COVID-19 gender policy changes support female scientists and improve research quality.

With more time being spent on caregiving responsibilities during the COVID-19 pandemic, female scientists' productivity dropped. When female scientists conduct research, identity factors are better incorporated in research content. In order to mitigate damage to the research enterprise, funding agencies can play a role by putting in place gender equity policies that support all applicants and ensure research quality. A national health research funder implemented gender policy changes that included extending deadlines and factoring sex and gender into COVID-19 grant requirements. Following these changes, the funder received more applications from female scientists, awarded a greater proportion of grants to female compared to male scientists, and received and funded more grant applications that considered sex and gender in the content of COVID-19 research. Further work is urgently required to address inequities associated with identity characteristics beyond gender.

A 10-year longitudinal evaluation of science policy interventions to promote sex and gender in health research.

BACKGROUND: Over the past decade, the Canadian Institutes of Health Research (CIHR) has implemented multicomponent interventions to increase the uptake of sex and gender in grant applications. Interventions included mandatory reporting on applicant forms, development of resources for applicants and evaluators, and grant review requirements. Here, we aim to inform science policy implementation by describing the 10-year outcomes and lessons learned from these interventions. METHODS: This is a prospective longitudinal study. The population is all applicants across 15 investigator-initiated CIHR competitions from 2011 to 2019 and grant evaluators from 2018 to 2019. Quantitative data were derived from applicants' and grant evaluators' mandatory reporting of sex and gender integration in the grants management database. The application was the unit of analysis. Trends in sex and gender uptake in applications were plotted over time, stratified by research area. Univariate logistic regression was used to assess associations between the sex of the applicant and the uptake of sex and gender, and the latter with funding success. Qualitative review of the quality and appropriateness of evaluators' comments informed the development of discipline-specific training to peer review committee members. Feedback was compiled from a subset of evaluators on the perceived usefulness of the educational materials using a brief questionnaire. RESULTS: Since 2011, 39,390 applications were submitted. The proportion that reported integration of sex rose from 22 to 83%, and gender from 12 to 33%. Population health research applications paid the greatest attention to gender (82%). Across every competition, applications with female principal investigators were more likely to integrate sex (odds ratio [OR] 1.60, 95% confidence interval [CI] 1.50-1.63) and gender (OR 2.40, 95% CI 2.29-2.51) than those who identified as male. Since 2018, applications that scored highly for the integration of sex (OR 1.92, 95% CI 1.50-2.50) and gender (OR 2.53, 95% CI 1.83-3.50) were more likely to be funded. Qualitative observations revealed persistent conflation of the terms sex and gender. Eighty-six percent of evaluators appreciated the tailored discipline-specific coaching. CONCLUSIONS: A number of policy interventions improved sex and gender uptake in grant applications, with higher success rates observed over time for applications that integrated sex and gender. Other funders' action plans around sex and gender integration may be informed from our experiences of the timing, type and targets of the different interventions, specifically those directed at evaluators.

Micronucleus formation causes perpetual unilateral chromosome inheritance in mouse embryos.

Chromosome segregation defects in cancer cells lead to encapsulation of chromosomes in micronuclei (MN), small nucleus-like structures within which dangerous DNA rearrangements termed chromothripsis can occur. Here we uncover a strikingly different consequence of MN formation in preimplantation development. We find that chromosomes from within MN become damaged and fail to support a functional kinetochore. MN are therefore not segregated, but are instead inherited by one of the two daughter cells. We find that the same MN can be inherited several times without rejoining the principal nucleus and without altering the kinetics of cell divisions. MN motion is passive, resulting in an even distribution of MN across the first two cell lineages. We propose that perpetual unilateral MN inheritance constitutes an unexpected mode of chromosome missegregation, which could contribute to the high frequency of aneuploid cells in mammalian embryos, but simultaneously may serve to insulate the early embryonic genome from chromothripsis.

Tri-directional anaphases as a novel chromosome segregation defect in human oocytes.

STUDY QUESTION: What are the chromosome segregation errors in human oocyte meiosis-I that may underlie oocyte aneuploidy? SUMMARY ANSWER: Multiple modes of chromosome segregation error were observed, including tri-directional anaphases, which we attribute to loss of bipolar spindle structure at anaphase-I. WHAT IS KNOWN ALREADY: Oocyte aneuploidy is common and associated with infertility, but mechanistic information on the chromosome segregation errors underlying these defects is scarce. Lagging chromosomes were recently reported as a possible mechanism by which segregation errors occur. STUDY DESIGN, SIZE, DURATION: Long-term confocal imaging of chromosome dynamics in 50 human oocytes collected between January 2015 and May 2016. PARTICIPANTS/MATERIALS, SETTING, METHODS: Germinal vesicle (GV) stage oocytes were collected from women undergoing intracytoplasmic sperm injection cycles and also CD1 mice. Oocytes were microinjected with complementary RNAs to label chromosomes, and in a subset of oocytes, the meiotic spindle. Oocytes were imaged live through meiosis-I using confocal microscopy. 3D image reconstruction was used to classify chromosome segregation phenotypes at anaphase-I. Segregation phenotypes were related to spindle dynamics and cell cycle timings. MAIN RESULTS AND THE ROLE OF CHANCE: Most (87%) mouse oocytes segregated chromosomes with no obvious defects. We found that 20% of human oocytes segregated chromosomes bi-directionally with no lagging chromosomes. The rest were categorised as bi-directional anaphase with lagging chromosomes (20%), bi-directional anaphase with chromatin mass separation (34%) or tri-directional anaphase (26%). Segregation errors correlated with chromosome misalignment prior to anaphase. Spindles were tripolar when tri-directional anaphases occurred. Anaphase phenotypes did not correlate with meiosis-I duration (P = 0.73). LARGE SCALE DATA: Not applicable. LIMITATIONS, REASONS FOR CAUTION: Oocytes were recovered at GV stage after gonadotrophin-stimulation, and the usual oocyte quality caveats apply. Whilst the possibility that imaging may affect oocyte physiology cannot be formally excluded, detailed controls and justifications are presented. WIDER IMPLICATIONS OF THE FINDINGS: This is one of the first reports of live imaging of chromosome dynamics in human oocytes, introducing tri-directional anaphases as a novel potential mechanism for oocyte aneuploidy. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by grants from Fondation Jean-Louis Lévesque (Canada), CIHR (MOP142334) and CFI (32711) to GF. JH is supported by Postdoctoral Fellowships from The Lalor Foundation and CIHR (146703). The authors have no conflict of interest.

Suppression of Sertoli cell tumour development during the first wave of spermatogenesis in inhibin α-deficient mice.

A dynamic partnership between follicle-stimulating hormone (FSH) and activin is required for normal Sertoli cell development and fertility. Disruptions to this partnership trigger Sertoli cells to deviate from their normal developmental pathway, as observed in inhibin α-knockout (Inha-KO) mice, which feature Sertoli cell tumours in adulthood. Here, we identified the developmental windows by which adult Sertoli cell tumourigenesis is most FSH sensitive. FSH was suppressed for 7 days in Inha-KO mice and wild-type littermates during the 1st, 2nd or 4th week after birth and culled in the 5th week to assess the effect on adult Sertoli cell development. Tumour growth was profoundly reduced in adult Inha-KO mice in response to FSH suppression during Weeks 1 and 2, but not Week 4. Proliferative Sertoli cells were markedly reduced in adult Inha-KO mice following FSH suppression during Weeks 1, 2 or 4, resulting in levels similar to those in wild-type mice, with greatest effect observed at the 2 week time point. Apoptotic Sertoli cells increased in adult Inha-KO mice after FSH suppression during Week 4. In conclusion, acute FSH suppression during the 1st or 2nd week after birth in Inha-KO mice profoundly suppresses Sertoli cell tumour progression, probably by inhibiting proliferation in the adult, with early postnatal Sertoli cells being most sensitive to FSH action.

Distinct classes of lagging chromosome underpin age-related oocyte aneuploidy in mouse.

Chromosome segregation errors that cause oocyte aneuploidy increase in frequency with maternal age and are considered a major contributing factor of age-related fertility decline in females. Lagging anaphase chromosomes are a common age-associated phenomenon in oocytes, but whether anaphase laggards actually missegregate and cause aneuploidy is unclear. Here, we show that lagging chromosomes in mouse oocytes comprise two mechanistically distinct classes of chromosome motion that we refer to as "class-I" and "class-II" laggards. We use imaging approaches and mechanistic interventions to dissociate the two classes and find that whereas class-II laggards are largely benign, class-I laggards frequently directly lead to aneuploidy. Most notably, a controlled prolongation of meiosis I specifically lessens class-I lagging to prevent aneuploidy. Our data thus reveal lagging chromosomes to be a cause of age-related aneuploidy in mouse oocytes and suggest that manipulating the cell cycle could increase the yield of useful oocytes in some contexts.

Considering how biological sex impacts immune responses and COVID-19 outcomes.

A male bias in mortality has emerged in the COVID-19 pandemic, which is consistent with the pathogenesis of other viral infections. Biological sex differences may manifest themselves in susceptibility to infection, early pathogenesis, innate viral control, adaptive immune responses or the balance of inflammation and tissue repair in the resolution of infection. We discuss available sex-disaggregated epidemiological data from the COVID-19 pandemic, introduce sex-differential features of immunity and highlight potential sex differences underlying COVID-19 severity. We propose that sex differences in immunopathogenesis will inform mechanisms of COVID-19, identify points for therapeutic intervention and improve vaccine design and increase vaccine efficacy.

Ca(2+) dynamics in oocytes from naturally-aged mice.

The ability of human metaphase-II arrested eggs to activate following fertilisation declines with advancing maternal age. Egg activation is triggered by repetitive increases in intracellular Ca(2+) concentration ([Ca(2+)]i) in the ooplasm as a result of sperm-egg fusion. We therefore hypothesised that eggs from older females feature a reduced ability to mount appropriate Ca(2+) responses at fertilisation. To test this hypothesis we performed the first examination of Ca(2+) dynamics in eggs from young and naturally-aged mice. Strikingly, we find that Ca(2+) stores and resting [Ca(2+)]i are unchanged with age. Although eggs from aged mice feature a reduced ability to replenish intracellular Ca(2+) stores following depletion, this difference had no effect on the duration, number, or amplitude of Ca(2+) oscillations following intracytoplasmic sperm injection or expression of phospholipase C zeta. In contrast, we describe a substantial reduction in the frequency and duration of oscillations in aged eggs upon parthenogenetic activation with SrCl2. We conclude that the ability to mount and respond to an appropriate Ca(2+) signal at fertilisation is largely unchanged by advancing maternal age, but subtle changes in Ca(2+) handling occur that may have more substantial impacts upon commonly used means of parthenogenetic activation.

Differential permeability of the blood-testis barrier during reinitiation of spermatogenesis in adult male rats.

The blood-testis barrier (BTB) sequesters meiotic spermatocytes and differentiating spermatids away from the vascular environment. We aimed to assess whether meiosis and postmeiotic differentiation could occur when the BTB is permeable. Using a model of meiotic suppression and reinitiation, BTB function was assessed using permeability tracers of small, medium, and large (0.6-, 70-, and 150-kDa) sizes to emulate blood- and lymphatic-borne factors that could cross the BTB. Adult rats (n = 9/group) received the GnRH antagonist acyline (10 wk) to suppress gonadotropins, followed by testosterone (24cm Silastic implant), for 2, 4, 7, 10, 15, and 35 days. In acyline-suppressed testes, all tracers permeated the seminiferous epithelium. As spermatocytes up to diplotene stage XIII reappeared, both the 0.6- and 70-kDa tracers, but not 150 kDa, permeated around these cells. Intriguingly, the 0.6- and 70-kDa tracers were excluded from pachytene spermatocytes at stages VII and VIII but not in subsequent stages. The BTB became progressively impermeable to the 0.6- and 70-kDa tracers as stages IV-VII round spermatids reappeared in the epithelium. This coincided with the appearance of the tight junction protein, claudin-12, in Sertoli cells and at the BTB. We conclude that meiosis can occur when the BTB is permeable to factors up to 70 kDa during the reinitiation of spermatogenesis. Moreover, BTB closure corresponds with the presence of particular pachytene spermatocytes and round spermatids. This research has implications for understanding the effects of BTB dynamics in normal spermatogenesis and also potentially in states where spermatogenesis is suppressed, such as male hormonal contraception or infertility.

Claudin-11 and connexin-43 display altered spatial patterns of organization in men with primary seminiferous tubule failure compared with controls.

OBJECTIVE: To assess the spatial organization of two proteins involved in the Sertoli cell junctional complex in men with primary seminiferous tubule failure. DESIGN: Retrospective study. SETTING: Medical research institute. PATIENT(S): Sixteen men total, six with meiotic arrest, seven with the Sertoli cell-only phenotype, and three with normal spermatogenesis. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Differences in claudin-11 and connexin-43 organization as detected using confocal microscopy. RESULT(S): In men with primary seminiferous tubule failure, four organizational patterns (I-IV) were recognized and quantified for claudin-11. Across these patterns, claudin-11 changed from a basal filamentous staining pattern to a punctate staining pattern with diffuse localization throughout the entire epithelium. Similar changes in staining patterns for connexin-43 were observed. Major differences were seen in the spatial organization of claudin-11 and connexin-43 in tubules from control men compared with tubules with primary seminiferous tubule failure, but we observed no differences in the spatial organization of these proteins in tubules from men with meiotic arrest and Sertoli cell-only phenotypes. CONCLUSION(S): The spatial organization of claudin-11 and connexin-43 is altered in men with primary seminiferous tubule failure. Disorganization of the proteins composing the Sertoli cell junctional complex may be involved in the spermatogenic impairment, possibly via loss of blood-testis barrier function.

Activin signaling regulates Sertoli cell differentiation and function.

Throughout development, activin A signaling stimulates proliferation and inhibits differentiation of testicular Sertoli cells. A decline in activin levels at puberty corresponds with the differentiation of Sertoli cells that is required to sustain spermatogenesis. In this study, we consider whether terminally differentiated Sertoli cells can revert to a functionally immature phenotype in response to activin A. To increase systemic activin levels, the right tibialis anterior muscle of 7-wk-old C57BL/6J mice was transduced with an adeno-associated virus (rAAV6) expressing activin A. We show that chronic activin signaling reduces testis mass by 23.5% compared with control animals and induces a hypospermatogenic phenotype, consistent with a failure of Sertoli cells to support spermatogenesis. We use permeability tracers and transepithelial electrical resistance measurements to demonstrate that activin potently disrupts blood-testis-barrier function in adult mice and ablates tight junction formation in differentiated primary Sertoli cells, respectively. Furthermore, increased activin signaling reinitiates a program of cellular proliferation in primary Sertoli cells as determined by 5-ethynyl-2'-deoxyuridine incorporation. Proliferative cells reexpress juvenile markers, including cytokeratin-18, and suppress mature markers, including claudin-11. Thus, activin A is the first identified factor capable of reprogramming Sertoli cells to an immature, dedifferentiated phenotype. This study indicates that activin signaling must be strictly controlled in the adult in order to maintain Sertoli cell function in spermatogenesis.

Teasing out the role of aromatase in the healthy and diseased testis.

Scientific discoveries over the past decade have shifted the stereotypical view of androgens as male hormones and estrogens as female hormones. It is now recognized that a delicate balance of both androgens and estrogens, a process controlled by aromatase, is fundamental for normal testicular development and fertility. While the site-specific actions of these two classes of steroids within the testis are becoming better documented, the role and regulation of estrogen biosynthesis by aromatase within the testis remains unclear. The majority of data comes from a wide range of animal species, particularly genetically modified mouse models; aromatase knockout (ArKO) and overexpressing (AROM(+)), with limited information on humans, however the existence of congenital aromatase mutations has provided some insight into its effects on individual parameters of the testis. This review dissects out the localization and activity of aromatase in the healthy and diseased testis, addresses the cellular insult to the testis that occurs in its absence and over abundance and proposes potential molecular mechanisms of aromatase regulation in the testis.