Poster Presentation ESA-SRB 2023 in conjunction with ENSA

Serum derived complement proteins play an important role in sperm-neutrophil binding (#360)

Sophie Warr 1 , Simon de Graaf 1 , Taylor Pini 2 , Jessica Rickard 1
  1. School of Life and Environmental Sciences, The University of Sydney, Sydney, NSW, Australia
  2. School of Veterinary Science, The University of Queensland, Gatton, QLD, Australia

Semen deposition in the female tract elicits an immune-mediated response, releasing an influx of polymorphonuclear leukocytes (PMNs)1. In addition to protecting the reproductive tract from pathogens, this response also acts to select an elite population of sperm, capable of achieving fertilisation2. It has been hypothesised that the reduced ability of frozen-thawed ram spermatozoa to traverse the cervix may be related to their increased susceptibility to neutrophil binding and phagocytosis3. As such in the present study, we examined the binding of PMNs to fresh and frozen-thawed ram spermatozoa in the presence of ewe serum. Heat-treated serum was also included to assess the role of complement proteins. PMNs isolated from the blood of Merino ewes (n=2) were incubated (37°C; 120mins) with fresh spermatozoa diluted in Tris-citrate-fructose supplemented with 15% egg yolk (FrEY) and frozen-thawed spermatozoa diluted in an egg yolk-based diluent (FtEY) either in the absence of serum, 7.5% (v/v) serum, or 7.5% (v/v) heat-treated serum.  Serum significantly increased sperm-neutrophil binding in FrEY and FtEY (84.85±1.6% and 80.23±2.1%, respectively) compared to that recorded in no serum (58.44±3.4% and 66.41±3.2%, respectively; p=0.025).  The presence of heat-treated serum significantly reduced binding in FrEY by 22.25% and FtEY by 23.56% compared to that recorded in serum.  In the absence of serum, FtEY recorded higher binding rates (66.41±3.2%) compared to FrEY (58.43±3.4%; p=0.025).  Results in the absence of serum suggest that cryopreservation may alter lectin-mediated PMN binding, potentially due to modified sperm-surface carbohydrates. The deactivation of complement in heat-treated serum reduced binding compared to untreated serum, suggesting that complement-mediated binding is a key sperm-neutrophil binding mechanism.  Further investigation is now required to explore factors which influence complement-mediated binding, such as seminal plasma, which could be used to suppress the immune response within the female environment, aiding the survival of frozen-thawed sperm in the cervix.

  1. Thompson LA, Barratt CL, Bolton AE, Cooke ID. The leukocytic reaction of the human uterine cervix. American journal of reproductive immunology. 1992 Sep;28(2):85-9.
  2. Hunter, R.H.F., 1981. Sperm transport and reservoirs in the pig oviduct in relation to the time of ovulation. Reproduction, 63(1), pp.109-117.
  3. Warr S, Pini T, de Graaf SP, Rickard JP. Molecular insights to the sperm–cervix interaction and the consequences for cryopreserved sperm. Biology of Reproduction. 2023 Feb;108(2):183-96.