their expression in CTCs.<h4>Methods</h4>The ex ...[more]

PMID: 19919679

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Project description:Hypoxia-inducible factor-1 (HIF-1) is an important transcriptional factor in mammalian cells for coordination of adaptive responses to hypoxia. It consists of a regulatory subunit HIF-1alpha, which accumulates under hypoxic conditions, and a constitutively expressed subunit HIF-1beta. In addition to the well characterized oxygen-dependent mode of action of HIF-1, recent work has shown that various growth factors and cytokines stimulate HIF-1alpha expression, thereby triggering transcription of numerous hypoxia-inducible genes by oxygen-independent mechanisms. In this study, we examined whether accumulation of HIF-1alpha induced by insulin-like growth factor-1 (IGF-1) has a regulatory role in excitatory synaptic transmission in hippocampal neuron cultures. Our results show that IGF-1 induced a time- and dose-dependent increase in HIF-1alpha expression that was blocked by pretreatment with selective IGF-1 receptor antagonist, transcriptional inhibitor, and translational inhibitors. In addition, pharmacological blockade of the phosphatidylinositol 3-kinase/Akt/mammalian target of the rapamycin signaling pathway, but not extracellular signal-regulated kinase, inhibited IGF-1-induced HIF-1alpha expression. More importantly, the increase in HIF-1alpha expression induced by IGF-1 was accompanied by increasing levels of vascular endothelial growth factor (VEGF) mRNA and protein, which enhanced excitatory synaptic transmission. In parallel, blockade of HIF-1alpha activity by echinomycin or lentiviral infection with dominant-negative mutant HIF-1alpha or short hairpin RNA targeting HIF-1alpha inhibited the increase in expression of VEGF and the enhancement of synaptic transmission induced by IGF-1. Conversely, transfection of constitutively active HIF-1alpha into neurons mimicked the effects of IGF-1 treatment. Together, these results suggest that HIF-1alpha accumulation can enhance excitatory synaptic transmission in hippocampal neurons by regulating production of VEGF.

Project description:Vascular endothelial growth factor, a critical factor in angiogenesis, mediates stem cell paracrine protective effects on ischemic myocardium. Studies on the role of sex in stem cell function have demonstrated that female mesenchymal stem cells produce greater vascular endothelial growth factor and provide better cardiac protection compared with male mesenchymal stem cells. The purpose of this study was to determine the mechanisms by which estrogen affects mesenchymal stem cell function as a potential therapeutic measure during ex vivo expansion, before therapeutic use.A single-step purification method using adhesion to cell culture plastic was adopted to isolate mesenchymal stem cells from wild-type, estrogen receptor-alpha knockout, estrogen receptor-beta knockout, and signal transducer and activator of transcription 3 knockout mice. Mesenchymal stem cells were treated with or without 17beta-estradiol, estrogen receptor-alpha agonist (propyl pyrazoletriol), and estrogen receptor-beta agonist (diarylpropionitrile).17beta-estradiol significantly increased mesenchymal stem cell vascular endothelial growth factor production in a dose-dependent manner. Both estrogen receptor-alpha and estrogen receptor-beta were expressed in mesenchymal stem cells. Administration of 17beta-estradiol or estrogen receptor-alpha agonist (not estrogen receptor-beta agonist) elevated mesenchymal stem cell vascular endothelial growth factor, hypoxia inducible factor-1alpha expression, and signal transducer and activator of transcription 3 activation. However, these effects were neutralized in estrogen receptor-alpha knockout mesenchymal stem cells, not estrogen receptor-beta knockout. Signal transducer and activator of transcription 3 knockout abolished estrogen receptor-alpha-induced hypoxia inducible factor-1alpha and subsequent vascular endothelial growth factor production.17beta-estradiol-induced vascular endothelial growth factor production from mesenchymal stem cells appears to be mediated through estrogen receptor-alpha-activated signal transducer and activator of transcription 3-mediated hypoxia inducible factor-1alpha expression.

Project description:BACKGROUND: Hypoxia-inducible factor 1 (HIF-1) is a master transcriptional regulator of genes regulating oxygen homeostasis. The HIF-1 protein is composed of two HIF-1alpha and HIF-1beta/aryl hydrocarbon receptor nuclear translocator (ARNT) subunits. The prognostic relevance of HIF-1alpha protein overexpression has been shown in breast cancer. The impact of HIF-1alpha alternative splice variant expression on breast cancer prognosis in terms of metastasis risk is not well known. METHODS: Using real-time quantitative reverse transcription PCR assays, we measured mRNA concentrations of total HIF-1alpha and 4 variants in breast tissue specimens in a series of 29 normal tissues or benign lesions (normal/benign) and 53 primary carcinomas. In breast cancers HIF-1alpha splice variant levels were compared to clinicopathological parameters including tumour microvessel density and metastasis-free survival. RESULTS: HIF-1alpha isoforms containing a three base pairs TAG insertion between exon 1 and exon 2 (designated HIF-1alphaTAG) and HIF-1alpha736 mRNAs were found expressed at higher levels in oestrogen receptor (OR)-negative carcinomas compared to normal/benign tissues (P = 0.009 and P = 0.004 respectively). In breast carcinoma specimens, lymph node status was significantly associated with HIF-1alphaTAG mRNA levels (P = 0.037). Significant statistical association was found between tumour grade and HIF-1alphaTAG (P = 0.048), and total HIF-1alpha (P = 0.048) mRNA levels. HIF-1alphaTAG mRNA levels were also inversely correlated with both oestrogen and progesterone receptor status (P = 0.005 and P = 0.033 respectively). Univariate analysis showed that high HIF-1alphaTAG mRNA levels correlated with shortened metastasis free survival (P = 0.01). CONCLUSIONS: Our results show for the first time that mRNA expression of a HIF-1alphaTAG splice variant reflects a stage of breast cancer progression and is associated with a worse prognosis.See commentary: http://www.biomedcentral.com/1741-7015/8/45.

Dataset Information

Hypoxia-inducible factor-1alpha and vascular endothelial growth factor expression in circulating tumor cells of breast cancer patients.

Introduction

Methods

Results

Conclusions

Publications

Hypoxia-inducible factor-1alpha and vascular endothelial growth factor expression in circulating tumor cells of breast cancer patients.

Similar Datasets

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