Short-term storage of semen samples in acidic extender increases the proportion of females in pigs.

BACKGROUND: Sex preselection is a desired goal of the animal industry to improve production efficiency, depending on industry demand. In the porcine industry, there is a general preference for pork from female and surgically castrated male pigs. Therefore, the birth of more females than males in a litter leads to economic benefits and improved animal welfare in the pig production industry. Our previous study suggested that the porcine semen extender (BTS) adjusted to pH 6.2 maximises the differences in viability between X-chromosome-bearing (X) spermatozoa and Y-chromosome-bearing (Y) spermatozoa without affecting sperm's functional parameters. In this study we aimed to evaluate whether the pH 6.2 extender is applicable at the farm level for increasing the number of female piglets without a decline in spermatozoa fertility. Artificial insemination (AI) was carried out with spermatozoa stored at pH 6.2 and pH 7.2 (original BTS) at day 1 and day 2 of storage. Next, the functional parameters of the spermatozoa, litter size, farrowing rate, and female-to-male ratio of offspring were determined. RESULTS: Although sperm motility decreased significantly after 2 d of storage, the viability of spermatozoa was preserved at pH 6.2 for 3 d. There was no significant difference in the farrowing rate and average litter size between the group inseminated with the spermatozoa stored in (pH 7.2) and that inseminated with spermatozoa stored in acidic BTS. The percentage of female piglets was approximately 1.5-fold higher in sows inseminated on day 1 in the pH 6.2 than in the pH 7.2 group. Furthermore, although there was no significant difference in the female-to-male ratio, the percentage of female piglets born was slightly higher in the pH 6.2 group than in the pH 7.2 group on day 2. CONCLUSIONS: The method optimised in our study is simple, economical, and may enhance the number of female births without any decline in spermatozoa fertility.

Bisphenol A exposure increases epididymal susceptibility to infection in mice.

Male fertility is linked with several well-orchestrated events including spermatogenesis, epididymal maturation, capacitation, the acrosome reaction, fertilization, and beyond. However, the detrimental effects of bisphenol A (BPA) on sperm maturation compared to spermatogenesis and sperm cells remain unclear. Therefore, this study was to investigate whether pubertal exposure to BPA induces male infertility via interruption of the immune response in the epididymis. CD-1 male mice (5 weeks old) were treated daily with vehicle (corn oil) and 50 mg BPA/kg-BW for 6 weeks by oral gavage. Following BPA exposure, we observed decreased intraepithelial projection of basal cells, indicative of changes to the luminal environment. We also observed decreased projection of macrophages and protrusion of apoptotic cells into the lumen induced by incomplete phagocytosis of apoptotic cells in the caput epididymis. Exposure to BPA also reduced the anti- and pro-inflammatory cytokines IL-10, IL-6, IFN-γ, and IL-7 in the epididymis, while the chemotaxis-associated cytokines CCL12, CCL17, CXCL16, and MCP-1 increased. This study suggests two possible mechanisms for BPA induction of male infertility. First, exposure to BPA may induce an imbalance of immune homeostasis by disrupting the ability of basal cells to perceive environmental changes. Second, exposure to BPA may lead to collapse of macrophage phagocytosis via downregulation of intraepithelial projection and inflammatory-related cytokines. In conclusion, the observed potential pathways can lead to autoimmune disorders such epididymitis and orchitis.

Importation and Transmission of SARS-CoV-2 B.1.1.529 (Omicron) Variant of Concern in Korea, November 2021.

In November 2021, 14 international travel-related severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) B.1.1.529 (omicron) variant of concern (VOC) patients were detected in South Korea. Epidemiologic investigation revealed community transmission of the omicron VOC. A total of 80 SARS-CoV-2 omicron VOC-positive patients were identified until December 10, 2021 and 66 of them reported no relation to the international travel. There may be more transmissions with this VOC in Korea than reported.

New technique of sex preselection for increasing female ratio in boar sperm model.

In this study, we tried to optimize the porcine semen extender conditions to maximize the differences between live X chromosome-bearing (X) spermatozoa and to Y chromosome-bearing (Y) spermatozoa without a decline in the fertility rate at different pH conditions during storage. We observed the viability of X and Y boar spermatozoa in acidic (pH 6.2), original (pH 7.2), and alkaline condition (pH 8.2) for 5 days to investigate the effect of storage conditions on the X to Y spermatozoa ratio. The functional parameters of spermatozoa were also examined to evaluate sperm quality. Sperm motility was preserved at pH 7.2 and pH 6.2 for 3 days, while sperm motility at pH 8.2 decreased significantly after 2 days. Non-capacitated spermatozoa increased while capacitated spermatozoa decreased during storage. Sperm viability decreased significantly duration-dependent under all pH conditions, but there was no significant difference during storage at pH 6.2 and 7.2. The X: Y ratio of live spermatozoa in acidic condition was maximized (1.2:1) without affecting the sperm function and fertility-related protein expression after 2 days compared to original conditions. Moreover, insemination of sows using acidic extender increased the number of female pups on days 1 and 2 of preservation. These results indicate that the production of female offspring may increase when acidic BTS is used for 2 days without affecting the success rate of AI. Above all, this method is simple and economical compared to other methods.

Endocrine-Disrupting Chemicals and Infectious Diseases: From Endocrine Disruption to Immunosuppression.

Endocrine-disrupting chemicals (EDCs) are hormonally active compounds in the environment that interfere with the body's endocrine system and consequently produce adverse health effects. Despite persistent public health concerns, EDCs remain important components of common consumer products, thus representing ubiquitous contaminants to humans. While scientific evidence confirmed their contribution to the severity of Influenza A virus (H1N1) in the animal model, their roles in susceptibility and clinical outcome of the coronavirus disease (COVID-19) cannot be underestimated. Since its emergence in late 2019, clinical reports on COVID-19 have confirmed that severe disease and death occur in persons aged ≥65 years and those with underlying comorbidities. Major comorbidities of COVID-19 include diabetes, obesity, cardiovascular disease, hypertension, cancer, and kidney and liver diseases. Meanwhile, long-term exposure to EDCs contributes significantly to the onset and progression of these comorbid diseases. Besides, EDCs play vital roles in the disruption of the body's immune system. Here, we review the recent literature on the roles of EDCs in comorbidities contributing to COVID-19 mortality, impacts of EDCs on the immune system, and recent articles linking EDCs to COVID-19 risks. We also recommend methodologies that could be adopted to comprehensively study the role of EDCs in COVID-19 risk.

Coronavirus Disease Exposure and Spread from Nightclubs, South Korea.

At least 246 cases of coronavirus disease (COVID-19) have been linked to nightclubs in Seoul, South Korea. During the April 30-May 5 holiday, young adults from across the country who visited nightclubs in Seoul contracted COVID-19 and spread it nationally. Nightclubs were temporarily closed to limit COVID-19 spread.

Coronavirus Disease Outbreak in Call Center, South Korea.

We describe the epidemiology of a coronavirus disease (COVID-19) outbreak in a call center in South Korea. We obtained information on demographic characteristics by using standardized epidemiologic investigation forms. We performed descriptive analyses and reported the results as frequencies and proportions for categoric variables. Of 1,143 persons who were tested for COVID-19, a total of 97 (8.5%, 95% CI 7.0%-10.3%) had confirmed cases. Of these, 94 were working in an 11th-floor call center with 216 employees, translating to an attack rate of 43.5% (95% CI 36.9%-50.4%). The household secondary attack rate among symptomatic case-patients was 16.2% (95% CI 11.6%- 22.0%). Of the 97 persons with confirmed COVID-19, only 4 (1.9%) remained asymptomatic within 14 days of quarantine, and none of their household contacts acquired secondary infections. Extensive contact tracing, testing all contacts, and early quarantine blocked further transmission and might be effective for containing rapid outbreaks in crowded work settings.

Multigenerational and transgenerational impact of paternal bisphenol A exposure on male fertility in a mouse model.

STUDY QUESTION: How does paternal exposure to bisphenol A (BPA) affect the fertility of male offspring in mice in future generations? SUMMARY ANSWER: Paternal exposure to BPA adversely affects spermatogenesis, several important sperm functions and DNA methylation patterns in spermatozoa, which have both multigenerational (in F0 and F1) and partial transgenerational (mainly noticed in F2, but F3) impacts on the fertility of the offspring. WHAT IS KNOWN ALREADY: BPA, a synthetic endocrine disruptor, is used extensively to manufacture polycarbonate plastics and epoxy resins. Growing evidence suggests that exposure to BPA during the developmental stages results in atypical reproductive phenotypes that could persist for generations to come. STUDY DESIGN, SIZE, DURATION: CD-1 male mice (F0) were treated with BPA (5 or 50 mg/kg body weight per day (bw/day)) or ethinylestradiol (EE) (0.4 µg/kg bw/day) for 6 weeks. Control mice were treated with vehicle (corn oil) only. The treated male mice were bred with untreated female mice to produce first filial generation (F1 offspring). The F2 and F3 offspring were produced similarly, without further exposure to BPA. PARTICIPANTS/MATERIALS, SETTING, METHODS: Histological changes in the testis along with functional, biochemical and epigenetic (DNA methylation) properties of spermatozoa were investigated. Subsequently, each parameter of the F0-F3 generations was compared between BPA-treated mice and control mice. MAIN RESULTS AND THE ROLE OF CHANCE: Paternal BPA exposure disrupted spermatogenesis by decreasing the size and number of testicular seminiferous epithelial cells, which eventually led to a decline in the total sperm count of F0-F2 offspring (P < 0.05). We further showed that a high BPA dose decreased sperm motility in F0-F2 males by mediating the overproduction of reactive oxygen species (F0-F1) and decreasing intracellular ATP (F0-F2) in spermatozoa (P < 0.05). These changes in spermatozoa were associated with altered global DNA methylation patterns in the spermatozoa of F0-F3 males (P < 0.05). Furthermore, we noticed that BPA compromised sperm fertility in mice from the F0-F2 (in the both dose groups) and F3 generations (in the high-dose group only). The overall reproductive toxicity of BPA was equivalent to or higher (high dose) than that of the tested dose of EE. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: Further research is required to determine the variables (e.g. lowest BPA dose) that are capable of producing changes in sperm function and fertility in future generations. WIDER IMPLICATIONS OF THE FINDINGS: These results may shed light on how occupational exposure to BPA can affect offspring fertility in humans. STUDY FUNDING/COMPETING INTEREST(S): This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (Grant No. NRF-2018R1A6A1A03025159). M.S.R. was supported by Korea Research Fellowship Program through the NRF funded by the Ministry of Science and ICT (Grant No. 2017H1D3A1A02013844). There are no competing interests.

Bisphenol A affects the maturation and fertilization competence of spermatozoa.

Although there are numerous studies on bisphenol A (BPA) on the testis and spermatozoa, the effect of BPA on the physiological link between the testis and maturation of spermatozoa has not been studied. To provide an optimal environment (acidic pH) for sperm maturation in the epididymis, clear cells secrete protons and principal cells reabsorb bicarbonate and the secreted proton. Because of its crucial role in sperm maturation and fertility, functional changes in the epididymis following BPA exposure must be considered to fully understand the mechanisms of BPA on male fertility. Here, we identified the adverse effects of BPA exposure during puberty in male mice. CD-1 male mice were gavaged daily with vehicle (corn oil) and 50 mg BPA/kg-BW for 6 weeks. We determined the changes in epididymis, functional sperm parameters including motility, capacitation status, tyrosine phosphorylation, and fertility-related protein expression and in vitro and in vivo fertility rate following BPA exposure. Expression of vacuolar-type H + -ATPase is necessary for the secretion of protons by clear cells of the caput epididymis and was directly down-regulated following BPA exposure, while there were no changes in the other epithelial cell types in the epididymis. Also, pERK 1/2 signaling pathway was increased significantly in the caput epididymis following BPA exposure. Consequently, the luminal pH slightly increased, resulting in premature capacitation of spermatozoa. Moreover, there was a significant loss of the acrosomal membrane following an increase of protein tyrosine phosphorylation, while PKA activity decreased during sperm capacitation. Fertility-related proteins also showed aberrant expression upon BPA exposure. These modifications resulted in decreased male fertility in vitro and in vivo.

Application of Testing-Tracing-Treatment Strategy in Response to the COVID-19 Outbreak in Seoul, Korea.

BACKGROUND: Following the coronavirus disease 2019 (COVID-19) outbreak in Wuhan, China, a total of 637 patients had been diagnosed with the disease in Seoul as of May 2, 2020. Our study aimed to describe the impact of the 3T strategies (preemptive testing, prompt tracing and proper treatment) on the epidemiological characteristics of COVID-19 in Seoul. METHODS: The descriptive and explanatory analysis was carried out on critical indicators such as epidemiological characteristics and key duration of patient status change from January 24 to May 2 in Seoul before and after preemptive testing for patients under investigation associated with COVID-19 clusters. RESULTS: Preemptive testing increased the positive test rate (3.9% to 4.2%), an asymptomatic case at diagnosis (16.9% to 30.6%), and reduced the time from symptom onset to quarantine (4.0 to 3.0 days). Prompt tracing decreased unknown sources of infection (6.9% to 2.8%), the mean number of contacts (32.2 to 23.6), and the time-varying reproduction number R(t) (1.3 to 0.6). With proper treatment, only 2 cases of mortality occurred, resulting in a fatality rate of just 0.3%. CONCLUSION: In the first wave of the COVID-19 pandemic lasting 100 days, the effect of the 3T strategies flattened the curve and decreased the time during which infected individuals were contagious, thereby lowering the R(t) below 1 in Seoul.

The expression and localization of V-ATPase and cytokeratin 5 during postnatal development of the pig epididymis.

OBJECTIVE: We examined the localization and expression of H+ pumping vacuolar ATPase (V-ATPase) and cytokeratin 5 (KRT5) in the epididymis of pigs, expressed in clear and basal cells, respectively, during postnatal development. METHODS: Epididymides were obtained from pigs at 1, 7, 21, 60, 120, and 180 days of age; we observed the localization and expression patterns of V-ATPase and KRT5 in the different regions of these organs, namely, the caput, corpus, and cauda. The differentiation of epididymal epithelial cells was determined by immunofluorescence labeling using cell-type-specific markers and observed using confocal microscopy. RESULTS: At postnatal day 5 (PND5), the localization of clear cells commenced migration from the cauda toward the caput. Although at PND120, goblet-shaped clear cells were detected along the entire length of the epididymis, those labeled for V-ATPase had disappeared from the corpus to cauda and were maintained only in the caput epididymis in adult pigs. In contrast, whereas basal cells labeled for KRT5 were only present in the vas deferens at birth, they were detected in all regions of the epididymis at PND60. These cells were localized at the base of the epithelium; however, no basal cells characterized by luminally extending cell projections were observed in any of the adult epididymides examined. CONCLUSION: The differentiation of clear and basal cells progressively initiates in a retrograde manner from the cauda to the caput epididymis. The cell-type-specific distribution and localization of the epithelial cells play important roles in establishing a unique luminal environment for sperm maturation and storage in the pig epididymis.

Unravelling purinergic regulation in the epididymis: activation of V-ATPase-dependent acidification by luminal ATP and adenosine.

KEY POINTS: In the epididymis, elaborate communication networks between epithelial cells are important with respect to establishing an optimal acidic luminal environment for the maturation and storage of spermatozoa, which is essential for male fertility. Proton secretion by epididymal clear cells is achieved via the proton pumping V-ATPase located in their apical membrane. In the present study, we dissect the molecular mechanisms by which clear cells respond to luminal ATP and adenosine to modulate their acidifying activity via the adenosine receptor ADORA2B and the pH-sensitive ATP receptor P2X4. We demonstrate that the hydrolysis of ATP to produce adenosine by ectonucleotidases plays a key role in V-ATPase-dependent proton secretion, and is part of a feedback loop that ensures acidification of the luminal compartment These results help us better understand how professional proton-secreting cells respond to extracellular cues to modulate their functions, and how they communicate with neighbouring cells. ABSTRACT: Cell-cell cross-talk is crucial for the dynamic function of epithelia, although how epithelial cells detect and respond to variations in extracellular stimuli to modulate their environment remains incompletely understood. In the present study, we used the epididymis as a model system to investigate epithelial cell regulation by luminal factors. In the epididymis, elaborate communication networks between the different epithelial cell types are important for establishing an optimal acidic luminal environment for the maturation and storage of spermatozoa. In particular, clear cells (CCs) secrete protons into the lumen via the proton pumping V-ATPase located in their apical membrane, a process that is activated by luminal alkalinization. However, how CCs detect luminal pH variations to modulate their function remains uncharacterized. Purinergic regulation of epithelial transport is modulated by extracellular pH in other tissues. In the present study, functional analysis of the mouse cauda epididymis perfused in vivo showed that luminal ATP and adenosine modulate the acidifying activity of CCs via the purinergic ADORA2B and P2X4 receptors, and that luminal adenosine content is itself regulated by luminal pH. Altogether, our observations illustrate mechanisms by which CCs are activated by pH sensitive P2X4 receptor and ectonucleotidases, providing a feedback mechanism for the maintenance of luminal pH. These novel mechanisms by which professional proton-secreting cells respond to extracellular cues to modulate their functions, as well as how they communicate with neighbouring cells, might be translatable to other acidifying epithelia.

Cigarette smoke extract may induce lysosomal storage disease-like adverse health effects.

Cigarette smoke is known to be associated with the incidence of a variety of pulmonary diseases, and alveolar macrophages are a key player in the defense mechanism against inhalable toxicants. Herein, we have found that a hydrophilic fraction in smoke extracts from 3R4F reference cigarettes (CSE) contains high concentrations of volatile substances compared to cigarette smoke condensate (amphoteric fraction). We also identified the toxic mechanism of CSE using MH-S, a mouse alveolar macrophage cell line. CSE decreased cell viability accompanying increased lactate dehydrogenase release. Additionally, mitochondrial volume and the potential increased along with enhanced expression of mitochondrial fusion proteins and decreased adenosine triphosphate production. Similarly, CSE clearly induced increase of catalase activity and intracellular calcium concentration and decrease of endoplasmic reticulum and lysosome volume at the highest dose. More interestingly, damaged organelles accumulated in the cytosol, and CSE-containing particles specifically penetrated to mitochondria. Meanwhile, any significant change in autophagy related protein expression was not found in CSE-treated cells. Subsequently, we evaluated the effects of CSE on secretion of inflammatory related cytokines and chemokines, considering the relationship between organelle damage and the disturbed immune response. Very importantly, we found that expression of innate and adaptive immunity related mediators is disrupted following CSE exposure. Taken together, we suggest that CSE may cause the accumulation of damaged organelles in the cytoplasm by impairing selective autophagic function. In addition, this accumulation is responsible for the inadequate ability of immune cells to repair the damage of lung tissue following exposure to CSE.

Repeated injection of KMRC011, a medical countermeasure for radiation, can cause adverse health effects in cynomolgus monkeys.

High-dose radiation-induced tissue damage is a major limiting factor in the medical application of nuclear technology. Herein, we tested 28-day repeated-dose toxicity of KMRC011, an agonist of toll-like receptor (TLR) 5, which is being developed as a medical countermeasure for radiation, using cynomolgus monkeys. KMRC011 (0.01, 0.02 or 0.04 mg/kg/day) was intramuscularly injected once daily for 4 weeks, and each two monkeys in both control and 0.04 mg/kg/day group were observed for an additional 2-week recovery period. There were no dose-related toxicological changes in mortality, clinical observations, body weight, food consumption, ophthalmological findings, electrocardiographs, coagulation, serum chemistry, organ weights, or urinalysis and urine chemistry. Although treatment-related changes, such as increased white blood cells, increased absolute and relative neutrophils, decreased relative lymphocytes and inflammatory lesions, were noted in the maximum dose group, these findings were not observed after the 2-week recovery period. Further, we considered that the kidneys and heart may be target organs of TLR5 agonists, as well as the spleen, and that autophagic signals can be triggered in tissue damage and the repair process. Importantly, accumulation of p62 protein, an indicator of autophagy, and a decrease of caveolin-1 protein, a regulator of TLR5 protein half-life, were found in both tissues from the highest dose group. Therefore, we conclude that the no-observed-adverse-effect level for KMRC011 may be greater than 0.04 mg/kg/day in male and female monkeys. Additionally, we propose that further studies are needed to identify the molecular signals, which are related to KMRC011-induced adverse effects.

Facile conversion of ATP-binding RNA aptamer to quencher-free molecular aptamer beacon.

We have developed RNA-based quencher-free molecular aptamer beacons (RNA-based QF-MABs) for the detection of ATP, taking advantage of the conformational changes associated with ATP binding to the ATP-binding RNA aptamer. The RNA aptamer, with its well-defined structure, was readily converted to the fluorescence sensors by incorporating a fluorophore into the loop region of the hairpin structure. These RNA-based QF-MABs exhibited fluorescence signals in the presence of ATP relative to their low background signals in the absence of ATP. The fluorescence emission intensity increased upon formation of a RNA-based QF-MAB·ATP complex.

Factors associated with intention to be tested for HIV among men who have sex with men in a country with a very low HIV prevalence.

This study examined factors associated with the intention to take an HIV test among men who have sex with men (MSM) in South Korea. An internet website-based survey was conducted among users of the only and largest online MSM website between 20 July 2016, and 20 August 2016. A total of 2915 participants completed the survey and answered questions related to sociodemographic information, health behaviors, sexual behaviors, and HIV testing history. Of these, 2587 (88.7%) participants responded as having an intention to take an HIV test. A multivariable logistic regression analysis revealed the following as having reduced the intention to undergo HIV testing: very good subjective health status and no sexual interactions during the last 6 months (Adjusted odds ratios [AOR] 0.45 and 0.54, respectively). In contrast, increased intention to take an HIV test was associated with being 20-29 years old, 30-39 years old, not paying or receiving money for sex, having a history of HIV testing, and taking an HIV test once per 12 months (AOR 2.64, 2.13, 1.54, 1.81, and 2.17, respectively). In conclusion, HIV testing among MSM in this study was associated with age, subjective health status, sex(es) of one's sexual partner(s) during the last 6 months, sexual risk behaviors, HIV testing history, and undergoing regular HIV testing.

Relative contribution of clear cells and principal cells to luminal pH in the mouse epididymis.

While spermatozoa undergo epididymal maturation, they remain quiescent thanks to the establishment of a low luminal pH. This study is aimed at determining how epithelial cells lining the epididymal lumen work together to maintain and regulate this acidic milieu. In particular, we examined the relative contribution of clear cells (CCs) and principal cells (PCs) to this process. Functional analysis in the mouse cauda epididymidis (Cd) perfused in vivo showed that the pH of a control solution remained constant at pH 6.6 after perfusion through the Cd lumen. In contrast, the pH of both an acidic (pH 5.8) and alkaline (pH 7.8) perfusate was progressively restored toward the control acidic pH. Pharmacological studies indicated the contribution of cystic fibrosis transmembrane regulator, previously shown to be present in the apical membrane of PCs, to the recovery from an acidic pH of 5.8. In addition, we found that CCs and PCs equally contribute to the recovery from an alkaline of 7.8, via the H+ pumping vacuolar ATPase (V-ATPase) located in CCs, and the Na+/H+ exchanger type 3 (NHE3) located in PCs. Immunofluorescence labeling showed apical membrane accumulation of the V-ATPase in CCs at pH 7.8, and its internalization at pH 5.8 compared to pH 6.6. Immunofluorescence showed expression of NHE3, but absence of NHE2, in PCs located in the Cd. RT-PCR and western blotting showed expression of NHE3 in all epididymal regions. Luminal 8-(4-chlorophenylthio)adenosine 3΄,5΄-cyclic monophosphate (cpt-cAMP) partially inhibited luminal pH recovery from pH 7.8. However, cpt-cAMP induced an increase in V-ATPase apical membrane accumulation at this pH. Cell fractionation studies showed the apical accumulation of NHE3 from intracellular vesicles at pH 7.8 versus 6.6, and prevention of this effect by cpt-cAMP. These results indicate the participation of both CCs and PCs in the regulation of luminal pH in the epididymis. Our study also shows the dual role of PCs in HCO3− and H+ secretion, and that this switch from base to acid secretion depends on the luminal environment. Characterization of the respective roles of CCs and PCs in the regulation of the optimal luminal condition for epididymal sperm maturation should provide new frameworks for the evaluation and treatment of male infertility.

Sex chromosome-dependent differential viability of human spermatozoa during prolonged incubation.

STUDY QUESTION: Are there significant differences in the ability of X chromosome-bearing (X) spermatozoa and Y chromosome-bearing (Y) spermatozoa to survive incubation under stressful conditions? SUMMARY ANSWER: Y spermatozoa are more vulnerable to stress than their X counterparts depending on culture period and temperature, and show higher expression of apoptotic proteins. WHAT IS KNOWN ALREADY: The primary sex ratio is determined by there being an equal number of spermatozoa carrying X and Y chromosomes. This balance can be skewed by exposure to stressful environmental conditions such as changes in pH, pollutants or endocrine disruptors. However, less is known about the ability of sperm carrying either sex chromosome to withstand environmental stress. STUDY DESIGN, SIZE, DURATION: The difference in survival between X and Y spermatozoa was evaluated by measuring motility, viability and Y:X chromosome ratio during incubation for 5 days, at three temperatures (4, 22 and 37°C), and three pH conditions (6.5, 7.5 and 8.5). To identify the critical factors that determine the survival of X and Y bearing spermatozoa, we analysed the expression levels of apoptosis-related proteins (Bcl, Bax and Caspase-3), as well as the extent of DNA damage under a subset of conditions. PARTICIPANTS/MATERIALS, SETTING, METHODS: Semen samples were obtained by masturbation from normozoospermic donors after 3 days of sexual abstinence. Four samples with >60% motility from different donors were mixed to obtain sufficient semen and eliminate sampling-related bias. Data are presented as mean ± SD of three independent experiments. Mean age of donors was 28.7 ± 3.2 years. MAIN RESULTS AND THE ROLE OF CHANCE: In total, 58 489 spermatozoa were scored. The viability of Y spermatozoa was lower after exposure to different temperatures and culture periods than that of X spermatozoa (P < 0.05). Increased expression of apoptotic proteins in live Y spermatozoa was observed, despite the addition of tocopherol to the culture medium (P < 0.05). LIMITATIONS, REASONS FOR CAUTION: Spermatozoa were cultured in vitro during the treatment period. It is difficult to extrapolate the observed lifespan differences to spermatozoa survival in vivo. The experiments were replicated only three times. WIDER IMPLICATIONS OF THE FINDINGS: The prolonged survival of X spermatozoa under stressful conditions might lead to shifts in the ratio of male-to-female births. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by the National Research Foundation of Korea (NRF) grant funded by the Korean government (MEST) (no. NRF-2014R1A2A2A01002706). The authors declare no competing financial interests.

pH-Responsive quencher-free molecular beacon systems containing 2'-deoxyuridine units labeled with fluorene derivatives.

Three fluorescent nucleosides-UFL, UAF, and UDAF, containing fluorene, 2-aminofluorene, and 2-dimethylaminofluorene units, respectively, covalently attached to 2'-deoxyuridine-have been incorporated into the central positions of oligodeoxynucleotides (ODNs) to examine the effects of their flanking bases (FBs) and pH on the emission properties upon hybridization with fully matched and single-base-mismatched targets. The ODN containing UFL and cytosine-FBs in the pH range from 5.5 to 8.0 and the ODN containing UDAF and cytosine-FBs under slightly acidic conditions (pH 6.0-6.5) exhibited dramatic increases in fluorescence only upon duplex formation with their fully matched target DNAs.

Discovery of predictive biomarkers for litter size in boar spermatozoa.

Conventional semen analysis has been used for prognosis and diagnosis of male fertility. Although this tool is essential for providing initial quantitative information about semen, it remains a subject of debate. Therefore, development of new methods for the prognosis and diagnosis of male fertility should be seriously considered for animal species of economic importance as well as for humans. In the present study, we applied a comprehensive proteomic approach to identify global protein biomarkers in boar spermatozoa in order to increase the precision of male fertility prognoses and diagnoses. We determined that l-amino acid oxidase, mitochondrial malate dehydrogenase 2, NAD (MDH2), cytosolic 5'-nucleotidase 1B, lysozyme-like protein 4, and calmodulin (CALM) were significantly and abundantly expressed in high-litter size spermatozoa. We also found that equatorin, spermadhesin AWN, triosephosphate isomerase (TPI), Ras-related protein Rab-2A (RAB2A), spermadhesin AQN-3, and NADH dehydrogenase [ubiquinone] iron-sulfur protein 2 (NDUFS2) were significantly and abundantly expressed in low-litter size spermatozoa (>3-fold). Moreover, RAB2A, TPI, and NDUFS2 were negatively correlated with litter size, whereas CALM and MDH2 were positively correlated. This study provides novel biomarkers for the prediction of male fertility. To the best of our knowledge, this is the first work that shows significantly increased litter size using male fertility biomarkers in a field trial. Moreover, these protein markers may provide new developmental tools for the selection of superior sires as well as for the prognosis and diagnosis of male fertility.