Oestrogen induced downregulation of TFPI expression is mediated by ERα.
BACKGROUND
Oestrogens influence the pathophysiology and development of hormone-sensitive cancers, such as breast cancer. Tissue factor pathway inhibitor (TFPI) is a serine protease inhibitor of the extrinsic coagulation pathway and has recently been associated with breast cancer cell development. Moreover, reduced TFPI levels have been reported in plasma of healthy post-menopausal women receiving hormone replacement therapy, indicating a possible link between oestrogen and TFPI. In our study, we aimed to examine the effects of oestrogen and oestrogen analogues on TFPI expression in breast cancer cells and to identify underlying mechanism(s).


METHODS
Oestrogen receptor alpha (ERα) positive MCF7 and negative MDA-MB-231 cells were treated with 17-β-oestradiol, 17-β-ethinyloestradiol, raloxifene and fulvestrant. TFPI mRNA and protein was measured using qRT-PCR and ELISA, respectively. Transient ERα knockdown was achieved using siRNA.


RESULTS
In ERα expressing MCF7 cells, but not in MDA-MB-231 cells, the TFPI mRNA and protein levels were significantly downregulated by more than 50% after four or six hours of incubation with 17-β-ethinyloestradiol and 17-β-oestradiol, respectively. Moreover, a significant increase in FXa generation was detected in response to oestrogens. Breast tissue ER antagonists, raloxifene and fulvestrant, did not affect TFPI mRNA, however, fulvestrant blocked oestrogen mediated reduction of TFPI mRNA. Transient knockdown of ERα abolished the oestrogenic effect on TFPI and co-treatment of MCF7 cells with the protein synthesis inhibitor cycloheximide and 17-β-oestradiol also led to reduction of TFPI mRNA.


CONCLUSIONS
Our data establish a direct and time dependent regulation of TFPI expression by oestrogens through the ERα at the transcriptional level.
Department of Haematology, Oslo University Hospital, Oslo, Norway; Research Institute of Internal Medicine, Oslo University Hospital, Oslo, Norway; Institute of Clinical Medicine, University of Oslo, Oslo, Norway.
Huda Omar
HO
Ali
Department of Haematology, Oslo University Hospital, Oslo, Norway; Research Institute of Internal Medicine, Oslo University Hospital, Oslo, Norway.
Benedicte
B
Stavik
Department of Haematology, Oslo University Hospital, Oslo, Norway; Research Institute of Internal Medicine, Oslo University Hospital, Oslo, Norway.
Elisabeth
E
Dørum
Department of Medical Genetics, Oslo University Hospital, Oslo, Norway.
Nina
N
Iversen
Department of Haematology, Oslo University Hospital, Oslo, Norway; Research Institute of Internal Medicine, Oslo University Hospital, Oslo, Norway; Institute of Clinical Medicine, University of Oslo, Oslo, Norway.
Per Morten
PM
Sandset
Department of Haematology, Oslo University Hospital, Oslo, Norway; Research Institute of Internal Medicine, Oslo University Hospital, Oslo, Norway. Electronic address: grethe.skretting@medisin.uio.no.
Grethe
G
Skretting
D000230:Adenocarcinoma / Q000235:genetics / Q000378:metabolism*; D014408:Biomarkers, Tumor / Q000097:blood / Q000235:genetics; D001943:Breast Neoplasms / Q000235:genetics / Q000378:metabolism*; D015536:Down-Regulation / Q000187:drug effects; D004958:Estradiol / Q000494:pharmacology*; D047628:Estrogen Receptor alpha / Q000235:genetics / Q000378:metabolism*; D004997:Ethinyl Estradiol / Q000494:pharmacology*; D005260:Female; D006801:Humans; D008074:Lipoproteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D061986:MCF-7 Cells; D013927:Thrombosis / Q000235:genetics / Q000378:metabolism*
D014408:Biomarkers, Tumor; D047628:Estrogen Receptor alpha; D008074:Lipoproteins; C051928:lipoprotein-associated coagulation inhibitor; D004997:Ethinyl Estradiol; D004958:Estradiol
ERα; FXa; TFPI; fulvestrant; oestrogen; raloxifene