The role of immune cell death in spermatogenesis and male fertility.

The male reproductive system provides a distinctive shield to the immune system, safeguarding germ cells (GCs) from autoimmune harm. The testis in mammals creates a unique immunological setting due to its exceptional immune privilege and potent local innate immunity. which can result from a number of different circumstances, including disorders of the pituitary gland, GC aplasia, and immunological elements. Apoptosis, or programmed cell death (PCD), is essential for mammalian spermatogenesis to maintain and ensure an appropriate number of GCs that correspond with the supporting capability of the Sertoli cells. Apoptosis is substantial in controlling the number of GCs in the testis throughout spermatogenesis, and any dysregulation of this process has been linked to male infertility. There is a number of evidence about the potential of PCD in designing novel therapeutic approaches in the treatment of infertility. A detailed understanding of PCD and the processes that underlie immunological infertility can contribute to the progress in designing strategies to prevent and treat male infertility. This review will provide a summary of the role of immune cell death in male reproduction and infertility and describe the therapeutic strategies and agents for treatment based on immune cell death.

Update on the research on the antigens of anti-sperm antibodies over the last decade.

This review summarizes the advancements over a decade of research on antigens of anti-sperm antibodies (ASAs), which are key to male immune infertility. Despite the progress in assisted reproductive technologies, understanding the roles and mechanisms of ASAs and their antigens remains vital for immune infertility management. We conducted a comprehensive literature search on PubMed from January 2013 to December 2023 using the following keywords: "anti-sperm antibody," "sperm antigen," and "immune infertility." In this review, we focus on the discoveries in sperm antigen identification and characterization through proteomics, gene disruption technology, and immunoinformatics, along with the development of fertility biomarkers. Here, we discuss the clinical applications of improved ASA detection methods and the progress in the development of immunocontraceptive vaccines. The intersection of advanced diagnostic techniques and vaccine development represents a promising frontier in reproductive health. The findings also highlight the need for standardized ASA detection methods and a comprehensive molecular-level approach to understanding ASA-related infertility. These insights underscore the significance of ongoing reproductive immunology research in enhancing clinical fertility outcomes and contraceptive vaccine development.

A comprehensive insight into the contribution of epigenetics in male infertility; focusing on immunological modifications.

Numerous recent studies have examined the impact epigenetics-including DNA methylation-has on spermatogenesis and male infertility. Differential methylation of several genes has been linked to compromised spermatogenesis and/or reproductive failure. Specifically, male infertility has been frequently associated with DNA methylation abnormalities of MEST and H19 inside imprinted genes and MTHFR within non-imprinted genes. Microbial infections mainly result in male infertility because of the immune response triggered by the bacteria' accumulation of immune cells, proinflammatory cytokines, and chemokines. Thus, bacterially produced epigenetic dysregulations may impact host cell function, supporting host defense or enabling pathogen persistence. So, it is possible to think of pathogenic bacteria as potential epimutagens that can alter the epigenome. It has been demonstrated that dysregulated levels of LncRNA correlate with motility and sperm count in ejaculated spermatozoa from infertile males. Therefore, a thorough understanding of the relationship between decreased reproductive capacity and sperm DNA methylation status should aid in creating new diagnostic instruments for this condition. To fully understand the mechanisms influencing sperm methylation and how they relate to male infertility, more research is required.

Anti-tubulin beta 4A (TBB4A) antibody immobilized sperm in a complement-dependent manner in humans.

Sperm-immobilizing antibodies (SI-Abs) are detected in the sera of 3 % of infertile women. SI-Abs are occasionally produced as allogeneic antibodies against sperm, causing immune infertility. SI-Abs inhibit the passage of sperm through the female reproductive tract. Research on anti-sperm antibodies (ASA) remains of great importance for population control. We aimed to identify the antigens recognized by SI-Abs and elucidate the pathogenesis of immune infertility. Twelve sperm-immobilization test (SIT)-positive and fourteen SIT-negative sera were analyzed by two-dimensional electrophoresis and western blotting. Antigenic materials were extracted from well-motile sperm prepared using 0.1 % sodium dodecyl sulfate. In total, 22 different spots were detected in the 12 positive sera. Among these, three positive serum samples showed two positive signals with similar migration patterns. The significant positive spots were Mr: 49 K, pI: 5.1 and Mr: 51 K, pI: 5.6. All these positive spots were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS); tubulin beta-4A (TBB4A) was identified from the spot Mr: 49 K, pI: 5.1. TBB4A is a major component of tubulin and constitutes the axoneme in the sperm tail and the centrosome in the sperm neck; it is generally located inside the cell. An authentic antibody against TBB4A showed a positive reaction in the sperm neck and tail regions in an immunofluorescence study. This antibody also inhibited sperm motility in a complement-dependent manner. Sperm membrane permeability reportedly changes during swimming and capacitation. We identified TBB4A as an antigenic molecule recognized by SI-Abs, which may be relevant to immunological contraception in the future.

The interlink between thyroid autoimmunity and type 1 diabetes and the impact on male and female fertility.

The aim of this review is to discuss the several interconnections between thyroid autoimmunity and type 1 diabetes in terms of epidemiology, immunoserology, genetic predisposition, and pathogenic mechanisms. We will also analyze the impact of these conditions on both male and female fertility. A literature search was carried out using the MEDLINE/PubMed, Scopus, Google Scholar, ResearchGate, and Clinical Trials Registry databases with a combination of keywords. It was found that the prevalence of thyroid autoantibodies in individuals with type 1 diabetes (T1DM) varied in different countries and ethnic groups from 7 to 35% in both sexes. There are several types of autoantibodies responsible for the immunoserological presentation of autoimmune thyroid diseases (AITDs) which can be either stimulating or inhibiting, which results in AITD being in the plus phase (thyrotoxicosis) or the minus phase (hypothyroidism). Different types of immune cells such as T cells, B cells, natural killer (NK) cells, antigen presenting cells (APCs), and other innate immune cells participate in the damage of the beta cells of the islets of Langerhans, which inevitably leads to T1D. Multiple genetic and environmental factors found in variable combinations are involved in the pathogenesis of AITD and T1D. In conclusion, although it is now well-known that both diabetes and thyroid diseases can affect fertility, only a few data are available on possible effects of autoimmune conditions. Recent findings nevertheless point to the importance of screening patients with immunologic infertility for AITDs and T1D, and vice versa.

The enigmatic interplay of immune cells and abnormal spermatozoa through Mendelian randomization.

PURPOSE: Abnormal spermatozoa significantly impact reproductive health, affecting fertility rates, potentially prolonging conception time, and increasing the risk of miscarriages. This study employs Mendelian randomization to explore their potential link with immune cells, aiming to reveal their potential causal association and wider implications for reproductive health. METHODS: We conducted forward and reverse Mendelian randomization analyses to explore the potential causal connection between 731 immune cell signatures and abnormal spermatozoa. Using publicly available genetic data, we investigated various immune signatures such as median fluorescence intensities (MFI), relative cell (RC), absolute cell (AC), and morphological parameters (MP). Robustness was ensured through comprehensive sensitivity analyses assessing consistency, heterogeneity, and potential horizontal pleiotropy. The MR study produced a statistically significant p-value of .0000684, Bonferroni-corrected for the 731 exposures. RESULTS: The Mendelian randomization analysis revealed strong indications of a reciprocal relationship between immune cell pathways and sperm integrity. When examining immune cell exposure, a potential causal link with abnormal sperm was observed in 35 different types of immune cells. Conversely, the reverse Mendelian randomization results indicated that abnormal sperm might causally affect 39 types of immune cells. These outcomes suggest a potential mutual influence between alterations in immune cell functionality and the quality of spermatozoa. CONCLUSION: This study highlights the close link between immune responses and sperm development, suggesting implications for reproductive health and immune therapies. Further research may offer crucial insights into male fertility and immune disorders.

Associations between cytokines and the risk of female and male infertility: A two-sample Mendelian randomization analysis.

PURPOSE: Observational studies have linked cytokines to the occurrence of female and male infertility. However, it is not clear how biomarkers of inflammation are causally related to infertility. To explore whether genetic variants in circulating cytokines are associated with the pathogenesis of infertility, we performed two-sample Mendelian randomization (MR) analysis. METHODS: A total of 31,112 individuals of European ancestry were included in a genome-wide association study (GWAS) of 47 circulating cytokines as instrumental variables (IVs). Outcome data were female infertility, including four different subtypes, and male infertility, from the FinnGen consortium. Five MR methods, including inverse-variance weighted (IVW), MR-Egger, simple mode, weighted median, and weighted mode were employed to examine the genetic association between cytokines and the risk of female infertility and male infertility. The false discovery rate (FDR) was controlled using the Benjamini-Hochberg method. RESULTS: After FDR correction, cis-protein quantitative trait locus (cis-pQTL) instruments showed that the cytokines GROa and MCSF were positively associated with female infertility. In analyses of subtypes of female infertility, eotaxin and sICAM were inversely associated with ovulation-related infertility; MCP3 alone was positively associated with uterus-related infertility; GROa and MCSF were positively correlated with infertility of cervical, vaginal, and other or unspecified origin; and MIP1a was negatively correlated with tubal origin infertility. The cytokines HGF, IL-2ra, and RANTES were positively correlated with male infertility. Similar findings were obtained in sensitivity analyses. There was no evidence of pleiotropy or heterogeneity in the results. CONCLUSION: These findings contribute to current understanding of the role of cytokine biomarkers in the etiology of female and male infertility. Furthermore clinical experimental validation is required to evaluate the potential of these cytokines as biomarkers.

Acute murine-betacoronavirus infection impairs testicular steroidogenesis and the quality of sperm production.

Although several testicular alterations promoted by coronavirus infection have been demonstrated, the extent, causes, and players of testicular pathogenesis are not totally understood. The present study aimed to investigate the short-term effects on male fertility of intranasally administered murine hepatitis virus strain 3 (MHV-3), a member of the genus Betacoronavirus, which causes a severe systemic acute infection. This mouse model might be used as a in vivo prototype for investigating the impact of betacoronavirus on the endocrine and exocrine testicular functions with the advantage to be performed in a biosafety level 2 condition. Herein, we performed virological, histopathological, and molecular studies regarding the testicular spermatogenesis and the spermatic quality analyses in an MHV-3-infected C57BL/6 mice. The main outcomes showed that MHV-3 infects mouse testis and induces a testicular inflammatory state, impairing the steroidogenic pathway. The infection led to several alterations in the testicular parenchyma, such as: seminiferous epithelium sloughing, retention of residual bodies, germ cell apoptosis, alterations in intercellular junction proteins, and worse spermatogenic parameters. Moreover, the levels of plasmatic testosterone as well as the quality of sperm production reduced. Therefore, the present data suggest that the viral/inflammatory impairment of the steroidogenic pathway and the consequent imbalance of androgen levels is critical in testicular pathology, disturbing the SC barrier function and the germ cell differentiation. Our study is important for comprehending the effects of beta coronavirus infections on testis function in order to develop treatments that could prevent virus-mediated male infertility.

Influence of inguinal hernia repair on sperm autoimmunity: The largest single center experience.

BACKGROUND: Studies on immunological infertility after inguinal hernia correction are few and not very representative. Anti-sperm antibodies have been shown to reduce male fertility. Although the extent of infertility due to anti-sperm antibodies alone is not very clear, data indicates that about 8%-10% of infertile patients have immunological infertility DESIGN: This retrospective study includes all infertile male patients (n = 2258) who underwent mixed antiglobulin reaction tests and urologic examination from 2000 to 2020. Sperm quality (assessed by the number of spermatozoa, their motility, vitality, and normal form) was also evaluated. Among these patients, 191 had previously undergone unilateral or bilateral inguinal hernia surgery repair. The aim of the study is to evaluate if there is a higher incidence of positive mixed antiglobulin reaction test among patients undergoing inguinal hernioplasty compared to the unselected infertile population. RESULTS: Anti-sperm antibodies would seem to increase in both patients who performed general andrological surgery and groin hernia correction, respectively 3.48 (95% Confidence Interval: 1.70-7.10; p < 0.001) and 2.45 (95% Confidence Interval: 1.01-5.99; p < 0.05) times more than the unselected infertile population. CONCLUSIONS: Mixed antiglobulin reaction test could be useful in patients undergone previous scrotal surgery or hernia correction men, to avoid false unexplained infertility diagnoses and to direct the couple to assisted reproductive technology procedures. Basal evaluation of spermatozoa does not actually consider andrological surgery as an indication to autoimmunity investigation.

Varicocele-Mediated Male Infertility: From the Perspective of Testicular Immunity and Inflammation.

Background: Varicocele (VC) is present in 35 - 40% of men with infertility. However, current surgical and antioxidant treatments are not completely effective. In addition to oxidative stress, it is likely that other factors such as testicular immune microenvironment disorder contribute to irreversible testicular. Evidence suggests that VC is associated with anti-sperm antibodies (ASAs), spermatogenesis and testosterone secretion abnormalities, and testicular cytokine production. Moreover, inhibition of inflammation can alleviate VC-mediated pathogenesis. The normal function of the testis depends on its immune tolerance mechanism. Testicular immune regulation is complex, and many infectious or non-infectious diseases may damage this precision system. Results: The testicular immune microenvironment is composed of common immune cells and other cells involved in testicular immunity. The former includes testicular macrophages, T cells, dendritic cells (DCs), and mast cells, whereas the latter include Leydig cells and Sertoli cells (SCs). In animal models and in patients with VC, most studies have revealed an abnormal increase in the levels of ASAs and pro-inflammatory cytokines such as interleukin (IL)-1 and tumor necrosis factor (TNF)-alpha in the seminal plasma, testicular tissue, and even peripheral blood. It is also involved in the activation of potential inflammatory pathways, such as the nucleotide-binding oligomerization domain-like receptor family pyrin domain containing (NLRP)-3 pathway. Finally, the development of VC-mediated infertility (VMI) may be facilitated by abnormal permeability of proteins, such as claudin-11, that constitute the blood-testis barrier (BTB). Conclusions: The testicular immune response, including the production of ASAs and inflammatory factors, activation of inflammatory pathways, and destruction of the BTB may be involved in the pathogenesis of VMI it is necessary to further explore how patient outcomes can be improved through immunotherapy.

Probiotic based-diet effect on the immune response and induced stress in irradiated mass reared Ceratitis capitata males (Diptera: Tephritidae) destined for the release in the sterile insect technique programs.

Ceratitis capitata (medfly) is one of the most devastating crop pests worldwide. The Sterile Insect Technique (SIT) is a control method that is based on the mass rearing of males, their sterilization, and release in the field. However, the effectiveness of the technique depends on the quality of the released males and their fitness. We previously isolated and selected a probiotic bacteria (Enterobacter sp.), from wild-caught medflies, according to criteria that improved biological quality traits of reared medfly males.We firstly evaluated the impact of the irradiation on the expression of different immune and stress genes in the medfly sterile males. Expression was measured at differents time points ranging from 0 to 168 h after irradiation to capture the response of genes with distinct temporal expression patterns. Then, we supplemented the larval diet with previously isolated Enterobacter sp.strain, live and autoclaved at various concentrations to see whether the probiotic treatments affect, through their protective role, the gene expression level, and quality traits. The irradiation had significant effect on the genes attacin, cecropin, PGPR-LC, hsp23, and hsp70 level expression. The expression of attacin and PGPR-LC was up-regulated while that of cecropin was down-regulated. Hsp genes showed decreased levels between 0 and 18 h to peak at 72 h. However, the supplementation of the probiotic strain, either live or autoclaved, was statistically significant only for attacingene. However, significant interaction time x probiotic was noticed for attacin, cecropin, hsp23 and hsp70. The probiotic treatments also improved the quality control parameters like pupal weight. From this work we can conclude that a consortium of parabiotics (autoclaved probiotics) treatment will be recommended in insectaries considering both the beneficial effects on mass reared insects and its general safety for insectary workers and for environment.

Pro-inflammatory cytokines and microRNAs in male infertility.

BACKGROUND: Male infertility is a problem that affects 10-15% of men of reproductive age. In particular, gametogenesis is a complex process in which inflammation may play a central role through the secretion of cytokines and the expression of microRNAs. We assessed the potential role of proinflammatory cytokines (TNF-α, IL-6 and IL-1α) and microRNAs (miR-146a-5p, miR-34a-5p and miR-23a-3p) in the seminal plasma of infertile men compared to controls, evaluating their correlation with seminal and biochemical parameters. METHODS AND RESULTS: Expression of cytokines and microRNAs was analyzed by ELISA and q-PCR. Our data shows that IL-1α was significantly increased in the azoospermic group compared to controls, TNF-α mRNA was more expressed in the oligozoospermic group than controls. There were no significant differences in miRNAs expression among the three groups. The correlations between sperm parameters and inflammatory markers were evaluated, however no significance was highlighted. CONCLUSIONS: The determination of each inflammatory marker separately in the seminal plasma of subfertile men, despite some significant differences, does not have a diagnostic value in male infertility even if an assay of selective pro-inflammatory cytokines and microRNAs in the semen may improve the diagnosis of male infertility.

Multiple Lesions Contribute to Infertility in Males Lacking Autoimmune Regulator.

Male factors, including those of autoimmune origin, contribute to approximately 50% of infertility cases in humans. However, the mechanisms underlying autoimmune male infertility are poorly understood. Deficiency in autoimmune regulator (AIRE) impairs central immune tolerance because of diminished expression of self-antigens in the thymus. Humans with AIRE mutations and mice with engineered ablation of Aire develop multiorgan autoimmunity and infertility. To determine the immune targets contributing to infertility in male Aire-deficient (-/-) mice, Aire-/- or wild-type (WT) males were paired with WT females. Aire-/- males exhibited dramatically reduced mating frequency and fertility, hypogonadism, and reduced serum testosterone. Approximately 15% of mice exhibited lymphocytic infiltration into the testis, accompanied by atrophy, azoospermia, and reduced numbers of mitotically active germ cells; the remaining mice showed normal testicular morphology, sperm counts, and motility. However, spermatozoa from all Aire-/- mice were defective in their ability to fertilize WT oocytes in vitro. Lymphocytic infiltration into the epididymis, seminal vesicle, and prostate gland was evident. Aire-/- male mice generated autoreactive antibodies in an age-dependent manner against sperm, testis, epididymis, prostate gland, and seminal vesicle. Finally, expression of Aire was evident in the seminiferous epithelium in an age-dependent manner, as well as in the prostate gland. These findings suggest that Aire-dependent central tolerance plays a critical role in maintaining male fertility by stemming autoimmunity against multiple reproductive targets.

Xiaokang Liuwei Dihuang decoction ameliorates the immune infertility of male rats induced by lipopolysaccharide through regulating the levels of sex hormones, reactive oxygen species, pro-apoptotic and immune factors.

Male immune infertility is a kind of disease that damages family life and happiness. The development of novel methods treating male immune infertility is of great significance. This study aimed to investigate the therapeutic effects of Chinese medicine Xiaokang Liuwei Dihuang decoction on immune infertility of male rats and explored the involved mechanisms. Model rats were established by lipopolysaccharide (LPS) injection. Anti-sperm antibody (AsAb) was detected by ELISA assay and testicular cell apoptosis was evaluated by TUNEL staining to verify the successful model establishment and screen suitable doses of Xiaokang Liuwei Dihuang decoction. Thirty rats were then divided into five groups (n = 6 per group): Control, LPS, Xiaokang Liuwei Dihuang decoction (15.12 g/kg, 30.24 g/kg and 45.36 g/kg). Results of HE staining showed that compared with LPS group, Xiaokang Liuwei Dihuang decoction treatments gradually improved the morphology of seminiferous tubules and elevated the number of spermatogenic cells as the doses increased. The sperm number and the levels of testosterone, luteinizing hormone (LH) and follicle stimulating hormone (FSH) in the serum of 15.12 g/kg, 30.24 g/kg and 45.36 g/kg Xiaokang Liuwei Dihuang decoction groups were much higher than those in LPS group. Results of TUNEL staining, ELISA assay and western blot showed that compared with LPS group, the testicular cell apoptosis and the levels of interleukin 1ß (IL-1ß), tumor necrosis factor α (TNF-α), AsAb, malondialdehyde (MDA) and toll-like receptor 2 (TLR2) in the testicular tissue significantly decreased in three Xiaokang Liuwei Dihuang decoction groups. Compared with LPS group, Bax expression in the 30.24 g/kg and 45.36 g/kg Xiaokang Liuwei Dihuang decoction groups was significantly down-regulated as well. In conclusion, Xiaokang Liuwei Dihuang decoction might ameliorate the immune infertility of male rats induced by LPS through regulating the levels of sex hormones, reactive oxygen species, pro-apoptotic and immune factors.

Evaluation of multi-epitope recombinant protein as a candidate for a contraceptive vaccine.

Contraceptive vaccine (CV) is a valuable, non-invasive, and alternative method for purposeful contraception. Sperm antigens are useful targets for producing CVs due to their specialized expression in sperm. In this study, a recombinant protein containing three main sperm epitopes (IZUMO1, SACA3, and PH-20) was designed and evaluated as CV to control fertility in male mice. The chimeric recombinant protein was expressed and purified in E. coli. Male mice were immunized by 100 µg purified protein and sera were collected to assess IgG antibodies. Evaluating the reproductive performance, immunized male mice mated with normal-fertile female mice and mating rate and the number of newborns was studied. Immunized mice were sacrificed and necropsy and histopathology studies were conducted. The results revealed that the designed chimeric protein stimulated the immune system of the mice effectively. The level of IgG antibody was significantly higher in vaccinated mouse rather than control mouse. Eighty percent of the vaccinated mice became infertile and in the remaining ones, the number of children decreased to 4-6 offspring instead of 10-12 in normal mice. Histopathological studies showed that no organs including heart, brain, lung, liver, kidney and intestine were damaged. However, Normal spermatogenesis has been disrupted and necrotic spermatogonia cells were reported in Seminiferous tubules. We concluded that the designed chimeric protein containing IZUMO1, SACA3, and PH-20 epitopes can stimulate the immune system and cause male contraception without any side effects.

Heat shock protein A4L is a potent autoantigen for testicular autoimmunity in mice.

Experimental autoimmune orchitis (EAO) may be used as a model to investigate immunological infertility in men. Murine EAO is induced via immunization with auto-immunogenic antigens (AIAgs) from testicular germ cells (TGCs). CD4 + T cells play a crucial role in EAO induction. However, whether AIAgs induce an immune response remains unclear. We aimed to identify self-antigens that induce EAO by screening a phage display library of random TGC peptides using IgG from EAO-induced A/J mice. Twenty TGC-specific AIAgs were detected, and G protein-coupled receptor kinase 2 interacting protein-1 (GIT1) and heat shock protein A4L (HSPA4L) were identified as candidate AIAgs that induce EAO. Immunization with GIT1 or HSPA4L, emulsified in complete Freund's adjuvant, resulted in 66 % or 100 % incidence of EAO, respectively, indicating that HSPA4L is a most potent AIAg that induces EAO in mice. These findings may expectedly help improve the diagnostic procedures and treatment of immunological infertility in men.

Mumps Orchitis: Clinical Aspects and Mechanisms.

The causative agent of mumps is a single-stranded, non-segmented, negative sense RNA virus belonging to the Paramyxoviridae family. Besides the classic symptom of painfully swollen parotid salivary glands (parotitis) in mumps virus (MuV)-infected men, orchitis is the most common form of extra-salivary gland inflammation. Mumps orchitis frequently occurs in young adult men, and leads to pain and swelling of the testis. The administration of MuV vaccines in children has been proven highly effective in reducing the incidence of mumps. However, a recent global outbreak of mumps and the high rate of orchitis have recently been considered as threats to male fertility. The pathogenesis of mumps orchitis remains largely unclear due to lack of systematic clinical data analysis and animal models studies. The alarming increase in the incidence of mumps orchitis and the high risk of the male fertility have thus become a major health concern. Recent studies have revealed the mechanisms by which MuV-host cells interact and MuV infection induces inflammatory responses in testicular cells. In this mini-review, we highlight advances in our knowledge of the clinical aspects and possible mechanisms of mumps orchitis.

Transient systemic inflammation in adult male mice results in underweight progeny.

PROBLEM: While the testes represent an immune-privileged organ, there is evidence that systemic inflammation is accompanied by local inflammatory responses. We therefore examined whether transient systemic inflammation caused any inflammatory and functional consequences in murine testes. METHOD OF STUDY: Using a single systemic administration of Toll-like receptor (TLR) agonists [lipopolysaccharide (LPS) or peptidoglycan (PG) or polyinosinic-polycytidylic acid (polyIC)] in young adult male mice, we assessed testicular immune-inflammatory landscape and reproductive functionality. RESULTS: Our findings demonstrated a significant induction of testicular TNF-α, IL-1ß and IL-6 transcripts within 24 h of TLR agonist injection. By day 6, these cytokine levels returned to baseline. While there was no change in caudal sperm counts at early time points, eight weeks later, twofold decrease in sperm count and reduced testicular testosterone levels were evident. When these mice were subjected to mating studies, no differences in mating efficiencies or litter sizes were observed compared with controls. Nonetheless, the neonatal weights of progeny from LPS/PG/polyIC-treated sires were significantly lower than controls. Postnatal weight gain up to three weeks was also slower in the progeny of LPS/polyIC-treated sires. Placental weights at 17.5 days post-coitum were significantly lower in females mated to LPS- and polyIC-treated males. Given this likelihood of an epigenetic effect, we found lower testicular levels of histone methyltransferase enzyme, mixed-lineage leukaemia-1, in mice given LPS/PG/polyIC 8 weeks earlier. CONCLUSION: Exposure to transient systemic inflammation leads to transient local inflammation in the testes, with persistent sperm-mediated consequences for foetal development.

Three-Dimensional Confocal Analysis of Chromosome Positioning Coupled with Immunofluorescence in Mouse Sperm Nuclei.

Male infertility is associated with several causes affecting the paternal nucleus such as DNA lesions (breaks, deletions, mutations, ...) or numerical chromosome anomalies. More recently, male infertility has also been associated with changes in the sperm epigenome, including modification in the topology of chromatin (Olszewska et al., Chromosome Research 16:875-890, 2008; Alladin et al., Syst Biol Reprod Med 59: 146-152, 2013) ref with number 1, 2. Indeed, the positioning of chromosomes in the sperm nucleus is nonrandom and defines chromosome territories (Champroux et al., Genes (Basel) 9:501, 2018) ref with number 3 whose optimal organization determines the success of embryonic development. In this context, the study of the spatial distribution of chromosomes in sperm cells could be relevant for clinical diagnosis. We describe here a in situ fluorescence hybridization (FISH) strategy coupled with a fluorescent immunocytochemistry approach followed by confocal analysis and reconstruction (2D/3D) as a powerful tool to analyze the location of chromosomes in the sperm nucleus using the mouse sperm as a model. Already, the two-dimensional (2D) analysis of FISH and immunofluorescence data reveal the location of chromosomes as well as the different markings on the spermatic nucleus. In addition, a good 3D rendering after Imaris software processing was obtained when Z-stacks of images were acquired over a defined volume (10 µm × 13 µm × 15 µm) with a sequential scanning mode to minimize bleed-through effects and avoid overlapping wavelengths.

Two populations of self-maintaining monocyte-independent macrophages exist in adult epididymis and testis.

Macrophages are the principal immune cells of the epididymis and testis, but their origins, heterogeneity, development, and maintenance are not well understood. Here, we describe distinct populations of epididymal and testicular macrophages that display an organ-specific cellular identity. Combining in vivo fate-mapping, chimeric and parabiotic mouse models with in-depth cellular analyses, we found that CD64hiMHCIIlo and CD64loMHCIIhi macrophage populations of epididymis and testis arise sequentially from yolk sac erythro-myeloid progenitors, embryonic hematopoiesis, and nascent neonatal monocytes. While monocytes were the major developmental source of both epididymal and testicular macrophages, both populations self-maintain in the steady-state independent of bone marrow hematopoietic precursors. However, after radiation-induced macrophage ablation or during infection, bone marrow-derived circulating monocytes are recruited to the epididymis and testis, giving rise to inflammatory macrophages that promote tissue damage. These results define the layered ontogeny, maintenance and inflammatory response of macrophage populations in the male reproductive organs.