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Thromb Haemost 2021; 121(07): 877-890
DOI: 10.1055/s-0040-1722225
Coagulation and Fibrinolysis

NETosis Markers in Pregnancy: Effects Differ According to Histone Subtypes

Sylvie Bouvier    *
1   Department of Haematology, University Hospital, Nîmes, France
2   Research Laboratory EA 2992, Montpellier University, Montpellier, France
3   Faculty of Pharmaceutical and Biological Sciences, Montpellier University, Montpellier, France
,
Mathieu Fortier‡   *
1   Department of Haematology, University Hospital, Nîmes, France
,
Laura Vincent
1   Department of Haematology, University Hospital, Nîmes, France
,
Christophe Demattei
4   Department of Biostatistics, Public Health and Innovation in Methodology, Nîmes University Hospital, Nîmes, France
,
Eve Mousty
5   Department of Gynecology and Obstetrics, University Hospital, Nîmes, France
,
Marielle Herzog
6   Belgian Volition SPRL, Parc Scientifique Crealys, Isnes, Belgium
,
Guillaume Rommelaere
6   Belgian Volition SPRL, Parc Scientifique Crealys, Isnes, Belgium
,
Eva Nouvellon
1   Department of Haematology, University Hospital, Nîmes, France
2   Research Laboratory EA 2992, Montpellier University, Montpellier, France
,
Eric Mercier
1   Department of Haematology, University Hospital, Nîmes, France
2   Research Laboratory EA 2992, Montpellier University, Montpellier, France
3   Faculty of Pharmaceutical and Biological Sciences, Montpellier University, Montpellier, France
,
Vincent Letouzey
5   Department of Gynecology and Obstetrics, University Hospital, Nîmes, France
7   Department of Artificial Polymers, Max Mousseron Institute of Biomolecules, Montpellier University, Montpellier, France
,
Jean-Christophe Gris 
1   Department of Haematology, University Hospital, Nîmes, France
2   Research Laboratory EA 2992, Montpellier University, Montpellier, France
3   Faculty of Pharmaceutical and Biological Sciences, Montpellier University, Montpellier, France
8   Department of Gynecology, I. M. Sechenov First Moscow State Medical University, Moscow, Russian Federation
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Abstract

NETosis is an innate immune response occurring after infection or inflammation: activated neutrophils expel decondensed DNA in complex with histones into the extracellular environment in a controlled manner. It activates coagulation and fuels the risk of thrombosis. Human pregnancy is associated with a mild proinflammatory state characterized by circulatory neutrophil activation which is further increased in complicated pregnancies, placenta-mediated complications being associated with an increased thrombotic risk. This aberrant activation leads to an increased release of nucleosomes in the blood flow. The aim of our study was to initially quantify nucleosome-bound histones in normal pregnancy and in placenta-mediated complication counterpart. We analyzed the role of histones on extravillous trophoblast function. Circulating nucleosome-bound histones H3 (Nu.QH3.1, Nu.QH3PanCit, Nu.QH3K27me3) and H4 (Nu.QH4K16Ac) were increased in complicated pregnancies. In vitro using the extravillous cell line HTR-8/SVNeo, we observed that free recombinant H2B, H3, and H4 inhibited migration in wound healing assay, but only H3 also blocked invasion in Matrigel-coated Transwell experiments. H3 and H4 also induced apoptosis, whereas H2B did not. Finally, the negative effects of H3 on invasion and apoptosis could be restored with enoxaparin, a low-molecular-weight heparin (LMWH), but not with aspirin. Different circulating nucleosome-bound histones are increased in complicated pregnancy and this would affect migration, invasion, and induce apoptosis of extravillous trophoblasts. Histones might be part of the link between the risk of thrombosis and pregnancy complications, with an effect of LMWH on both.

Keywords

histones - NETosis - pregnancy - preeclampsia - trophoblast

Authors' Contributions

S.B. designed the research, analyzed the data, and wrote the manuscript. M.F. performed the research, analyzed the data, and wrote the manuscript. L.V. performed the research. C.D. analyzed the data. E.Mo. and V.L. included participants in the main study. M.H and G.R. performed the ELISA tests. E.N. and E.Me analyzed the data. J.-C.G. designed the research and analyzed the data.


* These authors contributed equally in this work.


Supplementary Material



Publication History

Received: 30 June 2020

Accepted: 18 November 2020

Article published online:
10 January 2021

© 2021. Thieme. All rights reserved.

Georg Thieme Verlag KG
Rüdigerstraße 14, 70469 Stuttgart, Germany

 

  • References

  • 1 Lane-Cordova AD, Khan SS, Grobman WA, Greenland P, Shah SJ. Long-term cardiovascular risks associated with adverse pregnancy outcomes: JACC review topic of the week. J Am Coll Cardiol 2019; 73 (16) 2106-2116
    CrossrefPubMedGoogle Scholar
  • 2 Sattar N, Greer IA. Pregnancy complications and maternal cardiovascular risk: opportunities for intervention and screening?. BMJ 2002; 325 (7356): 157-160
    CrossrefPubMedGoogle Scholar
  • 3 Bellamy L, Casas JP, Hingorani AD, Williams DJ. Pre-eclampsia and risk of cardiovascular disease and cancer in later life: systematic review and meta-analysis. BMJ 2007; 335 (7627): 974
    CrossrefPubMedGoogle Scholar
  • 4 Bouvier S, Mousty E, Fortier M. et al. Placenta-mediated complications: nucleosomes and free DNA concentrations differ depending on subtypes. J Thromb Haemost 2020; DOI: 10.1111/jth.15105.
    CrossrefPubMedGoogle Scholar
  • 5 Zhong XY, Laivuori H, Livingston JC. et al. Elevation of both maternal and fetal extracellular circulating deoxyribonucleic acid concentrations in the plasma of pregnant women with preeclampsia. Am J Obstet Gynecol 2001; 184 (03) 414-419
    CrossrefPubMedGoogle Scholar
  • 6 Hu Y, Li H, Yan R. et al. Increased neutrophil activation and plasma dna levels in patients with pre-eclampsia. Thromb Haemost 2018; 118 (12) 2064-2073
    Article in Thieme ConnectPubMedGoogle Scholar
  • 7 Massberg S, Grahl L, von Bruehl ML. et al. Reciprocal coupling of coagulation and innate immunity via neutrophil serine proteases. Nat Med 2010; 16 (08) 887-896
    CrossrefPubMedGoogle Scholar
  • 8 Hahn S, Giaglis S, Hoesli I, Hasler P. Neutrophil NETs in reproduction: from infertility to preeclampsia and the possibility of fetal loss. Front Immunol 2012; 3: 362
    CrossrefPubMedGoogle Scholar
  • 9 Hahn S, Hasler P, Vokalova L. et al. The role of neutrophil activation in determining the outcome of pregnancy and modulation by hormones and/or cytokines. Clin Exp Immunol 2019; 198 (01) 24-36
    CrossrefPubMedGoogle Scholar
  • 10 Silk E, Zhao H, Weng H, Ma D. The role of extracellular histone in organ injury. Cell Death Dis 2017; 8 (05) e2812
    CrossrefPubMedGoogle Scholar
  • 11 Yang H, Antoine DJ, Andersson U, Tracey KJ. The many faces of HMGB1: molecular structure-functional activity in inflammation, apoptosis, and chemotaxis. J Leukoc Biol 2013; 93 (06) 865-873
    CrossrefPubMedGoogle Scholar
  • 12 Hu Y, Yan R, Zhang C. et al. High-mobility group box 1 from hypoxic trophoblasts promotes endothelial microparticle production and thrombophilia in preeclampsia. Arterioscler Thromb Vasc Biol 2018; 38 (06) 1381-1391
    CrossrefPubMedGoogle Scholar
  • 13 Thålin C, Daleskog M, Göransson SP. et al. Validation of an enzyme-linked immunosorbent assay for the quantification of citrullinated histone H3 as a marker for neutrophil extracellular traps in human plasma. Immunol Res 2017; 65 (03) 706-712
    CrossrefPubMedGoogle Scholar
  • 14 Leshner M, Wang S, Lewis C. et al. PAD4 mediated histone hypercitrullination induces heterochromatin decondensation and chromatin unfolding to form neutrophil extracellular trap-like structures. Front Immunol 2012; 3: 307
    CrossrefPubMedGoogle Scholar
  • 15 Bauden M, Pamart D, Ansari D. et al. Circulating nucleosomes as epigenetic biomarkers in pancreatic cancer. Clin Epigenetics 2015; 7: 106
    CrossrefPubMedGoogle Scholar
  • 16 Huppertz B, Peeters LL. Vascular biology in implantation and placentation. Angiogenesis 2005; 8 (02) 157-167
    CrossrefPubMedGoogle Scholar
  • 17 E Davies J, Pollheimer J, Yong HE. et al. Epithelial-mesenchymal transition during extravillous trophoblast differentiation. Cell Adhes Migr 2016; 10 (03) 310-321
    CrossrefPubMedGoogle Scholar
  • 18 Sibai B, Dekker G, Kupferminc M. Pre-eclampsia. Lancet 2005; 365 (9461): 785-799
    CrossrefPubMedGoogle Scholar
  • 19 Redman CW, Sargent IL. Latest advances in understanding preeclampsia. Science 2005; 308 (5728): 1592-1594
    CrossrefPubMedGoogle Scholar
  • 20 Demers M, Wagner DD. Neutrophil extracellular traps: a new link to cancer-associated thrombosis and potential implications for tumor progression. OncoImmunology 2013; 2 (02) e22946
    CrossrefPubMedGoogle Scholar
  • 21 Mena HA, Carestia A, Scotti L, Parborell F, Schattner M, Negrotto S. Extracellular histones reduce survival and angiogenic responses of late outgrowth progenitor and mature endothelial cells. J Thromb Haemost 2016; 14 (02) 397-410
    CrossrefPubMedGoogle Scholar
  • 22 Brown MA, Magee LA, Kenny LC. et al; International Society for the Study of Hypertension in Pregnancy (ISSHP). The hypertensive disorders of pregnancy: ISSHP classification, diagnosis & management recommendations for international practice. Pregnancy Hypertens 2018; 13: 291-310
    CrossrefPubMedGoogle Scholar
  • 23 ACOG Practice Bulletin No. ACOG Practice Bulletin No. 202: gestational hypertension and preeclampsia. Obstet Gynecol 2019; 133 (01) e1-e25
    CrossrefPubMedGoogle Scholar
  • 24 Gordijn SJ, Beune IM, Thilaganathan B. et al. Consensus definition of fetal growth restriction: a Delphi procedure. Ultrasound Obstet Gynecol 2016; 48 (03) 333-339
    CrossrefPubMedGoogle Scholar
  • 25 Price CP, Newall RG, Boyd JC. Use of protein:creatinine ratio measurements on random urine samples for prediction of significant proteinuria: a systematic review. Clin Chem 2005; 51 (09) 1577-1586
    CrossrefPubMedGoogle Scholar
  • 26 Papayannopoulos V, Metzler KD, Hakkim A, Zychlinsky A. Neutrophil elastase and myeloperoxidase regulate the formation of neutrophil extracellular traps. J Cell Biol 2010; 191 (03) 677-691
    CrossrefPubMedGoogle Scholar
  • 27 Wang F, Zhang N, Li B. et al. Heparin defends against the toxicity of circulating histones in sepsis. Front Biosci 2015; 20: 1259-1270
    CrossrefPubMedGoogle Scholar
  • 28 Giaglis S, Stoikou M, Sur Chowdhury C. et al. Multimodal regulation of NET formation in pregnancy: progesterone antagonizes the Pro-NETotic effect of estrogen and G-CSF. Front Immunol 2016; 7: 565
    CrossrefPubMedGoogle Scholar
  • 29 Mizugishi K, Yamashita K. Neutrophil extracellular traps are critical for pregnancy loss in sphingosine kinase-deficient mice on 129Sv/C57BL/6 background. FASEB J 2017; 31 (12) 5577-5591
    CrossrefPubMedGoogle Scholar
  • 30 Young CH, Rothfuss HM, Gard PF. et al. Citrullination regulates the expression of insulin-like growth factor-binding protein 1 (IGFBP1) in ovine uterine luminal epithelial cells. Reproduction 2017; 153 (01) 1-10
    CrossrefPubMedGoogle Scholar
  • 31 Nancy P, Siewiera J, Rizzuto G. et al. H3K27me3 dynamics dictate evolving uterine states in pregnancy and parturition. J Clin Invest 2018; 128 (01) 233-247
    CrossrefPubMedGoogle Scholar
  • 32 Rahat B, Sharma R, Bagga R, Hamid A, Kaur J. Imbalance between matrix metalloproteinases and their tissue inhibitors in preeclampsia and gestational trophoblastic diseases. Reproduction 2016; 152 (01) 11-22
    CrossrefPubMedGoogle Scholar
  • 33 Sacks GP, Studena K, Sargent K, Redman CW. Normal pregnancy and preeclampsia both produce inflammatory changes in peripheral blood leukocytes akin to those of sepsis. Am J Obstet Gynecol 1998; 179 (01) 80-86
    CrossrefPubMedGoogle Scholar
  • 34 Gupta A, Hasler P, Gebhardt S, Holzgreve W, Hahn S. Occurrence of neutrophil extracellular DNA traps (NETs) in pre-eclampsia: a link with elevated levels of cell-free DNA?. Ann N Y Acad Sci 2006; 1075: 118-122
    CrossrefPubMedGoogle Scholar
  • 35 Paules C, Youssef L, Rovira C. et al. Distinctive patterns of placental lesions in pre-eclampsia vs small-for-gestational age and their association with fetoplacental Doppler. Ultrasound Obstet Gynecol 2019; 54 (05) 609-616
    CrossrefPubMedGoogle Scholar
  • 36 De Pascalis C, Etienne-Manneville S. Single and collective cell migration: the mechanics of adhesions. Mol Biol Cell 2017; 28 (14) 1833-1846
    CrossrefPubMedGoogle Scholar
  • 37 Wildhagen KC, García de Frutos P, Reutelingsperger CP. et al. Nonanticoagulant heparin prevents histone-mediated cytotoxicity in vitro and improves survival in sepsis. Blood 2014; 123 (07) 1098-1101
    CrossrefPubMedGoogle Scholar
  • 38 Abrams ST, Zhang N, Dart C. et al. Human CRP defends against the toxicity of circulating histones. J Immunol 2013; 191 (05) 2495-2502
    CrossrefPubMedGoogle Scholar
  • 39 Kleine TJ, Lewis PN, Lewis SA. Histone-induced damage of a mammalian epithelium: the role of protein and membrane structure. Am J Physiol 1997; 273 (06) C1925-C1936
    CrossrefPubMedGoogle Scholar
  • 40 Hariton-Gazal E, Rosenbluh J, Graessmann A, Gilon C, Loyter A. Direct translocation of histone molecules across cell membranes. J Cell Sci 2003; 116 (Pt 22): 4577-4586
    CrossrefPubMedGoogle Scholar