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Thromb Haemost 2021; 121(11): 1483-1496
DOI: 10.1055/a-1382-9983
Stroke, Systemic or Venous Thromboembolism

Estradiol-Responsive miR-365a-3p Interacts with Tissue Factor 3′UTR to Modulate Tissue Factor-Initiated Thrombin Generation

Jiayin Tian
1   Western Australian Centre for Thrombosis and Haemostasis, Murdoch University, Murdoch, Perth, Australia
2   Perth Blood Institute, West Perth, Perth, Australia
3   College of Science, Health, Engineering and Education, Murdoch University, Murdoch, Perth, Australia
4   Centre for Molecular Medicine and Innovative Therapeutics, Health Futures Institute, Murdoch University, Murdoch, Perth, Australia
,
Murray J. Adams
3   College of Science, Health, Engineering and Education, Murdoch University, Murdoch, Perth, Australia
,
Jasmine Wee Ting Tay   *
1   Western Australian Centre for Thrombosis and Haemostasis, Murdoch University, Murdoch, Perth, Australia
2   Perth Blood Institute, West Perth, Perth, Australia
,
Ian James
5   Institute for Immunology and Infectious Diseases, Murdoch University, Murdoch, Perth, Australia
,
Suzanne Powell
1   Western Australian Centre for Thrombosis and Haemostasis, Murdoch University, Murdoch, Perth, Australia
,
Quintin W. Hughes   *
1   Western Australian Centre for Thrombosis and Haemostasis, Murdoch University, Murdoch, Perth, Australia
2   Perth Blood Institute, West Perth, Perth, Australia
,
Grace Gilmore
1   Western Australian Centre for Thrombosis and Haemostasis, Murdoch University, Murdoch, Perth, Australia
2   Perth Blood Institute, West Perth, Perth, Australia
4   Centre for Molecular Medicine and Innovative Therapeutics, Health Futures Institute, Murdoch University, Murdoch, Perth, Australia
,
Ross I. Baker
1   Western Australian Centre for Thrombosis and Haemostasis, Murdoch University, Murdoch, Perth, Australia
2   Perth Blood Institute, West Perth, Perth, Australia
4   Centre for Molecular Medicine and Innovative Therapeutics, Health Futures Institute, Murdoch University, Murdoch, Perth, Australia
,
Jim Yu-Hsiang Tiao
1   Western Australian Centre for Thrombosis and Haemostasis, Murdoch University, Murdoch, Perth, Australia
2   Perth Blood Institute, West Perth, Perth, Australia
4   Centre for Molecular Medicine and Innovative Therapeutics, Health Futures Institute, Murdoch University, Murdoch, Perth, Australia
› Author Affiliations Funding This study was supported by Murdoch University Small Grants and Perth Blood Institute.
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Abstract

Background High estradiol (E2) levels are linked to an increased risk of venous thromboembolism; however, the underlying molecular mechanism(s) remain poorly understood. We previously identified an E2-responsive microRNA (miR), miR-494–3p, that downregulates protein S expression, and posited additional coagulation factors, such as tissue factor, may be regulated in a similar manner via miRs.

Objectives To evaluate the coagulation capacity of cohorts with high physiological E2, and to further characterize novel E2-responsive miR and miR regulation on tissue factor in E2-related hypercoagulability.

Methods Ceveron Alpha thrombin generation assay (TGA) was used to assess plasma coagulation profile of three cohorts. The effect of physiological levels of E2, 10 nM, on miR expression in HuH-7 cells was compared using NanoString nCounter and validated with independent assays. The effect of tissue factor-interacting miR was confirmed by dual-luciferase reporter assays, immunoblotting, flow cytometry, biochemistry assays, and TGA.

Results Plasma samples from pregnant women and women on the contraceptive pill were confirmed to be hypercoagulable (compared with sex-matched controls). At equivalent and high physiological levels of E2, miR-365a-3p displayed concordant E2 downregulation in two independent miR quantification platforms, and tissue factor protein was upregulated by E2 treatment. Direct interaction between miR-365a-3p and F3-3′UTR was confirmed and overexpression of miR-365a-3p led to a decrease of (1) tissue factor mRNA transcripts, (2) protein levels, (3) activity, and (4) tissue factor-initiated thrombin generation.

Conclusion miR-365a-3p is a novel tissue factor regulator. High E2 concentrations induce a hypercoagulable state via a miR network specific for coagulation factors.

Keywords

estrogen - microRNA - thrombin generation - thrombosis - tissue factor

Author Contributions

J.T.: Drafted and edited the manuscript, performed the experiments, and analyzed the data. M.J.A.: Conceptualized, evaluated the data, edited and revised the manuscript. J.W.T.T.: Conceptualized, drafted the method sections of the manuscript, performed the experiments, and analyzed the data. I.J.: Performed statistical analyses on NanoString nCounter data and evaluated the data. S.P.: Performed the experiments and evaluated the data. Q.W.H.: Conceptualized, evaluated the data, and edited the manuscript. G.G.: Performed the experiments and evaluated the data. R.I.B.: Conceptualized, evaluated the data, and edited the manuscript. J.Y-H.T.: Conceptualized the experiments, evaluated the data, edited and revised the manuscript.


* Current affiliation: PathWest Laboratory Medicine, WA, Australia.


Supplementary Material



Publication History

Received: 14 July 2020

Accepted: 02 February 2021

Accepted Manuscript online:
04 February 2021

Article published online:
22 April 2021

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