Vaginal neutrophil infiltration is contingent on ovarian cycle phase and independent of pathogen infection.

The mucosa of the female reproductive tract must reconcile the presence of commensal microbiota and the transit of exogenous spermatozoa with the elimination of sexually transmitted pathogens. In the vagina, neutrophils are the principal cellular arm of innate immunity and constitute the first line of protection in response to infections or injury. Neutrophils are absent from the vaginal lumen during the ovulatory phase, probably to allow sperm to fertilize; however, the mechanisms that regulate neutrophil influx to the vagina in response to aggressions remain controversial. We have used mouse inseminations and infections of Neisseria gonorrhoeae, Candida albicans, Trichomonas vaginalis, and HSV-2 models. We demonstrate that neutrophil infiltration of the vaginal mucosa is distinctively contingent on the ovarian cycle phase and independent of the sperm and pathogen challenge, probably to prevent sperm from being attacked by neutrophils. Neutrophils extravasation is a multi-step cascade of events, which includes their adhesion through selectins (E, P and L) and integrins of the endothelial cells. We have discovered that cervical endothelial cells expressed selectin-E (SELE, CD62E) to favor neutrophils recruitment and estradiol down-regulated SELE expression during ovulation, which impaired neutrophil transendothelial migration and orchestrated sperm tolerance. Progesterone up-regulated SELE to restore surveillance after ovulation.

Vaginal neutrophils eliminate sperm by trogocytosis.

STUDY QUESTION: What is the vaginal polymorphonuclear (PMN) spermicidal mechanism to reduce the excess of sperm? SUMMARY ANSWER: We show that PMNs are very efficient at killing sperm by a trogocytosis-dependent spermicidal activity independent of neutrophil extracellular traps (NETs). WHAT IS KNOWN ALREADY: Trogocytosis has been described as an active membrane exchange between immune cells with a regulatory purpose. Recently, trogocytosis has been reported as a mechanism which PMNs use to kill tumour cells or Trichomonas vaginalis. STUDY DESIGN, SIZE, DURATION: We used in vivo murine models and human ex vivo sperm and PMNs to investigate the early PMN-sperm response. PARTICIPANTS/MATERIALS, SETTING, METHODS: We set up a live/dead sperm detection system in the presence of PMNs to investigate in vivo and ex vivo PMN-spermicidal activity by confocal microscopy, flow cytometry and computer-assisted sperm analysis (SCA). MAIN RESULTS AND THE ROLE OF CHANCE: We revealed that PMNs are highly efficient at killing sperm by way of a NETs-independent, contact-dependent and serine proteases-dependent engulfment mechanism. PMNs 'bite' sperm and quickly reduce sperm motility (within 5 min) and viability (within 20 min) after contact. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: This study was conducted using murine models and healthy human blood PMNs; whether it is relevant to human vaginal PMNs or to cases of infertility is unknown. WIDER IMPLICATIONS OF THE FINDINGS: Vaginal PMNs attack and immobilize excess sperm in the vagina by trogocytosis because sperm are exogenous and may carry pathogens. Furthermore, this mechanism of sperm regulation has low mucosal impact and avoids an exacerbated inflammatory response that could lead to mucosal damage or infertility. STUDY FUNDING/COMPETING INTEREST(S): This work was partially supported by Ministry of Economy and Competitiveness ISCIII-FIS grants, PI16/00050, and PI19/00078, co-financed by ERDF (FEDER) Funds from the European Commission, 'A way of making Europe' and IiSGM intramural grant II-PI-MRC-2017. M.R. holds a Miguel Servet II contract (CPII14/00009). M.C.L. holds IiSGM intramural contract. There are no competing interests.

Estradiol impairs epithelial CXCL1 gradient in the cervix to delay neutrophil transepithelial migration during insemination.

Female reproductive mucosa must allow allogenic sperm survival whereas at the same time, avoid pathogen infection. To preserve sperm from neutrophil attack, neutrophils disappear from the vagina during the ovulatory phase (high estradiol); although the mechanisms that regulate neutrophil influx to the vagina during insemination remain controversial. We investigated the sex hormone regulation of the neutrophil migration through the cervix during insemination and revealed that ovulatory estradiol dose fades the CXCL1 epithelial expression in the ectocervix and fornix; hence, retarding neutrophil migration and retaining them in the epithelium. These mechanisms spare sperm from neutrophil attack to preserve reproduction, but might compromise immunity. However, luteal progesterone dose promotes the CXCL1 gradient expression to restore neutrophil migration, to eliminate sperm and prevent sperm associated pathogen dissemination. Surprisingly, these mechanisms are hormone dependent and independent of the insemination. Thus, sex hormones orchestrate tolerance and immunity in the vaginal lumen by regulating neutrophil transepithelial migration in the fornix and ectocervix.