Epidemiological relationships between pesticide use and male infertility have been suggested for a long time. Etoxazole (ETX), an oxazoline pesticide, has been extensively used for pest eradication. It is considered relatively safe and has low mammalian toxicity because it specifically inhibits chitin synthesis. However, ETX may have toxic effects on the reproductive system. In this study, we examined the effects of ETX on the reproductive system using mouse testis cell lines (TM3 for Leydig cells and TM4 for Sertoli cells) and C57BL/6 male mice. We confirmed that ETX has anti-proliferative effects on the TM3 and TM4 cell lines. Moreover, ETX induced mitochondrial dysfunction and hampers calcium homeostasis. Western blot analysis of MAPK and Akt signaling cascades was performed to demonstrate the mode of action of ETX at a molecular level. Moreover, ETX induced misregulation of genes related to testicular function. Upon oral administration of ETX in C57BL/6 male mice, testis weight was reduced and transcriptional expression related to testis function was altered. These results indicate that ETX induces testicular toxicity by inducing mitochondrial dysfunction and calcium imbalance and regulating gene expression.

Propyl gallate (propyl 3,4,5-trihydroxybenzoate, PG) is a phenolic antioxidant that has been used for oil-containing foods to prevent acidification. Owing to its antioxidant properties, PG has been applied to various fields and active research is currently underway to prove PG as an anticancer agent. However, there are still concerns about PG as a possible reproductive toxicant. Therefore, we determined whether PG induced male infertility. Our results indicated that PG induced testicular dysfunction in both Leydig and Sertoli cells via suppression of cell viability and steroidogenesis. These normal testis functions were destroyed by PG-induced mitochondrial dysfunction and calcium homeostasis dysregulation. In addition, PG disrupted the expression of several genes associated with the testis function and induced endoplasmic reticulum stress. Furthermore, we verified PG-induced mRNA expression changes in steroidogenesis enzymes and hormone receptors in vitro and in vivo. From the results of the qPCR analysis, we further confirmed the PG-mediated reduction in the mRNA expression of genes related to testis functions by in situ hybridization. Finally, we demonstrated that PG induced testicular toxicity via the disruption of mitochondrial or ER function and the inhibition of testicular development-related genes in mice.

Exposure to endocrine disruptors such as bisphenols, can lead to and be the explanation for idiopathic infertility. In our study, we assessed the effect of exposure to bisphenol S (BPS) via breast milk on the testicular tissue health of adult male mice. Milking dams were exposed to BPS through drinking water (0.216 ng g bw/day and 21.6 ng g bw/day) from post-natal day 0–15. Although there was no significant difference in testicular histopathology between the control and experimental groups, we observed an increase in the number of tight and gap junctions in the blood-testis barrier (BTB) of adult mice after nursing BPS exposure. Moreover, there was an increase in oxidative stress markers in adult testicular tissue of mice exposed during nursing. Our nursing model indicates that breast milk is a route of exposure to an endocrine disruptor that can be responsible for idiopathic male infertility through the damage of the BTB and weakening of oxidative stress resistance in adulthood.

Microplastics (MPs), an emerging environmental pollutant, have been clarified to induce testicular disorder in mammals. And the current studies have delineated a correlation between gut microbiota and male reproduction. However, it's still unclear whether gut microbiota gets involved in MPs-induced reproductive toxicity. In this work, we constructed a mouse model drinking 5 μm polystyrene-MPs (PS-MPs) at the concentrations of 100 μg/L and 1000 μg/L for 90 days. Evident histological damage, spermatogenetic disorder and hormones synthesis inhibition were observed in PS-MPs exposed mice. With fecal microbiota transplantation (FMT) trial, the recipient mice exhibited gut microbial alteration, and the elevated abundance of Bacteroides and Prevotellaceae_UCG-001 were positively correlated with testicular disorder according to spearman correlation analysis. Mechanistically, increased proportion of pro-inflammatory bacteria may drive translocation of T helper 17 (Th17) cells, resulting in overproduced interleukin (IL)-17 A and downstream inflammatory response in both the mice exposed to PS-MPs and corresponding recipient mice. In summary, our findings revealed the critical role of gut microbiota in PS-MPs-induced reproductive toxicity, and tried to elucidate the underlying mechanism of gut microbial dysregulation-mediated IL-17 A signaling pathway. Furthermore, this study also provides the research basis for gut microbiota-targeted treatment of male infertility in the future.

Environmental cadmium (Cd) or high-fat diet (HFD) exposure alone are risk factors of male infertility. However, the effect and mechanism of co-exposure to HFD and Cd on sperm quality remain unclear. This study was aimed to explore the combined effects of HFD and Cd on spermatogenesis as well as its m6A-dependent mechanism in vivo and in vitro. As a result, co-exposure of HFD and Cd resulted in a significant decrease in the number of mature testicular seminiferous tubules and epididymis sperm quantity in mice, compared with Cd or HFD exposure alone. Correspondingly, the mRNAs expression of Smc3(spermatocytes marker), Acrv1(round spermatids marker) and Lzumo3(elongated spermatids marker) were downregulated in HFD and Cd group. Furthermore, combined exposure downregulated the expression of meiosis-related proteins (STRA8 and SYCP3), increased the m6A level of Stra8, and upregulated the expression of m6A-related proteins (METTL3 and YTHDF2) in mouse spermatocytes. Mechanistically, the above-mentioned impacts caused by co-exposure were markedly restored by Mettl3 siR and Ythdf2 siR. In addition, RNA stability assay showed that Ythdf2 siR obviously reversed co-exposure-increased Stra8 mRNA degradation rate in actinomycin-D-treated mouse spermatocytes. Meanwhile, excess ROS was observed in combined-exposure group, and a free radical scavenger N-tert-Butyl-α-phenylnitrone (PBN) attenuated co-exposure-upregulated expression of METTL3 and YTHDF2 in mouse spermatocytes. These results suggested that combination of HFD and Cd impaired spermatogenesis by degrading Stra8 in an m6A-YTHDF2-dependent manner via ROS activation.

Male infertility may be caused by genetic and/or environmental factors that impair spermatogenesis and sperm maturation. High-altitude (HA) hypoxic environments represent one of the most serious challenges faced by humans that reside in these areas. To assess the influence of the plateau environment on semen parameters, 2,798 males, including 1,111 native Tibetans and 1,687 Han Chinese individuals living in the plains (HCILP) who underwent pre-pregnancy checkups, were enrolled in this study. The semen samples of males were evaluated to determine conventional sperm parameters, sperm morphology, and sperm movement. Reproductive endocrine hormones (REHs) were detected in 474 males, including 221 Tibetans and 253 HCILP. Due to recurrent abortions in partners, the DNA fragmentation index (DFI) of 133 native Tibetans and 393 HCILP individuals was further compared. Luteinizing hormone (LH) (4.94 ± 2.12 vs. 3.29 ± 1.43 U/L), prolactin (11.34 ± 3.87 vs. 8.97 ± 3.48 nmol/L), E2/T (0.22 ± 0.11 vs 0.11 ± 0.05), median total sperm motility (61.20% vs. 51.56%), and DFI (23.11% vs. 7.22%) were higher in males from plateau areas while median progressive motility (PR) (35.60% vs. 41.12%), total number of PR sperms (51.61 vs. 59.63 mil/ejaculate), percentage of normal form sperms (3.70% vs. 6.00%), curvilinear velocity (36.10 vs. 48.97 μm/s), straight-line (rectilinear) velocity (14.70 vs. 31.52 μm/s), estradiol (103.82 ± 45.92 vs. 146.01 ± 39.73 pmol/L), progesterone (0.29 ± 0.27 vs. 2.22 ± 0.84 nmol/L), testosterone (4.90 ± 1.96 vs. 14.36 ± 5.24 nmol/L), and testosterone secretion index (ratio of testosterone to LH) (33.45 ± 22.86 vs 145.78 ± 73.41) were lower than those in males from the plains. There was no difference in median total sperm number (157.76 vs. 151.65 mil/mL), sperm concentration (52.40 vs. 51.79 mil/mL), volume (3.10 vs. 3.10 mL), total normal form sperms (5.91 vs. 6.58 mil/ejaculate, p50), and follicle-stimulating hormone (FSH) levels (4.13 ± 2.55 U/L vs 3.82 ± 2.35 U/L) between the two groups of males. The REH and sperm parameters of males from HA hypoxic environments were adaptively altered. Although the total number of PR sperm decreased and DFI increased, the Tibetan population that lives at HAs has been found to grown continuously and rapidly. These results supplement prior findings regarding the impact of HA on male reproductive function.

3-Monochloropropane-1,2-diol (3-MCPD) is a food contaminant formed during acid hydrolysis of vegetable proteins. The toxicological evaluation of smaller doses of 3-MCPD is essential for safety evaluation of this compound. The present study investigates the toxicologic potential of 3-MCPD on male genital organs of rats, applies a correlation between the induced infertility and developed lesions in testes, epididymis, and accessory glands and study the possible mechanisms of 3-MCPD-induced male infertility. Forty rats were randomly divided into four main groups of ten animals each: the control untreated group and three treated groups that were orally administered 3-MCPD at different doses (3, 7.5 and 15 mg/kg b.w) daily via stomach intubation for five successive days per week. Five rats from each group were euthanized after 30 days. The remaining rats were euthanized after 90 days to establish subacute and chronic toxicity studies. Oxidative stress markers, Nrf2 gene expression, semen analysis, and histopathological examination were performed at the end of each experimental period. Results indicated that 3-MCPD induces infertility in male rat via disruption of Nrf2 expression in the testicular tissue with subsequent increased oxidative stress indicators in the testis that affect spermatogenesis and induced testicular degeneration, in addition, induction of epididymal lesions that affect sperm motility and concentration and finally possible development of hyperplastic tissue reactions in accessory glands of intoxicated rats predicting the carcinogenic potential of this compound.

Male infertility is considered one of the most critical health problems that are expected to expand worldwide. Ulva lactuca is a species of green seaweeds which is known to be a rich source of many important nutrients. Accordingly, this study is designated to investigate the therapeutic role of Ulva lactuca water fraction (UL) against monosodium glutamate (MSG)-induced male reproductive system disorders in male rats. Ulva lactuca methanolic crude extract was prepared firstly, and then water-dissolved compounds of this crude methanolic extract were separated. Ulva lactuca water fraction active phenolic compounds were determined using high-performance liquid chromatography (HPLC). Thirty-two male rats were divided equally into four groups; male infertility was induced in sixteen experimental animals by MSG at dose of 15 mg/Kg for 45 days. Eight infertile animals were treated with 100 mg/Kg of Ulva lactuca water fraction for 30 days. The rest of the animals were divided into two control groups; one control group (eight animals) was used to study the effect of UL on healthy rats at dose of 100 mg/Kg for 30 days and healthy control group (eight animals). Semen quality parameters (concentration and motility ratio), serum testosterone, prostate-specific antigen (PSA), and phosphatases were estimated by using standard protocols. Moreover, prooxidants and endogenous antioxidant enzymes were measured in prostate and testis homogenates. In addition, relative expression of pro-inflammatory genes (inducible nitric oxide synthase (i-NOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-α alpha (TNF-α), and tumor protein (P53)) were assessed in testicular and prostatic tissues. Finally, histological alterations were measured by hematoxylin and eosin (H&E) stain. Results revealed that Ulva lactuca water fraction contains active phenolic constituents responsible for its antioxidant bioactivity. Oral administration of MSG significantly induced histological alterations. Oxidative stress was observed with elevated levels of nitric oxide (NO), thiobarbituric acid reactive substances (TBARS), and xanthine oxidase (XO) activity in both testis and prostate tissues. MSG adversely affected prostate function via elevation of PSA, prostatic acid phosphatases (PAPs), and total acid phosphatases (TAPs). In addition, it upregulated pro-inflammatory genes in testis and prostate tissues. Meanwhile, MSG reduced serum testosterone, semen quality, and antioxidant enzyme activities (glutathione peroxidase (GPx), glutathione-s-transferase (GST), and superoxide dismutase (SOD)). Treatment with UL notably ameliorated the state of oxidative stress and downregulated the expression of pro-inflammatory gene markers. This study highlighted the potential efficacy of Ulva lactuca water fraction on MSG-induced male infertility in rats. Therapeutic effect of UL on oxidative stress and inflammation induced by MSG in testicular and prostatic tissues.

The current research was constructed to throw the light on the protective possibility of Chlorella vulgaris (C. vulgaris) and Spirulina platensis (S. platensis) against lead acetate-promoted testicular dysfunction in male rats. Forty rats were classified into four groups: (i) control, (ii) rats received lead acetate (30 mg/kg bw), (iii) rats concomitantly received lead acetate and C. vulgaris (300 mg/kg bw), (vi) rats were simultaneously treated with lead acetate and S. platensis (300 mg/kg bw) via oral gavage for 8 weeks. Lead acetate promoted testicular injury as expressed with fall in reproductive organ weights and gonadosomatic index (GSI). Lead acetate disrupted spermatogenesis as indicated by sperm cell count reduction and increased sperm malformation percentage. Lead acetate-deteriorated steroidogenesis is evoked by minimized serum testosterone along with maximized follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels. Testicular oxidative, inflammatory, and apoptotic cascades are revealed by elevated acid phosphatase (ACP) and sorbitol dehydrogenase (SDH) serum leakage, declined testicular total antioxidative capacity (TAC) with elevated total oxidative capacity (TOC), tumor necrosis factor alpha (TNF-α), caspase-3 levels, lessened androgen receptor (AR) expression, and histopathological lesions against control. Our research highlights that C. vulgaris or S. platensis therapy can modulate lead acetate-promoted testicular dysfunction via their antioxidant activity as expressed by elevated TAC and reduced TOC, immunomodulatory effect as indicated by lessened TNF-α level, and anti-apoptotic potential that was revealed by minimized caspase-3 levels. As well as restoration of testicular histoarchitecture, androgen receptor, steroidogenesis, and spermatogenesis were detected with better impacts to S. platensis comparing with C. vulgaris. Therefore, further clinical trials are needed to test S. platensis and C. vulgaris as a promising candidate in treating male infertility.

Retrotransposons, as vital regulator of male fertility, are essential for spermatogenesis. Cadmium (Cd) is an environmental toxicant and endocrine disruptor, targeting the reproductive system. Growing evidence shows that Cd exposure can induce male infertility in mammals. In this study, we generated a male C57BL/6 J mice model with consecutive 35 days cadmium chloride (CdCl₂) in different concentrations of 0, 0.25, 0.5, 1.0, and 2.0 mg/kg. The results indicated that 1.0 and 2.0 mg/kg CdCl₂ significantly affected the body weight. Meanwhile, the highest dose group with 2.0 mg/kg CdCl₂ presented low fertility. Furthermore, the expression of retrotransposon mRNA was markedly increased in the higher doses group. We examined methylcytosine (mC) levels of the three active LINE-1 subfamilies TfI, A, and GfII in testis. Conclusively, Cd exposure probably undermines the male mice fertility by disrupting DNA methylation to regulate the retrotransposons. Further studies are required for identifying whether retrotransposon activation has any significant impacts on genome structure, stability, and expression in Cd-induced testicular injury, laying foundation for the treatment for male infertility.