Antimicrobial peptides (AMPs) are small proteins produced by epithelial surfaces and inflammatory cells, which have broad-spectrum antimicrobial and immunomodulatory activities. They are known to be important in a number of infectious and inflammatory conditions and have been shown to be present in a number of sites throughout the female reproductive tract. Inflammation and infection are associated with a number of complications of pregnancy including preterm labor, and AMPs may play a key role in maintaining and protecting pregnancy. The aim of this review is to describe the expression and function of AMPs in the pregnant female reproductive tract and their relation to preterm labor.

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hCAP18/LL-37 is the sole human cathelicidin; a family of host defence peptides with key roles in innate host defence. hCAP18/LL-37 is expressed primarily by neutrophils and epithelial cells, but its production and function in the lower genital tract is largely uncharacterised. Despite the significant roles for cathelicidin in multiple organs and inflammatory processes, its impact on infections that could compromise fertility and pregnancy is unknown. The aim of this study was to investigate cathelicidin production, regulation and function in the cervix. hCAP18/LL-37 was found to be present in cervicovaginal secretions collected from women in the first trimester of pregnancy and to be expressed at significantly higher levels in samples from women with alterations in vaginal bacterial flora characteristic of bacterial vaginosis. In endocervical epithelial cell lines, expression of the gene encoding hCAP18/LL-37 (CAMP) was not affected by TLR agonists, but was found to be up-regulated by both 1, 25 hydroxyvitamin D3 and 25 hydroxyvitamin D3. However, no association was found between serum levels of vitamin D and hCAP18/LL-37 concentrations in cervicovaginal secretions (n = 116). Exposure to synthetic LL-37 had a pro-inflammatory effect on endocervical epithelial cell lines, increasing secretion of inflammatory cytokine IL-8. Together these data demonstrate inducible expression of hCAP18/LL-37 in the female lower reproductive tract in vivo and suggest the capacity for this peptide to modulate host defence to infection in this system. Further investigation will elucidate the effects of hCAP18/LL-37 on the physiology and pathophysiology of labour, and may lead to strategies for the prevention of infection-associated preterm birth.

STUDY QUESTION What effects did treatment using hyaluronic acid (HA) binding/selection prior to ICSI have on clinical outcomes in the Hyaluronic Acid Binding sperm Selection (HABSelect) clinical trial? SUMMARY ANSWER Older women randomized to the trial’s experimental arm (selection of sperm bound to immobilized (solid-state) HA) had the same live birth rates as younger women, most likely a result of better avoidance of sperm with damaged DNA. WHAT IS KNOWN ALREADY Recent randomized controlled trials (RCTs) investigating the efficacy of HA-based sperm selection prior to ICSI, including HABSelect, have consistently reported reductions in the numbers of miscarriages among couples randomized to the intervention, suggesting a pathological sperm-mediated factor mitigated by prior HA-binding/selection. The mechanism of that protection is unknown. STUDY DESIGN, SIZE, DURATION The original HABSelect Phase 3 RCT ran from 2014 to 2017 and included 2752 couples from whom sperm samples used in control (ICSI) and intervention (Physiological IntraCytoplasmic Sperm Injection; PICSI) arms of the trial were stored frozen for later assessment of DNA quality (DNAq). The trial overlapped with its mechanistic arm, running from 2016 to 2018. PARTICIPANTS/MATERIALS, SETTING, METHODS As miscarriage reduction was a significant secondary outcome of the trial, samples (n = 1247) selected for the mechanistic analysis were deliberately enriched for miscarriage outcomes (n = 92 or 7.4%) from a total of 154 miscarriages (5.6%) among all (n = 2752) couples randomized by stratified random sampling. Values from fresh semen samples for sperm concentration (mml), percentage forward progressive motility and percentage HA-binding score (HBS) were obtained before being processed by differential density gradient centrifugation or (rarely) by swim-up on the day of treatment. Surplus sperm pellets were recovered, aliquoted and cryopreserved for later analysis of DNAq using slide-based Comet, TUNEL, acridine orange (AO) and the sperm chromatin dispersion (SCD) assays. Following their classification into normal and abnormal sample subcategories based on reference values for sperm concentration and motility, relationships with HBS and DNAq were examined by Spearman correlation, Student’s t-tests, Mann Whitney U tests, and logistic regression (univariable and multivariable). Parsimonious selection enabled the development of models for exploring and explaining data trends. Potential differences in future cumulative pregnancy rates relating to embryo quality were also explored. MAIN RESULTS AND THE ROLE OF CHANCE Results from the 1247 sperm samples assayed for HBS and/or DNAq, generated data that were considered in relation to standard physiological measures of (sperm) vitality and to treatment outcomes. All measures of HBS and DNAq discriminated normal from abnormal sperm samples (P < 0.001). SCD correlated negatively with the Comet (r = −0.165; P < 0.001) and TUNEL assays (r = −0.200; P < 0.001). HBS correlated negatively with AO (r = −0.211; P < 0.001), Comet (r = −0.127; P < 0.001) and TUNEL (r = −0.214; P < 0.001) and positively with SCD (r = 0.255; P < 0.001). A model for predicting live birth (and miscarriage) rates included treatment allocation (odds ratio: OR 2.167, 95% CI 1.084–4.464, P = 0.031), female age (OR 0.301, 95% CI 0.133–0.761, P = 0.013, per decade) and the AO assay (OR 0.79, 95% CI 0.60–1. 02.761, P = 0.073, per 10 points rise). A model predicting the expected rate of biochemical pregnancy included male age (OR 0.464, 95% CI 0.314–0.674, P < 0.001, per decade) and the SCD assay (OR 1.04, 95% CI 1.007–1.075, P = 0.018, per 10 point rise). A model for conversion from biochemical to clinical pregnancy did not retain any significant patient or assay variables. A model for post-injection fertilization rates included treatment allocation (OR 0.83, 95% CI 0.75–0.91, P < 0.001) and the Comet assay (OR 0.950, 95% CI 0.91–1.00, P = 0.041). LIMITATIONS, REASONS FOR CAUTION HABSelect was a prospective RCT and the mechanistic study group was drawn from its recruitment cohort for retrospective analysis, without the full benefit of randomization. The clinical and mechanistic aspects of the study were mutually exclusive in that measures of DNAq were obtained from residual samples and not from HA-selected versus unselected sperm. Models for fitting mechanistic with baseline and other clinical data were developed to compensate for variable DNAq data quality. HABSelect used a solid-state version of PICSI and we did not assess the efficacy of any liquid-state alternatives. PICSI reduced fertilization rates and did not improve the outlook for cumulative pregnancy rates. WIDER IMPLICATIONS OF THE FINDINGS Notwithstanding the interventional effect on fertilization rates and possibly blastocyst formation (neither of which influenced pregnancy rates), poor sperm DNAq, reflected by lower HBS, probably contributed to the depression of all gestational outcomes including live births, in the HABSelect trial. The interventional avoidance of defective sperm is the best explanation for the equalization in live birth rates among older couples randomized to the trial’s PICSI arm. As patients going forward for assisted conception cycles globally in future are likely to be dominated by an older demographic, HA-based selection of sperm for ICSI could be considered as part of their treatment plan. STUDY FUNDING/COMPETING INTEREST(S) The study was supported by the National Institute for Health Research (NIHR) EME (Efficacy and Mechanism Evaluation)-11-14-34. National Research Ethics Service approval 11/06/2013: 13/YH/0162. S.L. is CEO of ExamenLab Ltd (company number NI605309). TRIAL REGISTRATION NUMBER ISRCTN99214271.

Preterm birth, defined as delivery before 37 weeks of gestation, is the leading cause of neonatal mortality and morbidity. Infection and inflammation are frequent antecedents of spontaneous preterm birth. Cathelicidin, an antimicrobial host defence peptide, is induced by infection and inflammation and although expressed in the reproductive tract and fetal tissues, its role in the pathogenesis of spontaneous preterm birth is unknown. Here we demonstrate that cathelicidin expression is increased at RNA and protein level in the mouse uterus in a model of inflammation-induced labour, where ultrasound guided intrauterine injection of lipopolysaccharide (LPS) at E17 stimulates preterm delivery within 24 hours. Cathelicidin-deficient ( Camp −/− ) mice are less susceptible to preterm delivery than wild type mice following intrauterine injection of 1 μg of LPS, and this is accompanied by a decrease in circulating IL-6, an inflammatory mediator implicated in the onset of labour. We also show that the proportion of cathelicidin expressing cells in the myometrium is higher in samples obtained from women in labour at term than pre-labour. Together, these data suggest that cathelicidin has roles in mediating pro-inflammatory responses in a murine model of inflammation-induced labour, and in human term labour.

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(Abstracted from Hum Reprod 2022;37:1106–1125)With intracytoplasmic sperm injection (ICSI), many natural barriers that would normally prevent the entry of abnormal sperm are lost, increasing the importance of sperm DNA integrity for successful in vitro fertilization. There is currently no consensus on the optimal assay for measuring sperm DNA quality (DNAq), defined as any structural aspect of sperm chromatin that can compromise function if disrupted.

Background The fetal membranes contribute to the rise in pro-infl ammatory cytokines during parturition that play a role in the onset of labour. In pregnancy, dietary supplementation with ω-3 polyunsaturated fatty acids (PUFAs) can prolong gestation and decrease preterm birth. 1 However, mechanisms of these effects are unclear. There has been recent interest in anti-infl ammatory actions of ω-3 PUFAs in other tissues, but their effect on the fetal membranes is largely unstudied. Objective To determine whether eicosapentaenoic acid (EPA) and docosahexanoic acid (DHA) have an anti-infl ammatory effect on the fetal membranes. Method Amnion and chorion explants were obtained at elective caesarean section (n=6) and cultured with EPA 50 μM, DHA 50 μM or a combination of EPA and DHA for 24 h. Production of the infl ammatory cytokines IL-6, IL-8, IL-1 and MCP-1 was determined by enzyme-linked immunosorbent assay of culture media. Data was analyzed by analysis of variance. Results are expressed as median and interquartile range. Results In the amnion, treatment with a combination of EPA and DHA signifi cantly reduced production of IL-6 (11.75 (9.3-156.9) vs 265.2 (64.4-865.7) p<0.05), IL-8 (359.8 (241-2316) vs 7623 (4748-8761) p<0.05) and MCP-1 (213.4 (15.6-433.6) vs 1160 (186.9-2783) p<0.05) compared to control. Production of IL-1 was undetectable in both control and treated samples. In the chorion EPA and/or DHA treatment had no effect on IL-6, IL-8 or IL-1 production, but did signifi cantly reduce production of MCP-1 (448 (231.2-1311) vs 3601 (1724-5874) p<0.05). Conclusion ω-3 PUFAs have an anti-infl ammatory effect on fetal membranes. This may contribute to their ability to prolong gestation.