DNA integrity and stability are critical determinants of cell viability. This is especially true in the female germline, wherein DNA integrity underpins successful conception, embryonic development, pregnancy and the production of healthy offspring. However, DNA is not inert; rather, it is subject to assault from various environment factors resulting in chemical modification and/or strand breakage. If structural alterations result and are left unrepaired, they have the potential to cause mutations and propagate disease. In this regard, reduced genetic integrity of the female germline ranks among the leading causes of subfertility in humans. With an estimated 10% of couples in developed countries taking recourse to ART to achieve pregnancy, the need for ongoing research into the capacity of the oocyte to detect DNA damage and thereafter initiate cell cycle arrest, apoptosis or DNA repair is increasingly more pressing. This review documents our current knowledge of the quality control mechanisms utilised by the female germline to prevent and remediate DNA damage during their development from primordial follicles through to the formation of preimplantation embryos. The PubMed database was searched using the keywords: primordial follicle, primary follicle, secondary follicle, tertiary follicle, germinal vesical, MI, MII oocyte, zygote, preimplantation embryo, DNA repair, double-strand break and DNA damage. These keywords were combined with other phrases relevant to the topic. Literature was restricted to peer-reviewed original articles in the English language (published 1979–2018) and references within these articles were also searched. In this review, we explore the quality control mechanisms utilised by the female germline to prevent, detect and remediate DNA damage. We follow the trajectory of development from the primordial follicle stage through to the preimplantation embryo, highlighting findings likely to have important implications for fertility management, age-related subfertility and premature ovarian failure. In addition, we survey the latest discoveries regarding DNA repair within the metaphase II (MII) oocyte and implicate maternal stores of endogenous DNA repair proteins and mRNA transcripts as a primary means by which they defend their genomic integrity. The collective evidence reviewed herein demonstrates that the MII oocyte can engage in the activation of major DNA damage repair pathway(s), therefore encouraging a reappraisal of the long-held paradigm that oocytes are largely refractory to DNA repair upon reaching this late stage of their development. It is also demonstrated that the zygote can exploit a number of protective strategies to mitigate the risk and/or effect the repair, of DNA damage sustained to either parental germline; affirming that DNA protection is largely a maternally driven trait but that some aspects of repair may rely on a collaborative effort between the male and female germlines. The present review highlights the vulnerability of the oocyte to DNA damage and presents a number of opportunities for research to bolster the stringency of the oocyte’s endogenous defences, with implications extending to improved diagnostics and novel therapeutic applications to alleviate the burden of infertility.

: In brief: Post-ovulatory ageing of oocytes leads to poor oocyte and embryo quality as well as abnormalities in offspring. This review provides an update on the contributions of oxidative stress to this process and discusses the current literature surrounding the use of antioxidant media to delay post-ovulatory oocyte ageing. Following ovulation, the metaphase II stage oocyte has a limited functional lifespan before succumbing to a process known as post-ovulatory oocyte ageing. This progressive demise occurs both in vivo and in vitro and is accompanied by a deterioration in oocyte quality, leading to a well-defined sequelae of reduced fertilisation rates, poor embryo quality, post-implantation errors, and abnormalities in the offspring. Although the physiological consequences of post-ovulatory oocyte ageing have largely been characterised, less is known regarding the molecular mechanisms that drive this process. This review presents an update on the established relationships between the biochemical changes exhibited by the ageing oocyte and the myriad of symptoms associated with the ageing phenotype. In doing so, we consider the molecular events that are potentially involved in orchestrating post-ovulatory ageing with a particular focus on the role of oxidative stress. We highlight the mounting evidence that oxidative stress acts as an initiator for a cascade of events that create the aged oocyte phenotype. Specifically, oxidative stress has the capacity to disrupt mitochondrial function and directly damage multiple intracellular components of the oocyte such as lipids, proteins, and DNA. Finally, this review addresses emerging strategies for delaying post-ovulatory oocyte ageing with emphasis placed on the promise afforded by the use of selected antioxidants to guide the development of media tailored for the preservation of oocyte integrity during in vitro fertilisation procedures.

The unique biology of the oocyte means that accepted paradigms for DNA repair and protection are not of direct relevance to the female gamete. Instead, preservation of the integrity of the maternal genome depends on endogenous protein stores and/or mRNA transcripts accumulated during oogenesis. The aim of this study was to determine whether mature (MII) oocytes have the capacity to detect DNA damage and subsequently mount effective repair. For this purpose, DNA double strand breaks (DSB) were elicited using the topoisomerase II inhibitor, etoposide (ETP). ETP challenge led to a rapid and significant increase in DSB (P = 0.0002) and the consequential incidence of metaphase plate abnormalities (P = 0.0031). Despite this, ETP-treated MII oocytes retained their ability to participate in in vitro fertilisation, though displayed reduced developmental competence beyond the 2-cell stage (P = 0.02). To account for these findings, we analysed the efficacy of DSB resolution, revealing a significant reduction in DSB lesions 4 h post-ETP treatment. Notably, this response was completely abrogated by pharmacological inhibition of key elements (DNA-PKcs and DNA ligase IV) of the canonical non-homologous end joining DNA repair pathway, thus providing the first evidence implicating this reparative cascade in the protection of the maternal genome.

Artificially generated radiofrequency-electromagnetic energy (RF-EME) is now ubiquitous in our environment owing to the utilization of mobile phone and Wi-Fi based communication devices. While several studies have revealed that RF-EME is capable of eliciting biological stress, particularly in the context of the male reproductive system, the mechanistic basis of this biophysical interaction remains largely unresolved. To extend these studies, here we exposed unrestrained male mice to RF-EME generated via a dedicated waveguide (905 MHz, 2.2 W/kg) for 12 h per day for a period of 1, 3 or 5 weeks. The testes of exposed mice exhibited no evidence of gross histological change or elevated stress, irrespective of the RF-EME exposure regimen. By contrast, 5 weeks of RF-EME exposure adversely impacted the vitality and motility profiles of mature epididymal spermatozoa. These spermatozoa also experienced increased mitochondrial generation of reactive oxygen species after 1 week of exposure, with elevated DNA oxidation and fragmentation across all exposure periods. Notwithstanding these lesions, RF-EME exposure did not impair the fertilization competence of spermatozoa nor their ability to support early embryonic development. This study supports the utility of male germ cells as sensitive tools with which to assess the biological impacts of whole-body RF-EME exposure.

Forecasted increases in the prevalence and severity of extreme weather events accompanying changes in climatic behavior pose potential risk to the reproductive capacity of humans and animals of ecological and agricultural significance. While several studies have revealed that heat stress induced by challenges such as testicular insulation can elicit a marked negative effect on the male reproductive system, and particularly the production of spermatozoa, less is known about the immediate impact on male reproductive function following subchronic whole-body exposure to elevated ambient temperature. To address this knowledge gap, we exposed unrestrained male mice to heat stress conditions that emulate a heat wave (daily cycle of 8 h at 35 °C followed by 16 h at 25 °C) for a period of 7 d. Neither the testes or epididymides of heat-exposed male mice exhibited evidence of gross histological change, and similarly, spermatozoa of exposed males retained their functionality and ability to support embryonic development. However, the embryos generated from heat-exposed spermatozoa experienced pronounced changes in gene expression linked to acceleration of early embryo development, aberrant blastocyst hatching, and increased fetal:placental weight ratio. Such changes were causally associated with an altered sperm small noncoding RNA (sncRNA) profile, such that these developmental phenotypes were recapitulated by microinjection of wild-type embryos sired by control spermatozoa with RNAs extracted from heat-exposed spermatozoa. Such data highlight that even relatively modest excursions in ambient temperature can affect male reproductive function and identify the sperm sncRNA profile as a particular point of vulnerability to this imposed environmental stress.

BACKGROUND: Male factors contribute to ∼50% of all infertility cases globally and are a major contributor to escalating use of ART. In most instances, sub-fertile men retain the ability to produce spermatozoa, albeit with reduced quality and function. By necessity, an important feature of ART is the use of technologies that bypass the natural selection barriers that prevent poor-quality spermatozoa from participating in fertilization. This means that ART carries a significant risk of facilitating fertilization with poor-quality gametes harbouring undetected DNA damage and/or altered epigenomes. Such a scenario may account for the epidemiological links between the use of ‘high intervention’ technologies [e.g. ICSI] and an increased risk of adverse offspring outcomes. Such data highlight a pressing need for improved sperm selection tools that better mimic natural selection barriers, to ensure only the highest-quality spermatozoa are used for ART. OBJECTIVE AND RATIONALE: Current sperm selection techniques for ART and the processes underpinning sperm maturation have often been considered independently and therefore reviewed separately. Here we outline the requirement for connecting research paradigms towards advancing clinical outcomes. This review highlights the importance of combining our advancing knowledge of sperm maturation biology with the pursuit of rational sperm selection strategies for the clinic; specifically, this narrative review summarizes the current clinical technologies used for sperm selection with a focus on their physiological relevance and limitations. We have given consideration to the events associated with sperm maturation and the importance of zona pellucida (ZP) binding as inspiration to inform the development of the next generation of sperm selection strategies. The connections and information presented should provide utility and interest for both clinicians and reproductive biologists alike. SEARCH METHODS: The PubMed database was queried using the keywords: sperm selection/function/DNA quality/epigenome, ART, ICSI, male infertility, capacitation, zona pellucida, sperm–zona pellucida binding, DNA damage, and biofabrication. These keywords were combined with other relevant phrases. Literature was restricted to peer-reviewed articles in English (published between 1972 and 2024) with references within these articles also searched. OUTCOMES: During natural conception, high-quality sperm are ‘selected’, maximizing the chances of fertilization with healthy gametes carrying intact genomic/epigenetic cargo. This sub-population of spermatozoa possess the capacity to interact with the female reproductive tract and complete the suite of functional maturation processes required for successful fertilization and initiation of embryonic development. However, ART ‘high intervention’ strategies bypass these selective barriers leading to an increased risk of inadvertently transferring genomic defects to the offspring with potential downstream consequences for offspring health. This review contextualizes why current sperm selection technologies have provided only minor improvement to live birth rates following ART. We posit that capitalizing on sperm–ZP binding (the penultimate step of successful fertilization) with novel ZP mimetic technologies provides an attractive, but understudied, tool for clinical selection of fertilization-competent spermatozoa for ART improvement. LIMITATIONS, REASONS FOR CAUTION: The risk of bias in the interpretation of findings for a narrative review cannot be completely eliminated. Literature was limited to the language the authors speak: English. WIDER IMPLICATIONS: ART has provided transformative advancement for infertile couples, however, gaps in our fundamental understanding of how the best gametes are ‘selected’ during natural conception, which when unaccounted for during clinical conception, present a risk of continued reliance on ART and health consequences for the next generation. The purpose of this article was to contextualize our current knowledge across both sperm maturation events and current selection strategies for these cells in the clinic. We outline the potential for improved clinical outcomes through the advancement of our understanding in gamete biology in concert with the development of novel methods for artificial gamete selection. STUDY FUNDING/COMPETING INTEREST(S): No external funding, but financial support was received from the School of Environmental and Life Sciences, University of Newcastle, Australia. R.J.A. is a scientific advisor to Memphasys Ltd, a biotechnology company with interests in reproductive health and responsible for developing the Felix™ electrophoretic sperm isolation device. R.J.A. receives salary and grant from, and has stock in, Memphasys Ltd. The other authors declare no conflicts of interest.