Abstract
Purpose
Using a comprehensive flow cytometric panel, do endometrial immune profiles in adverse reproductive outcomes such as repeat implantation failure (RIF) and repeat pregnancy loss (RPL) differ from each other and male-factor controls?
Methods
Six-hundred and twelve patients had an endometrial biopsy to assess the immunophenotype. History on presentation was used to subdivide the population into recurrent implantation failure (RIF) [n = 178], recurrent pregnancy loss (RPL) [n = 155], primary infertility [n = 130] and secondary infertility [n = 114]. A control group was utilised for comparative purposes [n = 35] and lymphocyte subpopulations were described.
Results
Distinct lymphocyte percentage differences were noted across the populations. Relative to controls and RPL, patients with a history of RIF had significantly raised uterine NKs (53.2 vs 45.2 & 42.9%, p < 0.0001). All sub-fertile populations had increased percentage peripheral type NKs (p = 0.001), and exhibited increased CD69+ activation (p = 0.005), higher levels of B cells (p < 0.001), elevated CD4:CD8 ratio (p < 0.0001), lower T-regs (p = 0.034) and a higher proportion of Th1+ CD4s (p = 0.001). Patient aetiology confers some distinct findings, RPL; pNK, Bcells and CD4 elevated; RIF; uNK and CD56 raised while CD-8 and NK-T lowered.
Conclusions
Flow cytometric endometrial evaluation has the ability to provide a rapid and objective analysis of lymphocyte subpopulations. The findings show significant variations in cellular proportions of immune cells across the patient categories relative to control tissue. The cell types involved suggest that a potential differential pro-inflammatory bias may exist in patients with a history of adverse reproductive outcomes. Immunological assessment in appropriate populations may provide insight into the underlying aetiology of some cases of reproductive failure.
Similar content being viewed by others
Abbreviations
- PGT-A:
-
preimplantation genetic analysis-aneuploidy
- RPL:
-
recurrent pregnancy loss
- RM:
-
recurrent miscarriage
- RIF:
-
repeat implantation failure
- ART:
-
assisted reproductive technologies
- HRT:
-
hormone replacement therapy
- uNK:
-
uterine type natural killer cells
- pNK:
-
peripheral blood type natural killer cells
- CD:
-
cluster of differentiation
- Th1:
-
T helper type 1 (pro-inflammatory)
- Th2:
-
T helper type 2 (anti-inflammatory)
References
Lathi RB, Westphal LM, Milki AA. Aneuploidy in the miscarriages of infertile women and the potential benefit of preimplanation genetic diagnosis. Fertil Steril. 2008;89(2):353–7.
El Hachem H, et al. Recurrent pregnancy loss: current perspectives. Int J Womens Health. 2017;9:331–45.
Giakoumelou S, Wheelhouse N, Cuschieri K, Entrican G, Howie SEM, Horne AW. The role of infection in miscarriage. Hum Reprod Update. 2016;22(1):116–33.
Bozdag G, Aksan G, Esinler I, Yarali H. What is the role of office hysteroscopy in women with failed IVF cycles? Reprod BioMed Online. 2008;17(3):410–5.
Riccio L, et al. Immunology of endometriosis. Best Pract Res Clin Obstet Gynaecol. 2018;50:39–49.
Stern C, Chamley L. Antiphospholipid antibodies and coagulation defects in women with implantation failure after IVF and recurrent miscarriage. Reprod BioMed Online. 2006;13(1):29–37.
Di Simone N, et al. Antiphospholipid antibodies affect human endometrial angiogenesis: protective effect of a synthetic peptide (TIFI) mimicking the phospholipid binding site of beta(2) glycoprotein I. Am J Reprod Immunol. 2013;70(4):299–308.
Bourgain C, Devroey P. The endometrium in stimulated cycles for IVF. Hum Reprod Update. 2003;9(6):515–22.
Revel A. Defective endometrial receptivity. Fertil Steril. 2012;97(5):1028–32.
Kwak-Kim J, Bao S, Lee SK, Kim JW, Gilman-Sachs A. Immunological modes of pregnancy loss: inflammation, immune effectors, and stress. Am J Reprod Immunol. 2014;72(2):129–40.
Hill JA. Immunological contributions to recurrent pregnancy loss. Baillieres Clin Obstet Gynaecol. 1992;6(3):489–505.
Franasiak JM, Scott RT. Contribution of immunology to implantation failure of euploid embryos. Fertil Steril. 2017;107(6):1279–83.
Lucas ES, Dyer NP, Murakami K, Hou Lee Y, Chan YW, Grimaldi G, et al. Loss of endometrial plasticity in recurrent pregnancy loss. Stem Cells. 2016;34(2):346–56.
Steer CV, Campbell S, Tan SL, Crayford T, Mills C, Mason BA, et al. The use of transvaginal color flow imaging after in vitro fertilization to identify optimum uterine conditions before embryo transfer. Fertil Steril. 1992;57(2):372–6.
Diaz-Gimeno P, et al. A genomic diagnostic tool for human endometrial receptivity based on the transcriptomic signature. Fertil Steril. 2011;95(1):50–60. 60.e1–15
Moffett A, Shreeve N. Reply: first do no harm: continuing the uterine NK cell debate. Hum Reprod. 2016;31(1):218–9.
Maecker HT, McCoy JP, Nussenblatt R. Standardizing immunophenotyping for the human immunology project. Nat Rev Immunol. 2012;12(3):191–200.
van Mourik MS, Macklon NS, Heijnen CJ. Embryonic implantation: cytokines, adhesion molecules, and immune cells in establishing an implantation environment. J Leukoc Biol. 2009;85(1):4–19.
Savasi VM, Mandia L, Laoreti A, Cetin I. Maternal and fetal outcomes in oocyte donation pregnancies. Hum Reprod Update. 2016;22(5):620–33.
Tang AW, Alfirevic Z, Quenby S. Natural killer cells and pregnancy outcomes in women with recurrent miscarriage and infertility: a systematic review. Hum Reprod. 2011;26(8):1971–80.
Russell P, Sacks G, Tremellen K, Gee A. The distribution of immune cells and macrophages in the endometrium of women with recurrent reproductive failure. III: further observations and reference ranges. Pathology. 2013;45(4):393–401.
Laird, L., Li, Bulmer, <RCOG 2016 guidelines.pdf>. 2016.
Hanna J, Goldman-Wohl D, Hamani Y, Avraham I, Greenfield C, Natanson-Yaron S, et al. Decidual NK cells regulate key developmental processes at the human fetal-maternal interface. Nat Med. 2006;12(9):1065–74.
Loke YW, King A, Burrows TD. Decidua in human implantation. Hum Reprod. 1995;10(Suppl 2):14–21.
Lédée N, Petitbarat M, Chevrier L, Vitoux D, Vezmar K, Rahmati M, et al. The uterine immune profile may help women with repeated unexplained embryo implantation failure after in vitro fertilization. Am J Reprod Immunol. 2016;75(3):388–401.
Lachapelle MH, et al. Endometrial T, B, and NK cells in patients with recurrent spontaneous abortion. Altered profile and pregnancy outcome. J Immunol. 1996;156(10):4027–34.
Ledee N, et al. Specific and extensive endometrial deregulation is present before conception in IVF/ICSI repeated implantation failures (IF) or recurrent miscarriages. J Pathol. 2011;225(4):554–64.
Moffett A, Colucci F. Uterine NK cells: active regulators at the maternal-fetal interface. J Clin Invest. 2014;124(5):1872–9.
Quenby S, Farquharson R. Uterine natural killer cells, implantation failure and recurrent miscarriage. Reprod BioMed Online. 2006;13(1):24–8.
Vassiliadou N, Bulmer JN. Immunohistochemical evidence for increased numbers of ‘classic’ CD57+ natural killer cells in the endometrium of women suffering spontaneous early pregnancy loss. Hum Reprod. 1996;11(7):1569–74.
Alecsandru D, Garcia-Velasco JA. Why natural killer cells are not enough: a further understanding of killer immunoglobulin-like receptor and human leukocyte antigen. Fertil Steril. 2017;107(6):1273–8.
Laufer N, Simon A. Recurrent implantation failure: current update and clinical approach to an ongoing challenge. Fertil Steril. 2012;97(5):1019–20.
Medicine, T.P.C.o.t.A.S.f.R., Evaluation and treatment of recurrent pregnancy loss: a committee opinion. Fertility and Sterility, 2012. 98(5): p. 1103–1111.
ESHRE, <ESHRE RPL Guideline_28112017_FINAL.pdf>. 2017.
Burkhard SH, Mair F, Nussbaum K, Hasler S, Becher B. T cell contamination in flow cytometry gating approaches for analysis of innate lymphoid cells. PLoS One. 2014;9(4):e94196.
Dosiou C, Giudice LC. Natural killer cells in pregnancy and recurrent pregnancy loss: endocrine and immunologic perspectives. Endocr Rev. 2005;26(1):44–62.
Marci R, Gentili V, Bortolotti D, Lo Monte G, Caselli E, Bolzani S, et al. Presence of HHV-6A in endometrial epithelial cells from women with primary unexplained infertility. PLoS One. 2016;11(7):e0158304.
Coulam CB, Bilal M, Salazar Garcia MD, Katukurundage D, Elazzamy H, Fernandez EF, et al. Prevalence of HHV-6 in endometrium from women with recurrent implantation failure. Am J Reprod Immunol. 2018;80:e12862.
Siewiera J, el Costa H, Tabiasco J, Berrebi A, Cartron G, Bouteiller P, et al. Human cytomegalovirus infection elicits new decidual natural killer cell effector functions. PLoS Pathog. 2013;9(4):e1003257.
Fehniger TA, Cooper MA, Nuovo GJ, Cella M, Facchetti F, Colonna M, et al. CD56bright natural killer cells are present in human lymph nodes and are activated by T cell-derived IL-2: a potential new link between adaptive and innate immunity. Blood. 2003;101(8):3052–7.
Petitbarat M, Rahmati M, Sérazin V, Dubanchet S, Morvan C, Wainer R, et al. TWEAK appears as a modulator of endometrial IL-18 related cytotoxic activity of uterine natural killers. PLoS One. 2011;6(1):e14497.
Fukui A, Funamizu A, Yokota M, Yamada K, Nakamua R, Fukuhara R, et al. Uterine and circulating natural killer cells and their roles in women with recurrent pregnancy loss, implantation failure and preeclampsia. J Reprod Immunol. 2011;90(1):105–10.
Thiruchelvam U, Wingfield M, O'Farrelly C. Natural killer cells: key players in endometriosis. Am J Reprod Immunol. 2015;74(4):291–301.
Gleicher N. Expected advances in human fertility treatments and their likely translational consequences. J Transl Med. 2018;16(1):149.
Hviid MM, Macklon N. Immune modulation treatments—where is the evidence? Fertil Steril. 2017;107(6):1284–93.
Alecsandru D, Garcia-Velasco JA. Immune testing and treatment: still an open debate. Hum Reprod. 2015;30(8):1994.
Sacks G. Enough! Stop the arguments and get on with the science of natural killer cell testing. Hum Reprod. 2015;30(7):1526–31.
Ledee N, et al. Impact of prednisone in patients with repeated embryo implantation failures: beneficial or deleterious? J Reprod Immunol. 2018;127:11–5.
Potdar N, Gelbaya T, Nardo LG. Endometrial injury to overcome recurrent embryo implantation failure: a systematic review and meta-analysis. Reprod BioMed Online. 2012;25(6):561–71.
Gnainsky Y, Granot I, Aldo PB, Barash A, Or Y, Schechtman E, et al. Local injury of the endometrium induces an inflammatory response that promotes successful implantation. Fertil Steril. 2010;94(6):2030–6.
Nastri, C.O., et al., Endometrial injury in women undergoing assisted reproductive techniques. Cochrane Database Syst Rev, 2015(3): p. Cd009517.
Reljic M, et al. Endometrial injury, the quality of embryos, and blastocyst transfer are the most important prognostic factors for in vitro fertilization success after previous repeated unsuccessful attempts. J Assist Reprod Genet. 2017;34(6):775–9.
Acknowledgements
Thanks to the Sims IVF patients who contributed tissue samples to facilitate the analysis and the Staff of the Clinic for their support. Thanks also to Dr. Renate Van der Molen, UMC Radboud, for critical appraisal of the manuscript.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Informed consent
Informed consent was obtained from all individual participants included in the study.
Conflict of interest
The authors declare no conflict of interest is present.
Rights and permissions
About this article
Cite this article
Marron, K., Walsh, D. & Harrity, C. Detailed endometrial immune assessment of both normal and adverse reproductive outcome populations. J Assist Reprod Genet 36, 199–210 (2019). https://doi.org/10.1007/s10815-018-1300-8
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10815-018-1300-8