Fertility In Mice Research Articles

CCDC146 is required for sperm flagellum biogenesis and male fertility in mice.

Multiple morphological abnormalities of the flagella (MMAF) is a severe disease of male infertility, while the pathogenetic mechanisms of MMAF are still incompletely understood. Previously, we found that the deficiency of Ccdc38 might be associated with MMAF. To understand the underlying mechanism of this disease, we identified the potential partner of this protein and found that the coiled-coil domain containing 146 (CCDC146) can interact with CCDC38. It is predominantly expressed in the testes, and the knockout of this gene resulted in complete infertility in male mice but not in females. The knockout of Ccdc146 impaired spermiogenesis, mainly due to flagellum and manchette organization defects, finally led to MMAF-like phenotype. Furthermore, we demonstrated that CCDC146 could interact with both CCDC38 and CCDC42. It also interacts with intraflagellar transport (IFT) complexes IFT88 and IFT20. The knockout of this gene led to the decrease of ODF2, IFT88, and IFT20 protein levels, but did not affect CCDC38, CCDC42, or ODF1 expression. Additionally, we predicted and validated the detailed interactions between CCDC146 and CCDC38 or CCDC42, and built the interaction models at the atomic level. Our results suggest that the testis predominantly expressed gene Ccdc146 is essential for sperm flagellum biogenesis and male fertility, and its mutations might be associated with MMAF in some patients.

Cylicins are a structural component of the sperm calyx being indispensable for male fertility in mice and human.

Cylicins are testis-specific proteins, which are exclusively expressed during spermiogenesis. In mice and humans, two Cylicins, the gonosomal X-linked Cylicin 1 (Cylc1/CYLC1) and the autosomal Cylicin 2 (Cylc2/CYLC2) genes, have been identified. Cylicins are cytoskeletal proteins with an overall positive charge due to lysine-rich repeats. While Cylicins have been localized in the acrosomal region of round spermatids, they resemble a major component of the calyx within the perinuclear theca at the posterior part of mature sperm nuclei. However, the role of Cylicins during spermiogenesis has not yet been investigated. Here, we applied CRISPR/Cas9-mediated gene editing in zygotes to establish Cylc1- and Cylc2-deficient mouse lines as a model to study the function of these proteins. Cylc1 deficiency resulted in male subfertility, whereas Cylc2-/-, Cylc1-/yCylc2+/-, and Cylc1-/yCylc2-/- males were infertile. Phenotypical characterization revealed that loss of Cylicins prevents proper calyx assembly during spermiogenesis. This results in decreased epididymal sperm counts, impaired shedding of excess cytoplasm, and severe structural malformations, ultimately resulting in impaired sperm motility. Furthermore, exome sequencing identified an infertile man with a hemizygous variant in CYLC1 and a heterozygous variant in CYLC2, displaying morphological abnormalities of the sperm including the absence of the acrosome. Thus, our study highlights the relevance and importance of Cylicins for spermiogenic remodeling and male fertility in human and mouse, and provides the basis for further studies on unraveling the complex molecular interactions between perinuclear theca proteins required during spermiogenesis.

Dnali1 is required for sperm motility and male fertility in mice

BackgroundThe sperm flagellum is an evolutionarily conserved specialized organelle responsible for sperm motility and male fertility. Deleterious mutations in genes involved in the sperm flagellum assembly can often cause sperm motility defects and male infertility. The murine Dnali1 gene encodes a protein that is known to interact with the cytoplasmic dynein heavy chain 1.ResultsA Dnali1-mutated mouse model was generated by inducing a nonsense mutation in the Dnali1 gene. The Dnali1-mutated male mice presented impaired sperm motility and were completely infertile. Although no obviously abnormal sperm morphology was observed in Dnali1-mutated male mice, the ultrastructural structure of sperm flagellum was disrupted, displaying as an asymmetrical distribution of the longitudinal columns (LCs). Notably, infertile Dnali1-mutated male mice were able to obtain offspring via ICSI.ConclusionsOur results uncover a role of DNALI1 in sperm motility and male fertility in mice, and demonstrate that ICSI overcomes Dnali1-associated male infertility, thus providing guidance for the diagnosis and genetic counseling of DNALI1-associated human infertility.

FAM71D is dispensable for spermatogenesis and male fertility in mice.

In mammals, the generation of sperm cells capable of fertilization is a highly complex process including spermatogenesis in the testis and maturation in the epididymis. In our previous study, we have demonstrated that FAM71D (Family with sequence similarity 71, member D), which could interact with calmodulin, was highly expressed in human and mouse testis. To investigate the physiological role of FAM71D in spermatogenesis, we next generateFam71d loss-of-function mouse model using CRISPR/Cas9 technology. We performed immunofluorescence and RT-qPCR to examine the protein and mRNA expression in testicular cells. We found that FAM71D was predominantly localized in the round and elongated spermatids. And FAM71D KO mice displayed normal development of germ cell and fertility. Furthermore, testicular histology and sperm concentration showed no significant difference between WT and KO mice. These data demonstrate that FAM71D is dispensable for mouse spermatogenesis and male fertility.

Asperosaponin VI protects against spermatogenic dysfunction in mice by regulating testicular cell proliferation and sex hormone disruption

Ethnopharmacological relevanceStudies have found that the causes of male infertility are complex, and spermatogenic dysfunction accounts for 30%–65% of male infertility causes, which is the main cause of male infertility. Asperosaponin VI (ASVI) is a saponin extracted from the traditional Chinese herb Dipsacus asperoides C.Y.Cheng & T.M.Ai. However, the precise protective impact and underlying mechanism of ASVI in the therapy of spermatogenic dysfunction remain unknown. Aim of the studyTo investigate the impact of ASVI on the spermatogenic dysfunction induced by cytoxan (CTX) in mice, as well as explore any potential mechanisms. Materials and methodsPotential ASVI targets were screened using the Pharmapper and Uniprot databases, while genes related to spermatogenic dysfunction were collected from the GeneCards database. The String and Cytoscape databases were then used for PPI analysis for the common targets of ASVI and spermatogenic dysfunction. Meanwhile, the Metascape database was used for KEGG and GO analysis. In vivo experiments, spermatogenic dysfunction was induced in male mice by intraperitoneal administration of CTX (80 mg/kg). To demonstrate the possible protective effects of ASVI on reproductive organs, CTX-induced spermatogenic dysfunction mice with different dosages of ASVI (0.8, 4, 20 mg/kg per day) treatment were collected and gonad weight was detected. The testis and epididymis were detected again by H&E. To assess the impact of ASVI on fertility in male mice, we analyzed sperm quality, serum hormones, sexual behavior, and fertility. The mechanism was investigated using WB, IF, IHC, and Co-IP technology. ResultsThe ASVI exhibited interactions with 239 associated targets. Furthermore, 1555 targets associated with spermatogenic dysfunction were predicted, and further PPI analysis identified 6 key targets. Among them, the EGFR gene exhibited the highest degree of connection and was at the core of the network. Based on the GO and KEGG enrichment analysis, ASVI may affect spermatogenic dysfunction through the EGFR pathway. In vivo experiments, ASVI significantly improved CTX-induced damage to male fertility and reproductive organs, increasing sperm quality. At the same time, ASVI can resist CTX-induced testicular cell damage by increasing p-EGFR, p-ERK, PCNA, and p-Rb in the testis and by promoting the interaction of CyclinD1 with CDK4. In addition, ASVI can also regulate sex hormone disorders and protect male fertility. ConclusionsASVI improves CTX-induced spermatogenesis dysfunction by activating the EGFR signaling pathway and regulating sex hormone homeostasis, which may be a new potential protective agent for male spermatogenic dysfunction.

Autophagy core protein BECN1 is vital for spermatogenesis and male fertility in mice†.

Mammalian spermatogenesis is a highly complex multi-step biological process, and autophagy has been demonstrated to be involved in the process of spermatogenesis. Beclin-1/BECN1, a core autophagy factor, plays a critical role in many biological processes and diseases. However, its function in spermatogenesis remains largely unclear. In the present study, germ cell-specific Beclin 1 (Becn1) knockout mice were generated and were conducted to determine the role of Becn1 in spermatogenesis and fertility of mice. Results indicate that Becn1 deficiency leads to reduced sperm motility and quantity, partial failure of spermiation, actin network disruption, excessive residual cytoplasm, acrosome malformation, and aberrant mitochondrial accumulation of sperm, ultimately resulting in reduced fertility in male mice. Furthermore, inhibition of autophagy was observed in the testes of germ cell-specific Becn1 knockout mice, which may contribute to impaired spermiogenesis and reduced fertility. Collectively, our results reveal that Becn1 is essential for fertility and spermiogenesis in mice.

Non-monotonic effects of Bisphenol A Dimethacrylate on male mouse reproductive system and fertility leads to impaired conceptive performance

As an estrogenic agent, Bisphenol A Dimethacrylate (Bis-DMA) may incite alterations in both the reproductive tract and the neuroendocrine axis, and thus have the potential to affect the proper development, maturity and conceptive performance in animals. We investigated the consequences of 14 weeks of exposure to different concentrations of Bis-DMA on male mouse conceptive performance. Male mice were exposed to Bis-DMA (0, 0.1 mg/L, 1.0 mg/L or 10 mg/L) via drinking water, and the effects on fertility, reproductive organ weights, reproductive hormone levels, sperm counts and testicular histology were assessed. We clearly demonstrate that prolonged exposure of male mice to Bis-DMA negatively affects fertility and reproduction causing significant reductions in sperm counts, non-monotonic effects on serum LH and testosterone levels, increased seminal vesicle weights, lower number of embryonic implantations and viable fetuses, as well as, increased embryonal resorptions in females mated by Bis-DMA treated males. Furthermore, Bis-DMA caused abnormalities in testicular infrastructure with atrophic seminiferous tubules exhibiting intraepithelial vacuolization and disorganization, loss and shedding of germ cells into the lumen, and presence of apoptotic cells. Our data collectively suggest that Bis-DMA adversely affects male fertility and reproduction by interference with normal hormone signaling in the testis, inducing changes in testicular infrastructure and ultimately leading to impaired reproductive function and fertility.

In pre-clinical study fetal hypoxia caused autophagy and mitochondrial impairment in ovary granulosa cells mitigated by melatonin supplement

IntroductionFetal hypoxia has long-term effects on postnatal reproductive functions and the mitochondrial impairments of ovarian granulosa cells may be one of the causes. Melatonin applied to mitigate mitochondrial dysfunction and autophagy in mammalian cells has been reported. However, the potential mechanisms by which fetal hypoxia damages reproductive function in neonatal female mice and the melatonin effects on this problem remain unclear. ObjectivesThis research aimed to explore the mechanism that fetal hypoxia damages reproductive function in neonatal female mice and attempt to improve the reproductive function by treating with melatonin in vivo and in vitro. MethodsWe established a fetal hypoxia model and confirmed that fetal hypoxia affects ovarian function by inducing GC excessive autophagy. Transcriptomic analysis, gene interference, cell immunofluorescence, immunohistochemistry and western blot were conducted to explore and verify the underlying mechanisms in mice GCs and KGN cells. Finally, melatonin treatment was executed on hypoxia-treated mice GCs and KGN cells and melatonin injection to fetal-hypoxia-treated mice to determine its effect. ResultsThe results of in vitro experiments found that fetal hypoxia led to mitochondrial dysfunction in ovarian GCs causing autophagic cell death. And the PI3K/Akt/FoxO pathway mediated the occurrence of this process by transcriptome analysis of ovarian GCs from normal and fetal hypoxia mice, which was further verified in mice GCs and KGN cells. Additionally, melatonin administration prevented autophagic injuries and mitochondrial impairments in hypoxia-treated mice GCs and KGN cells. Meanwhile, in vivo experiments by melatonin injection ameliorated oxidative stress of ovary in fetal-hypoxia-treated mice and improved their low fertility. ConclusionOur data found that fetal hypoxia causes ovarian GCs excessive autophagy leading to low fertility in neonatal female mice and mitigated by melatonin. These results provide a potential therapy for hypoxic stress-related reproductive disorders.

Classical Estrogen Signaling in Ciliated Epithelial Cells of the Oviduct Is Nonessential for Fertility in Female Mice.

Ciliary action performs a critical role in the oviduct (Fallopian tube) during pregnancy establishment through sperm and egg transport. The disruption of normal ciliary function in the oviduct affects oocyte pick-up and is a contributing factor to female infertility. Estrogen is an important regulator of ciliary action in the oviduct and promotes ciliogenesis in several species. Global loss of estrogen receptor α (ESR1) leads to infertility. We have previously shown that ESR1 in the oviductal epithelial cell layer is required for female fertility. Here, we assessed the role of estrogen on transcriptional regulation of ciliated epithelial cells of the oviduct using single-cell RNA-sequencing analysis. We observed minor variations in ciliated cell genes in the proximal region (isthmus and uterotubal junction) of the oviduct. However, 17β-estradiol treatment had little impact on the gene expression profile of ciliated epithelial cells. We also conditionally ablated Esr1 from ciliated epithelial cells of the oviduct (called ciliated Esr1d/d mice). Our studies showed that ciliated Esr1d/d females had fertility rates comparable to control females, did not display any disruptions in preimplantation embryo development or embryo transport to the uterus, and had comparable cilia formation to control females. However, we observed some incomplete deletion of Esr1 in the ciliated epithelial cells, especially in the ampulla region. Nevertheless, our data suggest that ESR1 expression in ciliated cells of the oviduct is dispensable for ciliogenesis and nonessential for female fertility in mice.

High-Salt Diet Causes Defective Oocyte Maturation and Embryonic Development to Impair Female Fertility in Mice.

High salinity has been reported to induce many human disorders in tissues and organs to interfere with their normal physiological functions. However, it is unknown how salinity affects the development of female germ cells. This study suggests that a high-salt diet (HSD) may weaken oocyte quality to impair female fertility in mice and investigates the underlying mechanisms. C57BL/6 female mice are fed with a regular diet (Control) or a high-salt diet (HSD). Oocyte maturation, fertilization rate, embryonic development, and female fertility are evaluated. In addition, the spindle organization, actin polymerization, and kinetochore-microtubule attachment of oocytes are examined in both groups. Moreover, single-cell transcriptome data are used to demonstrate how HSD alters the transcript levels of genes. The observations confirm that HSD leads to female subfertility due to the deterioration of oocyte and embryo quality. The mechanism underlying reveals HSD compromises the oocytes' autophagy, apoptosis level, and mitochondrial function. The work illustrates that a high concentration of salt diet results in oocyte meiotic arrest, fertilization failure, and early developmental defection that embryos undergo to reduce female fertility in mice by perturbing the level of autophagy and apoptosis, mitochondrial function in oocytes.

Spermidine promotes fertility in aged female mice.

Spermidine promotes fertility in aged female mice.

Polyamine metabolite spermidine rejuvenates oocyte quality by enhancing mitophagy during female reproductive aging.

Advanced age is a primary risk factor for female infertility due to reduced ovarian reserve and declining oocyte quality. However, as an important contributing factor, the role of metabolic regulation during reproductive aging is poorly understood. Here, we applied untargeted metabolomics to identify spermidine as a critical metabolite in ovaries to protect oocytes against aging. In particular, we found that the spermidine level was reduced in ovaries of aged mice and that supplementation with spermidine promoted follicle development, oocyte maturation, early embryonic development and female fertility of aged mice. By microtranscriptomic analysis, we further discovered that spermidine-induced recovery of oocyte quality was mediated by enhancement of mitophagy activity and mitochondrial function in aged mice, and this mechanism of action was conserved in porcine oocytes under oxidative stress. Altogether, our findings suggest that spermidine supplementation could represent a therapeutic strategy to ameliorate oocyte quality and reproductive outcome in cis-gender women and other persons trying to conceive at an advanced age. Future work is needed to test whether this approach can be safely and effectively translated to humans.

The Effect of Phenolic Extract of Mint (Mentha spicata) in the Fertility of Male Laboratory Mice.

The current study was conducted to investigate the effect of the phenolic extract of the Mentha spicata plant on the fertility of laboratory mice. The phenolic compounds were extracted using the Reflex condenser from the leaves of the Mentha spicata plant, and it was chemically detected. In the current study,)Albino Mice of the musculus Mus type was used at an age ranging between (12-11) months. The group was randomly divided into three groups. The control group was injected with a physiological solution (0.9% NaCl), the second group was injected with the phenolic extract of the Mentha spicata plant at (100 mg/kg) concentration, and the third group was injected by concentration (200 mg/kg) from the phenolic extract of the Mentha spicata. The phenols were dissolved in a physiological solution (0.9% NaCl), and all individuals of these groups were injected through the peritoneal membrane I.P. at a rate of 0.5 ml per animal per day for 30 days. The phenolic extract of Mentha spicata leaves caused a significant decrease (P<0.05) in the level of Testosterone and Luteinizing hormone (ng/ml) in the blood serum of the first and second treatment group when compared with the control group. The study also indicates a significant decrease (P<0.05) in the concentration of Follicle-Stimulating Hormone in the third treatment group compared with its level in the control group. At the same time, there were no significant changes (P<0.05) for the second treatment group compared with the control group. The average concentration of sperms taken from the caudal epididymis did not change significantly (P<0.05) for both treatments compared to the control group. The percentage of motile sperm decreased significantly (P<0.05) as a result of treatment with the phenolic extract of Mentha spicata leaves, and this percentage is directly proportional to the increase in the doses used of the extract. Sperm vitality compared to the control group, as for the percentage leading to abnormal sperms, the results showed a significant increase (P<0.05) in the average percentage of abnormal sperms in the caudal epididymis of the second treatment animals when compared with the rates of the first and control groups, while there was no There were no significant changes (P<0.05) in the rate of the first treatment compared to the control. Histologically, the results indicate that treatment with phenolic extract of Mentha spicata leaves caused simple histological changes, represented by a simple vacuole of the interstitial layer between the tubules, slight congestion and partial detachment of the germ cells from the basement membrane with a thick fibrous capsule surrounding the epididymis tubes and congestion of blood vessels. In contrast, the seminal tubules and seminal lumen remained normal. This compared to the control group, which showed the semi- Bionatura http://dx.doi.org/10.21931/RB/CSS/2023.08.03.48 2 niferous tubules, seminal cavity and germ layers commonly with normal spermatozoa stages. Keywords: Phenolic extract, Mentha spicata, Fertility

Intergenerational effects on fertility in male and female mice after chronic exposure to environmental doses of NSAIDs and 17α-ethinylestradiol mixtures

Non-steroidal anti-inflammatory drugs (NSAIDs) and 17α-ethinylestradiol (EE2) are extensively used in human and veterinary medicine. Due to their partial removal by wastewater treatment plants, they are frequent environmental contaminants, particularly in drinking water. Here, we investigated the adverse outcomes of chronic exposure to mixtures of NSAIDs (ibuprofen, 2hydroxy-ibuprofen, diclofenac) and EE2 at two environmentally relevant doses in drinking water, on the reproductive organ development and fertility in F1-exposed male and female mice and in their F2 offspring. In male and female F1 mice, which were exposed to these mixtures, reproductive organ maturation, estrous cyclicity, and spermiogenesis were altered. These defects were observed also in F2 animals, in addition to some specific sperm parameter alterations in F2 males. Transcriptomic analysis revealed significant changes in gene expression patterns and associated pathways implicated in testis and ovarian physiology. Chronic exposure of mice to NSAID and EE2 mixtures at environmental doses intergenerationally affected male and female fertility (i.e. total number of pups and time between litters). Our study provides new insights into the adverse effects of these pharmaceuticals on the reproductive health and will facilitate the implementation of a future regulatory environmental risk assessment of NSAIDs and EE2 for human health.

Common as well as unique methylation-sensitive DNA regulatory elements in three mammalian SLC9C1 genes

The SLC9C1 gene (which encodes the NHE10 protein) is essential for male fertility in both mice and humans, however the epigenetic mechanisms regulating its testis/sperm-specific gene expression have yet to be studied. Here we identify and characterize DNA regulatory elements of the SLC9C1 gene across three mammalian species: mouse, rat, and human. First, in silico analysis of these mammalian SLC9C1 genes identified a CpG island located upstream of the transcription start site in the same relative position in all three genes. Further analysis reveals that this CpG island behaves differently, with respect to gene regulatory activity, in the mouse SLC9C1 gene than it does in the rat and human SLC9C1 gene. The mouse SLC9C1 CpG island displays strong promoter activity by itself and seems to have a stronger gene regulatory effect than either the rat or human SLC9C1 CpG islands. While the function of the upstream SLC9C1 CpG island may be divergent across the three studied species, it appears that the promoters of these three mammalian SLC9C1 genes share similar DNA methylation-sensitive regulatory mechanisms. All three SLC9C1 promoter regions are differentially methylated in lung and testis, being more hypermethylated in lung relative to the testis, and DNA sequence alignments provide strong evidence of primary sequence conservation. Luciferase assays reveal that in vitro methylation of constructs containing different elements of the three SLC9C1 genes largely exhibit methylation-sensitive promoter activity (reduced promoter activity when methylated) in both HEK 293 and GC-1spg cells. In total, our data suggest that the DNA methylation-sensitive elements of the mouse, rat, and human SLC9C1 promoters are largely conserved, while the upstream SLC9C1 CpG island common to all three species seems to perform a different function in mouse than it does in rat and human. This work provides evidence that while homologous genes can all be regulated by DNA methylation-dependent epigenetic mechanisms, the location of the specific cis-regulatory elements responsible for this regulation can differ across species.

Na+/H+ Exchangers (NHEs) in Mammalian Sperm: Essential Contributors to Male Fertility.

Na+/H+ exchangers (NHEs) are known to be important regulators of pH in multiple intracellular compartments of eukaryotic cells. Sperm function is especially dependent on changes in pH and thus it has been postulated that NHEs play important roles in regulating the intracellular pH of these cells. For example, in order to achieve fertilization, mature sperm must maintain a basal pH in the male reproductive tract and then alkalize in response to specific signals in the female reproductive tract during the capacitation process. Eight NHE isoforms are expressed in mammalian testis/sperm: NHE1, NHE3, NHE5, NHE8, NHA1, NHA2, NHE10, and NHE11. These NHE isoforms are expressed at varying times during spermatogenesis and localize to different subcellular structures in developing and mature sperm where they contribute to multiple aspects of sperm physiology and male fertility including proper sperm development/morphogenesis, motility, capacitation, and the acrosome reaction. Previous work has provided evidence for NHE3, NHE8, NHA1, NHA2, and NHE10 being critical for male fertility in mice and NHE10 has recently been shown to be essential for male fertility in humans. In this article we review what is known about each NHE isoform expressed in mammalian sperm and discuss the physiological significance of each NHE isoform with respect to male fertility.

The effect of letrozole treatment on the fertility and embryos parameters in female mice: Histological and hormonal study

Objective: To evaluate how letrozole affects reproductive hormones and embryos in mature female mice.Method: The investigative study was conducted from December 2021 to February 2022 at the Pathology Departmentof Kut Technical Institute, Al-Kut, Iraq, and comprised adult female albino mice weighing 23-25g. The mice were dividedinto control group A, those receiving 1.5mg/kg/day letrozole in group B, and those receiving 2.5mg/kg/day letrozolein group C. The intervention lasted 14 days. Blood samples were collected under anaesthesia by heart puncture forhormonal testing. Data was analysed using SPSS 15.Results: Of the 30 mice, 10(33.3%) were in each of the 3 groups. Right and left ovaries of mice in groups B and Cshowed a significant rise in multi-follicular cells compared to group A (p<0.05). Follicle-stimulating hormone, oestrogenand progesterone concentrations as well as luteinizing hormone increased significantly in the intervention groupscompared to the control group (p<0.05). The impact of letrozole on embryos was significant in groups B and Ccompared to group A.Conclusions: Letrozole was found to have a positive effect on reproductive parameters and hormones in mice.Keywords: Mice, Ovary, Letrozole, oestrogen, Progesterone, Luteinizing hormone.

SAT362 Low-dose DHT’s Effects On Gluconeogenesis And Energy Metabolism Are Ameliorated By Central Nervous System Knockout Of The Androgen Receptor

Abstract Disclosure: V. Ubba: None. S. joseph: None. R. Akbar: None. M. Dsilva: None. S. Wu: None. Polycystic ovarian syndrome (PCOS) is a leading cause of infertility in women of reproductive age. Mostly, the level of androgens are deviated from their physiological values in females, resulting in hyperandrogenemia. Androgen manifests its function in the cells via the Androgen Receptor (AR), making it a prime factor for studying PCOS pathophysiology. We generated central nervous system (CNS) specific AR knockout mouse (SynARKO), using synapsin promoter driven cre, to examine its role under normal and hyperandrogenemic conditions in female mice. Assessment of reproductive parameters under physiological levels of androgens, revealed no differences in the cyclicity and fertility of SynARKO mice when compared to Control (Con-veh) mice. These parameters were altered under hyperandrogenemia (induced using DHT) in Control mice (Con-DHT) mice and were not rescued in the SynARKO- DHT mice, suggesting CNS AR is dispensable to reproductive features in females. Evaluation of metabolic functions by Glucose tolerance test, we observed alleviated glucose intolerance in SynARKO-DHT (4 months). Energy expenditure and food intake were increased in Con-DHT mice compared to Con-veh, while showing no further differences in SynARKO-DHT mice compared to SynARKO-veh. At the molecular level, DHT treatment showed increased phosphorylated Akt in hypothalamus of control mice. Interestingly, it was restored in synARKO upon DHT treatment. Further, the inflammation marker, phosphorylated NF-kB p-65, was increased in the hypothalamus of Con-DHT mice. Additionally, IBA, a marker of macrophage activation was also increased upon DHT treatment in control mice. Taken together, our results suggest that while the reproductive functions are not mediated by the CNS AR, metabolic effects caused by hyperandrogenemia, is regulated by the central nervous system AR. Presentation Date: Saturday, June 17, 2023

Targeted mutation and inactivation of the kinesin light chain 3 protein-encoding gene have no impact on mouse fertility†.

The kinesin light chain 3 protein (KLC3) is the only member of the kinesin light chain protein family that was identified in post-meiotic mouse male germ cells. It plays a role in the formation of the sperm midpiece through its association with both spermatid mitochondria and outer dense fibers (ODF). Previous studies showed a significant correlation between its expression level and sperm motility and quantitative semen parameters in humans, while the overexpression of a KLC3-mutant protein unable to bind ODF also affected the same traits in mice. To further assess the role of KLC3 in fertility, we used CRISPR/Cas9 genome editing in mice and investigated the phenotypes induced by the invalidation of the gene or of a functional domain of the protein. Both approaches gave similar results, i.e. no detectable change in male or female fertility. Testis histology, litter size and sperm count were not altered. Apart from the line-dependent alterations of Klc3 mRNA levels, testicular transcriptome analysis did not reveal any other changes in the genes tested. Western analysis supported the absence of KLC3 in the gonads of males homozygous for the inactivating mutation and a strong decrease in expression in males homozygous for the allele lacking one out of the five tetratricopeptide repeats. Overall, these observations raise questions about the supposedly critical role of this kinesin in reproduction, at least in mice where its gene mutation or inactivation did not translate into fertility impairment.

Zika virus infection leads to hormone deficiencies of the hypothalamic-pituitary-gonadal axis and diminished fertility in mice.

Zika virus (ZIKV) infection in pregnant women during the third trimester can cause neurodevelopmental delays and cryptorchidism in children without microcephaly. However, the consequences of congenital ZIKV infection on fertility in these children remain unclear. Here, using an immunocompetent mouse model, we reveal that congenital ZIKV infection can cause hormonal disorders of the hypothalamic-pituitary-gonadal axis, leading to reduced fertility and decreased sexual preference. Our study has for the first time linked the hypothalamus to the reproductive system and social behaviors after ZIKV infection. Although the extent to which these observations in mice translate to humans remains unclear, these findings did suggest that the reproductive health and hormone levels of ZIKV-exposed children should receive more attention to improve their living quality.