Effect of hypoxia on tissue factor pathway inhibitor expression in breast cancer

X Y Cui; M Tinholt; B Stavik; A E A Dahm; S Kanse; Y Jin; S Seidl; K K Sahlberg; N Iversen; G Skretting; P M Sandset

doi:10.1111/jth.13206

Effect of hypoxia on tissue factor pathway inhibitor expression in breast cancer

J Thromb Haemost. 2016 Feb;14(2):387-96. doi: 10.1111/jth.13206. Epub 2016 Jan 30.

Authors

X Y Cui^{1

2

3

4}, M Tinholt^{1

3

4

5}, B Stavik^{1

3}, A E A Dahm^{2

4

6}, S Kanse^{1

3

7}, Y Jin⁸, S Seidl^{1

3

7}, K K Sahlberg^{9

10

11}, N Iversen⁵, G Skretting^{1

3}, P M Sandset^{1

3

4}

Affiliations

¹ Department of Hematology, Oslo University Hospital, Oslo, Norway.
² Department of Hematology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.
³ Research Institute of Internal Medicine, Oslo University Hospital, Oslo, Norway.
⁴ Institute of Clinical Medicine, University of Oslo, Oslo, Norway.
⁵ Department of Medical Genetics, Oslo University Hospital, Oslo, Norway.
⁶ Department of Hematology, Akershus University Hospital, Lørenskog, Norway.
⁷ Institute of Basal Medical Sciences, University of Oslo, Oslo, Norway.
⁸ Department of Biosciences, University of Oslo, Oslo, Norway.
⁹ Department of Research, Vestre Viken Hospital Trust, Drammen, Norway.
¹⁰ K. G. Jebsen Center for Breast Cancer Research, University of Oslo, Oslo, Norway.
¹¹ Oslo Breast Cancer Research Consortium (OSBREAC), Oslo, Norway.

PMID: 26598923
DOI: 10.1111/jth.13206

Abstract

ESSENTIALS: A hypoxic microenvironment is a common feature of tumors that may influence activation of coagulation. MCF-7 and SK-BR-3 breast cancer cells and breast cancer tissue samples were used. The results showed transcriptional repression of tissue factor pathway inhibitor expression in hypoxia. Hypoxia-inducible factor 1α may be a target for the therapy of cancer-related coagulation and thrombosis.

Background: Activation of coagulation is a common finding in patients with cancer, and is associated with an increased risk of venous thrombosis. As a hypoxic microenvironment is a common feature of solid tumors, we investigated the role of hypoxia in the regulation of tissue factor (TF) pathway inhibitor (TFPI) expression in breast cancer.

Objectives: To explore the transcriptional regulation of TFPI by hypoxia-inducible factor (HIF)-1α in breast cancer cells and their correlation in breast cancer tissues.

Methods and results: MCF-7 and SK-BR-3 breast cancer cells were cultured in 1% oxygen or treated with cobalt chloride (CoCl2 ) to mimic hypoxia. Time-dependent and dose-dependent downregulation of TFPI mRNA (quantitative RT-PCR) and of free TFPI protein (ELISA) were observed in hypoxia. Western blotting showed parallel increases in the levels of HIF-1α protein and TF. HIF-1α inhibitor abolished or attenuated the hypoxia-induced downregulation of TFPI. Luciferase reporter assay showed that both hypoxia and HIF-1α overexpression caused strong repression of TFPI promoter activity. Subsequent chromatin immunoprecipitation and mutagenesis analysis demonstrated a functional hypoxia response element within the TFPI promoter, located at -1065 to -1060 relative to the transcriptional start point. In breast cancer tissue samples, gene expression analyses showed a positive correlation between the mRNA expression of TFPI and that of HIF-1α.

Conclusions: This study demonstrates that HIF-1α is involved in the transcriptional regulation of the TFPI gene, and suggests that a hypoxic microenvironment inside a breast tumor may induce a procoagulant state in breast cancer patients.

Keywords: HIF-1α protein; blood coagulation; breast cancer; human; hypoxia; tissue factor pathway inhibitor.

MeSH terms

Adult
Aged
Binding Sites
Blood Coagulation*
Breast Neoplasms / genetics
Breast Neoplasms / metabolism*
Breast Neoplasms / pathology
Cell Hypoxia
Down-Regulation
Female
Gene Expression Regulation, Neoplastic
HEK293 Cells
Humans
Hypoxia-Inducible Factor 1, alpha Subunit / genetics
Hypoxia-Inducible Factor 1, alpha Subunit / metabolism
Lipoproteins / genetics
Lipoproteins / metabolism*
MCF-7 Cells
Middle Aged
Oxygen / metabolism*
Promoter Regions, Genetic
RNA, Messenger / genetics
RNA, Messenger / metabolism
Time Factors
Transcription, Genetic
Transfection
Tumor Microenvironment

Substances

HIF1A protein, human
Hypoxia-Inducible Factor 1, alpha Subunit
Lipoproteins
RNA, Messenger
lipoprotein-associated coagulation inhibitor
Oxygen