Reduced binding activity of vaccine serum to omicron receptor-binding domain.

Coronavirus disease 2019 (COVID-19) vaccination regimens contribute to limiting the spread of severe acute respiratory syndrome Coronavirus-2 (SARS-CoV-2). However, the emergence and rapid transmission of the SARS-CoV-2 variant Omicron raise a concern about the efficacy of the current vaccination strategy. Here, we expressed monomeric and dimeric receptor-binding domains (RBDs) of the spike protein of prototype SARS-CoV-2 and Omicron variant in E. coli and investigated the reactivity of anti-sera from Chinese subjects immunized with SARS-CoV-2 vaccines to these recombinant RBDs. In 106 human blood samples collected from 91 participants from Jiangxi, China, 26 sera were identified to be positive for SARS-CoV-2 spike protein antibodies by lateral flow dipstick (LFD) assays, which were enriched in the ones collected from day 7 to 1 month post-boost (87.0%) compared to those harvested within 1 week post-boost (23.8%) (P < 0.0001). A higher positive ratio was observed in the child group (40.8%) than adults (13.6%) (P = 0.0073). ELISA results showed that the binding activity of anti-SARS-CoV-2 antibody-positive sera to Omicron RBDs dropped by 1.48- to 2.07-fold compared to its homogeneous recombinant RBDs. Thus, our data indicate that current SARS-CoV-2 vaccines provide restricted humoral protection against the Omicron variant.

Physiologically detectable bisphenol A impairs human sperm functions by reducing protein-tyrosine phosphorylation.

BACKGROUND: Bisphenol A (BPA), a widely used plastic monomer and plasticizer, is detectable in blood, urine and semen of a healthy people, with concentrations ranging from 0.1 nM to 10 nM. It has been shown that in vitro exposure of BPA as low as 0.001 nM could significantly inhibited mouse sperm motility and acrosome reaction. However, it is still unclear whether BPA at those physiologically detectable concentration affects human sperm. METHODS: The effects of different concentrations of BPA (0, 10-3, 10-2, 10-1, 10, 103 nM) on sperm functions were examined, including human sperm viability, kinematic parameters, hyperactivation and capacitation. RESULTS: BPA caused a remarkable decline in human sperm viability, motility and progressive motility, hyperactivation, capacitation and progesterone-induced acrosome reaction. Mechanism studies showed that BPA could suppress the protein tyrosine phosphorylation level of human sperm, but had no effect on sperm calcium signaling. CONCLUSIONS: Physiologically detectable concentrations of BPA may impair human sperm functions via suppressing protein tyrosine phosphorylation of human sperm, implying that environmental pollution of BPA might be a factor contributing to male infertility.

Novel Japanese encephalitis virus NS1-based vaccine: Truncated NS1 fused with E. coli heat labile enterotoxin B subunit.

BACKGROUND: Current vaccines against Japanese encephalitis virus (JEV) of flaviviruses have some disadvantages, such as the risk of virulent reversion. Non-structural protein NS1 is conserved among flaviviruses and confers immune protection without the risk of antibody-dependent enhancement (ADE). Therefore, NS1 has become a promising vaccine candidate against flaviviruses. METHODS: A NS1-based vaccine (LTB-NS1∆63) with a truncated NS1 protein (NS1∆63) fused to E. coli heat-labile enterotoxin B subunit (LTB) was expressed in E.coli and explored for its ability to induce immune responses. Safety of LTB-NS1∆63 was assessed by determining its toxicity in vitro and in vivo. Protective capability of LTB-NS1∆63 and its-induced antisera was evaluated in the mice challenged with JEV by analyzing mortality and morbidity. FINDINGS: LTB-NS1∆63 induced immune responses to a similar level as LTB-NS1, but more robust than NS1∆63 alone, particularly in the context of oral immunization of mice. Oral vaccination of LTB-NS1∆63 led to a higher survival rate than that of NS1∆63 or live-attenuated JEV vaccine SA14-14-2 in the mice receiving lethal JEV challenge. LTB-NS1∆63 protein also significantly decreases the morbidity of JEV-infected mice. In addition, passive transfer of LTB-NS1∆63-induced antisera provides a protection against JEV infection in mice. INTERPRETATION: NS1∆63 bears JEV NS1 antigenicity. Besides, LTB-NS1∆63 could serve as a novel protein-based mucosa vaccine targeting JEV and other flaviviruses. FUNDING: This work was supported by the National Natural Science Foundation, Jiangxi Province Science and Technology Committee, Education Department of Jiangxi Province.

Testis developmental related gene 1 (TDRG1) encodes a progressive motility-associated protein in human spermatozoa.

STUDY QUESTION: Is there an association between the human testis-specific gene, testis developmental related gene 1 (TDRG1) and human sperm motility? SUMMARY ANSWER: TDRG1 is associated with asthenozoospermia and involved in regulating human sperm motility. WHAT IS KNOWN ALREADY: Many testis-specific proteins potentially regulate spermatogenesis and sperm motility. We have identified a novel human testis-specific gene, TDRG1, which encodes a 100-amino-acid protein localized in the human sperm tail, yet little is known about its role in human spermatozoa. STUDY DESIGN, SIZE, DURATION: Sperm samples were obtained from normozoospermic men and asthenozoospermic men who visited the reproductive medical center at Jiangxi Maternal and Child Health Hospital, Nanchang, Jiangxi, China between February 2018 and January 2019. In total, 27 normozoospermic men and 25 asthenozoospermic men were recruited to participate in the study. PARTICIPANTS/MATERIALS, SETTING, METHODS: The level of TDRG1 in sperm of normozoospermic and asthenozoospermic men was examined by immunoblotting and immunofluorescence assays. Progressive motility was examined by computer-aided sperm analysis. The correlation between the TDRG1 protein level and progressive motility was analyzed by linear regression. TDRG1 was imported into the sperm of normozoospermic and asthenozoospermic men using a cell-penetrating peptide (CPP)-fused TDRG1 recombinant protein (CPP-TDRG1), and the progressive motility was examined. Also, the altered proteins associated with TDRG1 in asthenozoospermic sperm were detected using label-free quantification method-based quantitative proteomic technology. TDRG1-interacting proteins were identified by co-immunoprecipitation coupled with tandem mass spectrometry analysis. MAIN RESULTS AND THE ROLE OF CHANCE: The mean level of TDRG1 was significantly decreased in sperm of asthenozoospermic men compared with normozoospermic men (P < 0.05) and was positively correlated with percentage of progressively motile sperm (r2 = 0.75, P = 0.0001). The introduction of TDRG1 into human sperm, using CPP, significantly increased progressive motility (P < 0.05) and improved the progressive motility of sperm from asthenozoospermic men to the normal level. TDRG1 forms a protein complex with sperm-motility related proteins in human sperm and its downregulation was associated with a decrease in other motility-related proteins. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The sample size was limited and larger cohorts are needed for verifying the positive effect of CPP-TDRG1 on human sperm motility. Furthermore, the caution should be paid that a comprehensive safety examination would be performed to evaluate whether CPP-TDRG1 is a possible treatment approach for asthenozoospermia. WIDER IMPLICATIONS OF THE FINDINGS: Our results provide new insights into the mechanisms of sperm motility which may contribute to the diagnosis and treatment for asthenozoospermia. STUDY FUNDING/COMPETING INTEREST(S): National Natural Science Foundation of China (81501317 and 81871207 to H.C.; 81771644 to T.L.; 31671204 to X.Z.; 81571432 to Y.T.). The authors have no conflicts of interest to declare.

Dendritic Macrosurfactant Assembly for Physical Functionalization of HIPE-Templated Polymers.

High-internal-phase emulsion-templated macroporous polymers (polyHIPEs) have attracted much interest, but their surface functionalization remains a primary concern. Thus, competitive surface functionalization via physical self-assembly of macrosurfactants was reviewed. Dendritic and diblock-copolymer macrosurfactants were tested, and the former appeared to be more topologically competitive in terms of solubility, viscosity, and versatility. In particular, hyperbranched polyethyleneimine (PEI) was transformed into dendritic PEI macrosurfactants through click-like N-alkylation with epoxy compounds. Free-standing PEI macrosurfactants were used as molecular nanocapsules for charge-selective guest encapsulation and robustly dictated the surface of a macroporous polymer through the HIPE technique, in which the macroporous polymer could act as a well-recoverable adsorbent. Metal nanoparticle-loaded PEI macrosurfactants could similarly lead to polyHIPE, whose surface was dictated by its catalytic component. Unlike conventional Pickering stabilizer, PEI macrosurfactant-based metal nanocomposite resulted in open-cellular polyHIPE, rendering the catalytic sites well accessible. The active amino groups on the polyHIPE could also be transformed into functional groups of aminopolycarboxylic acids, which could efficiently eliminate trace and heavy metal species in water.

Effects of the environmental endocrine disruptors di-2-ethylhexyl phthalate and mono-2-ethylhexyl phthalate on human sperm function in vitro.

Di-2-ethylhexyl phthalate (DEHP), a plastic-derived, endocrine-disrupting chemical, has been shown to exhibit male reproductive toxicity. However, its effects on human mature spermatozoa are largely unknown. In this study we investigated the invitro effects of DEHP and mono-2-ethylhexyl phthalate (MEHP; the main metabolite of DEHP) on sperm function and the mechanisms involved. Human spermatozoa were exposed to phthalates invitro at the doses that cover the concentrations detected in human semen: 20nM-8 µM DEHP, 1nM-20 µM MEHP or a mixture of 20nM-8 µM DEHP and 1nM-20 µM MEHP. DEHP and MEHP, alone or in combination, had no effect on the viability, membrane integrity, motility, homeostasis of reactive oxygen species or mitochondrial activity of human spermatozoa. Interestingly, 1nM-20 µM MEHP and combinations of 20nM-8 µM DEHP and 1nM-20 µM MEHP enhanced penetration ability, hyperactivation and the spontaneous acrosome reaction of human spermatozoa, and increased intracellular free Ca2+ concentrations ([Ca2+]i) and tyrosine phosphorylation, two key signalling pathways that regulate sperm function. The findings of this study suggest that invitro exposure to MEHP metabolised from DEHP affects human sperm function by inducing increases in sperm [Ca2+]i and tyrosine phosphorylation, which adds to our understanding of the effects of DEHP on male reproduction.

Anethole compromises human sperm function by affecting the sperm intracellular calcium concentration and tyrosine phosphorylation.

Anethole is a natural anisole derivative that has been widely used in food and daily chemical industries, agricultural applications and the traditional medicine. It is closely related to aspects of daily life, and humans can easily be exposed to it. Although the reproductive toxicity of anethole was shown in the rat, its effect on human reproduction remains unknown. In this study, we examined the effect of anethole on human sperm in vitro. Different anethole doses (0.1, 1, 10, and 100 µM) were applied to ejaculated human sperm. Fertilization-essential functions, as well as the intracellular calcium concentration ([Ca2+]i) and tyrosine phosphorylation, two vital factors for regulating sperm function, were measured. The results indicated that 10 and 100 µM anethole significantly reduced the motility, hyperactivation, and penetration ability of human sperm (P < 0.05) and inhibited the increase in human sperm functions induced by progesterone, a hormone essential for sperm function activation. Additionally, 10 and 100 µM anethole decreased both basal and progesterone-increased tyrosine phosphorylation, [Ca2+]i, and the current of CATSPER, a cation channel of sperm predominant for Ca2+ influx. These results suggest that anethole inhibits human sperm functions by reducing sperm [Ca2+]i through CATSPER and suppressing tyrosine phosphorylation in vitro, raising the fact that the caution is needed when overtaking anethole.

In vitro exposure to metformin activates human spermatozoa at therapeutically relevant concentrations.

BACKGROUND: Metformin, a drug used to treat type 2 diabetes, has gained attention for its multiple therapeutic applications. However, little is known about its effects on human sperm function at therapeutically relevant concentration. OBJECTIVES: The aim of this study was to elucidate the in vitro actions of metformin on human sperm function and explore the underlying mechanism of any effects. MATERIALS AND METHODS: Human ejaculated spermatozoa were treated with therapeutically relevant concentrations (0.25, 5, 10, 20, 40, and 80 µM) of metformin in vitro. Fertilization-essential functions of spermatozoa were examined, including viability, motility, capacitation, acrosome reaction, hyperactivation, and penetration ability. The signaling pathways mediated by 5'-AMP-activated protein kinase (AMPK), intracellular calcium concentration ([Ca2+ ]i ), and tyrosine phosphorylation of spermatozoa were also measured. RESULTS: Although metformin did not affect sperm viability, motility, and [Ca2+ ]i , it significantly increased the percentages of capacitated spermatozoa, acrosomal-reacted spermatozoa, and hyperactivated spermatozoa as well as penetration ability of human spermatozoa at the concentrations of 40 and 80 µM (P < .05). These concentrations of metformin also elevated the levels of phosphorylated AMPK and tyrosine phosphorylation in human spermatozoa. In addition, activation of AMPK by A769662 (an AMPK activator) had similar effects to metformin on human spermatozoa, while inhibition of AMPK by Compound C (an AMPK inhibitor) suppressed the enhancement of metformin on human spermatozoa. CONCLUSION: Our findings indicate that metformin activates human sperm function through an AMPK-related mechanism which increases tyrosine phosphorylation at therapeutically relevant concentrations, thereby suggesting its improvement on human sperm function when treating subfertile males of type 2 diabetes.

A novel copy number variation in CATSPER2 causes idiopathic male infertility with normal semen parameters.

STUDY QUESTION: Are genetic abnormalities in CATSPER (cation channel of sperm) genes associated with idiopathic male infertility with normal semen parameters and, if so, how do they affect male fertility? SUMMARY ANSWER: A novel copy number variation (CNV) in CATSPER2 causes idiopathic male infertility with normal semen parameters by disrupting the ability of sperm to penetrate viscous media, undergo hyperactivation and respond to progesterone. WHAT IS KNOWN ALREADY: CATSPER is the principle Ca2+ channel mediating extracellular Ca2+ influx into spermatozoa. Although several case reports have suggested a causal relationship between CATSPER disruption and human male infertility, whether genetic abnormalities in CATSPER genes are associated with idiopathic male infertility with normal semen parameters remains unclear. STUDY DESIGN, SIZE, DURATION: Spermatozoa were obtained from men attending the reproductive medical center at Jiangxi Provincial Maternal and Child Health Hospital, Nanchang, Jiangxi, China between January 2014 and June 2016. In total, 120 men from infertile couples and 20 healthy male donors were selected to take part in the study, based on their normal semen parameters. PARTICIPANTS/MATERIALS, SETTING, METHODS: CATSPER and KSPER currents were assessed using the whole-cell patch-clamp technique. Whole-genome sequencing and TaqMan® CNV assays were performed to identify genetic variations. The expression levels of genes encoding the CATSPER complex were measured by quantitative real-time PCR and Western blot. Sperm motion characteristics and hyperactivation were examined with a computer-aided sperm analysis (CASA) system. Sperm responses to progesterone, assessed as increases in CATSPER current and intercellular Ca2+ concentrations ([Ca2+]i), as well as inducement of penetration ability and acrosome reaction, were examined by means of whole-cell patch-clamp technique, single-sperm [Ca2+]i imaging, penetration into methylcellulose assay and chlortetracycline staining, respectively. MAIN RESULTS AND THE ROLE OF CHANCE: An infertile man with complete disruption of CATSPER current was identified. This individual has a novel CNV which disrupts one gene copy in the region 43894500-43950000 in chromosome 15 (GRCh37.p13 Primary Assembly, nsv3067119), containing the whole DNA sequence of CATSPER2. This CNV affected the expression of CATSPER2, resulting in dramatically reduced levels of CATSPER2 proteins in the individual's spermatozoa. Although this individual exhibited normal semen parameters, his spermatozoa showed impaired penetration ability, deficient hyperactivation, and did not respond to progesterone, in terms of monovalent current potentiation, [Ca2+]i increase, penetration ability enhancement and acrosome reaction inducement, which may explain the individual's idiopathic infertility. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Our novel findings require more cases to support the CATSPER2 CNV identified in this study as a common cause of idiopathic male infertility in patients with normal semen parameters. Therefore, caution must be taken when extrapolating the use of this CNV as a potential biomarker for idiopathic male infertility. WIDER IMPLICATIONS OF THE FINDINGS: The findings from the unique human CATSPER 'knockout' model in this study not only confirm the essential roles of CATSPER in mediating progesterone response and regulating hyperactivation in human spermatozoa but also reveal that disruption of CATSPER current is a significant factor causing idiopathic male infertility. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by National Natural Science Foundation of China (81771644 and 31400996 to T.L.; 31230034 to X.Z.); National Basic Research Program of China (973 Program, 2015CB943003 to X.Z.); National Key Research and Development Program of China (2016YFC1000905 to T.L.); Natural Science Foundation of Jiangxi, China (20121BBG70021 and GJJ12015 to X.Z.; 20161BAB204167 and 20171ACB21006 to T.L.) and the open project of National Population and Family Planning Key Laboratory of Contraceptives and Devices Research (No. 2016KF07 to T.L.). The authors have no conflicts of interest to declare.

Effects of two environmental endocrine disruptors di-n-butyl phthalate (DBP) and mono-n-butyl phthalate (MBP) on human sperm functions in vitro.

Di-n-butyl phthalate (DBP), a plastic-derived, endocrine-disrupting chemical, is regarded as a male reproductive toxicant. In this study, we investigated the in vitro actions of DBP and mono-n-butyl phthalate (MBP, the main metabolite of DBP) on human sperm functions. Human sperm were treated with DBP (2 nM-6 µM), MBP (1 nM-3 µM), and a mixture of DBP and MBP in vitro. The results showed that only DBP at 6 µM, a dose reported in semen of infertile men, MBP at 3 µM (three times of the reported maximum MBP concentration in semen), and their mixture, had obvious adverse effects on sperm motility, penetration ability and capacitation. In addition, these doses of phthalates suppressed human sperm tyrosine phosphorylation, a key signaling pathway that regulates sperm functions. Our findings indicate that DBP and MBP may compromise human sperm functions by inhibiting sperm tyrosine phosphorylation once they accumulate in semen at high levels.

ß-Elemene Inhibits Human Sperm Function by Affecting Sperm Vitality and Intracellular Calcium.

BACKGROUND/AIMS: ß-Elemene is a bioactive sesquiterpene compound that exhibits a potent anti-tumor effect and is used in various clinical applications. However, little is known about its effect on the male reproductive system. The objective of this study was to investigate the in vitro actions of ß-elemene on human sperm function and elucidate the underlying mechanism. METHODS: The cytotoxicity of ß-elemene toward MCF-10A, MDA-MD-231, and A549 cells was evaluated with cell proliferation and colony formation assays. Additionally, human sperm were treated with different concentrations (0, 10, 20, 40, 80, 160, and 320 µM) of ß-elemene in vitro. The characteristics in human sperm essential for fertilization, including vitality, motility, capacitation, acrosome reaction, responsiveness to progesterone, and intracellular calcium concentration ([Ca2+]i) were examined with a computer-assisted sperm analysis system, chlortetracycline staining, and a fluorescent Ca2+ indicator. RESULTS: A comprehensive evaluation of sperm motility, especially hyperactivated motility, revealed that treatments with 40-320 µM ß-elemene decreased human sperm vitality, motility (total motility, progressive motility, and curvilinear velocity), and penetrating ability in a dose-dependent manner, but were non-toxic or minimally toxic toward MCF-10A, MDA-MD-231, and A549 cells. Although 10 and 20 µM ß-elemene did not affect sperm vitality and motility, these concentrations increased the spontaneous acrosome reaction and inhibited progesterone-induced sperm functions by affecting sperm [Ca2+]i. CONCLUSION: These results suggest that ß-elemene inhibits human sperm function by affecting sperm vitality and [Ca2+]i. These observations must be considered when using ß-elemene to treat cancer patients who may wish to preserve their fertility.

Rosmarinic acid compromises human sperm functions by an intracellular Ca2+ concentration-related mechanism.

Rosmarinic acid (RA), a natural phenolic ester, is cytoprotective for male reproduction in animal models. The present study investigated the in vitro actions of RA on human sperm functions. Human sperm were exposed to 1, 10, 100, and 1000 µM RA in vitro and sperm functions were examined. The results showed that although RA did not affect human sperm viability, RA at 10-1000 µM dose-dependently reduced sperm motility, penetration ability, capacitation, and spontaneous acrosome reaction. In addition, the intracellular Ca2+ concentration ([Ca2+]i), which serve as a key regulator of sperm function, was decreased by RA (10-1000 µM) in a dose-dependent manner. Furthermore, the current of the sperm-specific potassium channel, KSPER, which is predominant for Ca2+ influx in sperm, was dose-dependently inhibited by 10-1000 µM RA. Therefore, we conclude that in vitro exposure to RA can compromise human sperm functions by decreasing sperm [Ca2+]i through the suppression of KSPER current.

Diethylstilbestrol activates CatSper and disturbs progesterone actions in human spermatozoa.

STUDY QUESTION: Is diethylstilbestrol (DES), a prototypical endocrine-disrupting chemical (EDC), able to induce physiological changes in human spermatozoa and affect progesterone actions? SUMMARY ANSWER: DES promoted Ca2+ flux into human spermatozoa by activating the cation channel of sperm (CatSper) and suppressed progesterone-induced Ca2+ signaling, tyrosine phosphorylation and sperm functions. WHAT IS KNOWN ALREADY: DES significantly impairs the male reproductive system both in fetal and postnatal exposure. Although various EDCs affect human spermatozoa in a non-genomic manner, the effect of DES on human spermatozoa remains unknown. STUDY DESIGN, SIZE, DURATION: Sperm samples from normozoospermic donors were exposed in vitro to a range of DES concentrations with or without progesterone at 37°C in a 5% CO2 incubator to mimic the putative exposure to this toxicant in seminal plasma and the female reproductive tract fluids. The incubation time varied according to the experimental protocols. All experiments were repeated at least five times using different individual sperm samples. PARTICIPANTS/MATERIALS, SETTING, METHODS: Human sperm intracellular calcium concentrations ([Ca2+]i) were monitored with a multimode plate reader following sperm loading with Ca2+ indicator Fluo-4 AM, and the whole-cell patch-clamp technique was performed to record CatSper and alkalinization-activated sperm K+ channel (KSper) currents. Sperm viability and motility parameters were assessed by an eosin-nigrosin staining kit and a computer-assisted semen analysis system, respectively. The ability of sperm to penetrate into viscous media was examined by penetration into 1% methylcellulose. The sperm acrosome reaction was measured using chlortetracycline staining. The level of tyrosine phosphorylation was determined by western blot assay. MAIN RESULTS AND THE ROLE OF CHANCE: DES exposure rapidly increased human sperm [Ca2+]i dose dependently and even at an environmentally relevant concentration (100 pM). The elevation of [Ca2+]i was derived from extracellular Ca2+ influx and mainly mediated by CatSper. Although DES did not affect sperm viability, motility, penetration into viscous media, tyrosine phosphorylation or the acrosome reaction, it suppressed progesterone-stimulated Ca2+ signaling and tyrosine phosphorylation. Consequently, DES (1-100 µM) significantly inhibited progesterone-induced human sperm penetration into viscous media and acrosome reaction. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Although DES has been shown to disturb progesterone actions on human spermatozoa, this study was performed in vitro, and caution must be taken when extrapolating the results in practical applications. WIDER IMPLICATIONS OF THE FINDINGS: The present study revealed that DES interfered with progesterone-stimulated Ca2+ signaling and tyrosine phosphorylation, ultimately inhibited progesterone-induced human sperm functions and, thereby, might impair sperm fertility. The non-genomic manner in which DES disturbs progesterone actions may be a potential mechanism for some estrogenic endocrine disruptors to affect human sperm function. STUDY FUNDING/COMPETING INTERESTS: National Natural Science Foundation of China (No. 31400996); Natural Science Foundation of Jiangxi, China (No. 20161BAB204167 and No. 20142BAB215050); open project of National Population and Family Planning Key Laboratory of Contraceptives and Devices Research (No. 2016KF07) to T. Luo; National Natural Science Foundation of China (No. 81300539) to L.P. Zheng. The authors have no conflicts of interest to declare.

Toxic effects of 2,4-dichlorophenoxyacetic acid on human sperm function in vitro.

The herbicide 2,4-Dichlorophenoxyacetic acid (2,4-D) is globally used in agriculture and has been linked to human sperm abnormalities in vivo. However, its effects on ejaculated human spermatozoa in vitro have not been characterized. Therefore, we examined the effects of 2,4-D on the functions of ejaculated human spermatozoa in vitro, including: sperm motility, the ability to move through a viscous medium, capacitation, and the acrosome reaction. Different doses of 2,4-D (10 nM, 100 nM, 1 µM, 10 µM, 100 µM, and 200 µM) were applied to human spermatozoa prepared from normal fresh semen samples. The results indicated that 2,4-D did not affect the viability, capacitation, or spontaneous acrosome reactions of human spermatozoa, but it dose-dependently inhibited the total motility, progressive motility, ability to penetrate viscous medium, and progesterone-induced capacitation and acrosome reaction rates. These results suggest that exposure to 2,4-D and its accumulation in the seminal plasma and follicular fluid might increase the risk of infertility. Our findings provide new insights for understanding the male reproductive toxicity of 2,4-D.

Ketamine inhibits human sperm function by Ca(2+)-related mechanism.

Ketamine, a dissociative anesthetic, which was widely used in human and animal medicine, has become a popular recreational drug, as it can induce hallucinatory effects. Ketamine abuse can cause serious damage to many aspects of the organism, mainly reflected in the nervous system and urinary system. It has also been reported that ketamine can impair the male genital system. However, the detailed effect of ketamine on human spermatozoa remains unclear. Thus, we investigated the in vitro effects of ketamine on human sperm functions, to elucidate the underlying mechanism. Human sperm were treated in vitro with different concentrations of ketamine (0, 0.125, 0.25, 0.5, 1 g/L). The results showed that 0.25-1 g/L ketamine inhibited sperm total motility, progressive motility and linear velocity, in a dose-dependent manner. In addition, the sperm's ability to penetrate viscous medium and the progesterone-induced acrosome reaction were significantly inhibited by ketamine. Ketamine did not affect sperm viability, capacitation and spontaneous acrosome reaction. The intracellular calcium concentration ([Ca(2+)]i), which is a central factor in the regulation of human sperm function, was decreased by ketamine (0.125-1 g/L) in a dose-dependent manner. Furthermore, the currents of the sperm-specific Ca(2+) channel, CatSper, which modulates Ca(2+) influx in sperm, were inhibited by ketamine (0.125-1 g/L) in a dose-dependent manner. Our findings suggest that ketamine induces its toxic effects on human sperm functions by reducing sperm [Ca(2+)]i through inhibition of CatSper channel.

Lead Inhibits Human Sperm Functions by Reducing the Levels of Intracellular Calcium, cAMP, and Tyrosine Phosphorylation.

It is well known that there has been a worldwide decrease in human male fertility in recent years. One of the main factors affecting this is environmental pollution. Lead is one of the major heavy metal contaminants that threaten the health of animals and human beings in China. It preferentially accumulates in male reproductive organs and can be up to 10 µM in human seminal plasma. Lead impairs mammalian spermatogenesis and sperm quality in vivo. It also inhibits sperm functions in vitro but the underlying mechanisms remain unclear. Therefore, we aimed to investigate the in vitro toxicity of lead on human sperm functions and to elucidate the underlying mechanisms. Semen samples were collected from 20 healthy volunteers with different careers and backgrounds living in Nanchang, Jiangxi. Human sperm suspensions were treated with different concentrations of lead acetate (0, 0.5, 2.5, 10, 50, and 100 µM) and the viability, motility, capacitation and progesterone-induced acrosome reaction were examined. Treatment with 10-100 µM lead acetate dose-dependently inhibited total and progressive motility measures, capacitation and progesterone-induced acrosome reaction. It also dose-dependently decreased the intracellular concentrations of cyclic adenosine monophosphate (cAMP) and calcium ([Ca(2+)]i), and reduced the tyrosine phosphorylation of sperm proteins, all of which are thought to be key factors in the regulation of sperm function. Our findings suggest that lead inhibits human sperm functions by reducing the levels of sperm intracellular cAMP, [Ca(2+)]i and tyrosine phosphorylation of sperm proteins in vitro.

Emodin inhibits human sperm functions by reducing sperm [Ca(2+)]i and tyrosine phosphorylation.

Emodin, a bioactive anthraquinone widely used in Chinese traditional medicine, disrupts mouse testicular gene expression in vivo. In this study, we investigated the toxicity of emodin to human sperm in vitro. Different doses of emodin (25, 50, 100, 200 and 400µM) were applied to ejaculated human sperm. The results indicated that 100, 200 and 400µM emodin significantly inhibited the total motility, progressive motility and linear velocity of human sperm. In addition, sperm's ability to penetrate viscous medium together with progesterone induced capacitation and acrosome reaction was also adversely affected by emodin. In contrast, emodin did not affect sperm viability. Furthermore, intracellular Ca(2+) concentration ([Ca(2+)]i) and tyrosine phosphorylation, which serve as key regulators of sperm function, were dose-dependently reduced by emodin (50-400µM). These results suggest that emodin inhibits human sperm functions by reducing sperm [Ca(2+)]i and suppressing tyrosine phosphorylation in vitro.