Transcriptomic analysis of mechanism underlying the effect of induced molting on semen quality and reproductive performance in aged Houdan roosters.

The reproductive performance of breeder roosters has significant economic importance in the poultry industry. Breeder roosters have severely reduced semen quality with age and will be at risk of culling in the following years. In order to extend the use of breeder roosters, we drew on the induced molting model of hens and selected 35 Houdan roosters aged 50 wk for induced molting. By comparing the body weight, testicular weight, semen quality, and reproductive performance before and after induced molting, we found that induced molting could restore the body weight and testicular weight to the levels before molting (P > 0.05). At the same time, it significantly improved sperm motility (P < 0.05) and also improved reproductive performance such as fertilization rate and hatching rate. To further reveal the mechanism underlying the effects of induced molting on semen quality and reproductive performance in aged Houdan roosters, we collected testes from 3 periods: 1 d before fasting (F0), 15 d after fasting (F15), and 32 d after recovery feeding (R32) for transcriptome sequencing analysis. A total of 5,671 genes were detected in F0, F15, and R32, and trend analysis of the 5,671 differential genes showed 2 significant trends (profile 5 and profile 2). KEGG enrichment analysis of the genes in the 2 profiles, revealed significantly enriched pathway regulation of actin cytoskeleton. In the regulation of actin cytoskeleton pathway, we found a protein kinase gene (SRC) and a senescence gene (ROCK2). SRC was highly expressed at F15, leading to the phosphorylation of key substrates, which in turn disrupted the Sertoli cell spermatid connection and the spermiogenesis process, resulting in no mature spermatozoa produced from F15, SRC expression was inhibited at R32, the expression level was reduced, and mature spermatozoa reappeared. The senescence gene ROCK2 was highly expressed at F15 compared to F0 and R32, which may have been responsible for inducing senescence atrophy in the testes.

Counteract severe heat stress by including different forms of zinc in the rabbit bucks' diet.

The harmful influences of global warming on rabbit reproduction and industry attract global attention. Zinc (Zn) is an important trace element with a wide list of functions in the male reproductive system. The aim of this study was to estimate the effects of different forms of zinc supplementation, as organic (Zn methionine), nano (nano Zn oxide) as indirect way to minimized it impact on environment, and inorganic (Zn sulphate) on physiological parameters, semen quality, anti-oxidative status, hormonal profiles of male rabbits subjected to server heat stress. Thirty-six V-line bucks (6-7 months old, 2842.29 ± 34.46 g weight) were randomly distributed to 4 groups, bucks in 1st group (control group, Con) fed basal diet without Zn supplementation, the 2nd, 3rd and 4th groups fed basial diet with 30 mg/kg of zinc methionine (Zn-Met), nano zinc oxide (ZnO-NPs), and zinc sulphate (Zn-S), respectively for a period of 10 weeks suffered from severe heat stress of environmental ambient temperature (over 32 of temperature humidity index, THI). Semen samples were collected and evaluated for volume, pH, motility, concentration, viability, initial fructose, and seminal plasma antioxidant concentration. Liver, kidney function, hormonal and antioxidant profile were measured in blood serum. The results revealed that, compared to control, all forms of Zn supplementation used significantly improved kidney function (creatinine), serum antioxidant (SOD and CAT), physiological parameters, especially on 1st month of the experiment, and seminal plasma antioxidant (SOD and CAT) of heat stressed bucks. Likewise, semen quality in terms of sperm concentration, sperm viability, and initial fructose enhanced significantly (P ≤ 0.05) by ZnO-NPs supplementation. Zinc methionine supplementation significantly improved liver function and decreased seminal plasma TBARs. Treatments with Zn-Met and ZnO-NPs increased seminal TAC and blood testosterone levels with reduced blood cortisol levels compared to other groups. Severe heat stress could be counteract by inclusion Zn with studied forms Zn-Met and ZnO-NPs at recommended dose (30 mg/kg diet) to improve semen quality and antioxidant status.

Total fertilization failure after ICSI: insights into pathophysiology, diagnosis, and management through artificial oocyte activation.

BACKGROUND: Total fertilization failure (TFF) is the failure of all metaphase II oocytes to fertilize in ART cycles. The phenomenon represents a known cause of infertility, affecting 1-3% of ICSI cycles. Oocyte activation deficiency (OAD) is the leading cause of fertilization failure, attributed to sperm- or oocyte-related issues, although until recently little attention has been given to oocyte-related deficiencies. Different strategies for overcoming TFF have been proposed in clinical settings, mainly using artificial oocyte activation (AOA) by calcium ionophores. Typically, AOA has been blindly applied with no previous diagnosis testing and, therefore, not considering the origin of the deficiency. The scarcity of data available and the heterogeneous population subjected to AOA make it challenging to draw firm conclusions about the efficacy and safety of AOA treatments. OBJECTIVE AND RATIONALE: TFF leads to an unexpected, premature termination of ART, which inflicts a substantial psychological and financial burden on patients. This review aims to provide a substantial update on: the pathophysiology of fertilization failure, focusing both on sperm- and oocyte-related factors; the relevance of diagnostic testing to determine the cause of OAD; and the effectiveness and safety of AOA treatments to overcome fertilization failure. SEARCH METHODS: Relevant studies were identified in the English-language literature using PubMed search terms, including fertilization failure, AOA, phospholipase C zeta (PLCζ), PLCZ1 mutations, oocyte-related factors, wee1-like protein kinase 2 (WEE2) mutations, PAT1 homolog 2 (PATL2) mutations, tubulin beta-8 chain (TUBB8) mutations, and transducin-like enhancer protein 6 (TLE6) mutations. All relevant publications until November 2022 were critically evaluated and discussed. OUTCOMES: Fertilization failure after ART has been predominantly associated with PLCζ deficiencies in sperm. The reason relates to the well-established inability of defective PLCζ to trigger the characteristic pattern of intracellular Ca2+ oscillations responsible for activating specific molecular pathways in the oocyte that lead to meiosis resumption and completion. However, oocyte deficiencies have recently emerged to play critical roles in fertilization failure. Specifically, mutations have been identified in genes such as WEE2, PATL2, TUBB8, and TLE6. Such mutations translate into altered protein synthesis that results in defective transduction of the physiological Ca2+ signal needed for maturation-promoting factor (MPF) inactivation, which is indispensable for oocyte activation. The effectiveness of AOA treatments is closely related to identifying the causal factor of fertilization failure. Various diagnostic tests have been developed to determine the cause of OAD, including heterologous and homologous tests, particle image velocimetry, immunostaining, and genetic tests. On this basis, it has been shown that conventional AOA strategies, based on inducing the calcium oscillations, are highly effective in overcoming fertilization failure caused by PLCζ-sperm deficiencies. In contrast, oocyte-related deficiencies might be successfully managed using alternative AOA promoters that induce MPF inactivation and meiosis resumption. Such agents include cycloheximide, N,N,N',N'-tetrakis(2-pyridylmethyl)ethane-1,2-diamine (TPEN), roscovitine, and WEE2 complementary RNA. In addition, when OAD is caused by oocyte dysmaturity, applying a modified ovarian stimulation protocol and trigger could improve fertilization. WIDER IMPLICATIONS: AOA treatments represent a promising therapy to overcome fertilization failure caused by sperm- and oocyte-related factors. Diagnosing the cause of fertilization failure will be essential to improve the effectiveness and safe utilization of AOA treatments. Even though most data have not shown adverse effects of AOA on pre- and post-implantation embryo development, the literature is scarce on the matter concerned and recent studies, mainly using mice, suggest that AOA might cause epigenetic alterations in the resulting embryos and offspring. Until more robust data are available, and despite the encouraging results obtained, AOA should be applied clinically judiciously and only after appropriate patient counseling. Currently, AOA should be considered an innovative treatment, not an established one.

Sexual behaviour response, testicular development and semen quality of New Zealand white rabbit bucks as influenced by dietary chitosan.

The present study was conducted to determine the effects of dietary chitosan supplementation on sexual behaviour responses, testicular development, and semen quality traits of New Zealand White (NZW) rabbit bucks. Twenty-four 5-week-old rabbit bucks were used in this experiment. Animals were grouped into four equal experimental groups: the control group was fed only on a basal diet, whereas the other groups were fed the basal diet supplemented with three levels of chitosan at 0.2, 0.4, or 0.6 g/kg, respectively. Also, bucks that received chitosan at 0.2 and 0.4 g/kg had a significantly earlier time of sexual libido (p ≤ .05) and had significantly higher ejaculate volume and sperm concentration than other groups (p ≤ .001). Furthermore, basic and sexual behaviours were significantly improved in bucks fed chitosan at 0.2 and 0.4 g/kg compared with other groups. Therefore, it could be concluded that using chitosan at 0.2 and 0.4 g/kg enhanced sexual behaviour, improved semen quality, and reproductive efficiency in the NZW rabbit bucks.

Administration of carbetocin-a long-acting oxytocin analogue-before sperm collection by transrectal ultrasound-guided massage of the accessory sex glands in bucks (Capra hircus) and ibexes (Capra pyrenaica).

Transrectal ultrasonic-guided massage of the accessory sex glands (TUMASG) is a technique that allows collecting semen requiring few electrical stimuli or even no pulse. A long-acting analogue of oxytocin (carbetocin, 0.1 mg) was i.v. administered before TUMASG in 10 conscious bucks (Experiment 1) and 10 anaesthetized Iberian ibexes (Experiment 2) to shorten the time of semen collection, decrease the number of electrical stimuli and/or improve the semen quality. The ejaculated volume, concentration, quality parameters and kinetics variables of the sperm were determined in fresh semen. The time length of the procedures and the number of electric pulses applied were recorded. Furthermore, stress response indicators (number of vocalizations in Experiment 1; heart and respiratory rates, rectal temperature, cortisol levels, totals proteins and neutrophil-to-lymphocyte ratio in Experiment 2) were documented. In bucks, the administration of carbetocin tended to shorten the time needed for semen collection but no-showed differences in the fresh seminal quality. In the Iberian ibexes, there were no significant differences between groups in the time length of procedures or in the number of animals that ejaculated. Carbetocin administration only reduced the respiratory rate, did it modify fresh semen characteristics in ibexes. In conclusion, the administration of carbetocin did not appear as a useful tool to improve welfare during semen collection with TUMASG or semen quality in conscious bucks and anaesthetized ibexes, having only slight advantages related to the procedure.

Boar semen cryopreserved with trehalose-containing liposomes: disaccharide determination and rheological behaviour.

This study aimed to detect intracellular trehalose in boar sperm that were cryopreserved with liposomes and conduct an analysis of its effects on some characteristics of thawed sperm, including rheological properties. First, soybean lecithin cholesterol-based liposomes were produced and characterized in the presence of 300 mM trehalose. Next, semen samples were frozen in two freezing media: a control medium with 300 mM trehalose and an experimental medium supplemented with 300 mM trehalose and 10% liposomes, both of which were thawed and then studied to ascertain their integrity, motility, rheological response, and trehalose quantities by testing two methods of spermatic lysis via high-performance liquid chromatography with an evaporative light-scattering detector (HPLC-ELSD). The results found spherical liposomes measuring 357 nm that were relatively stable in an aqueous medium and had an entrapment efficiency of 73%. An analysis of the cryopreserved ejaculates showed that their viability and motility did not significantly differ between groups (P > 0.05). The viscous response of the samples was influenced by the extracellular medium rather than by the freezing-thawing process, which resulted in a loss of interaction between the cells and cryoprotectants. Finally, intracellular trehalose levels were determined using HPLC-ELSD, with no differences observed (P > 0.05) when comparing both sperm lysis methods. The use of liposomes with trehalose appears to be a promising option for boar semen cryopreservation, with a marked effect on rheological properties. The proposed HPLC-ELSD method was effective for measuring trehalose in cryopreserved cell samples.

Sperm flow cytometric parameters, antioxidant status, and testicular histomorphology in roosters fed diets supplemented with camphor.

Genetic selection based on the high growth rate and consequently high slaughter weight in broiler chickens has caused many problems in broiler breeders. A negative correlation between growth and reproductive traits has declined semen quality and fertility in roosters. The present study aimed to evaluate the effects of camphor levels on some reproductive parameters included semen parameters, antioxidant status, and testicular development in broiler breeder roosters. Thirty-five ROSS 308 broiler breeder roosters were divided into 5 groups to receive camphor (C) levels: C0, C50, C250, C750, and C1000 ppm for 12 consecutive weeks (31-43 wk). Body weight, seminal volume, sperm concentration, and percentage of live and morphologically normal sperm were not affected by diets (P > 0.05), however, significantly were changed by bird age over the experiment (P < 0.05). Semen quality factor (SQF) significantly was affected by both diets and age (P < 0.05). Mitochondrial activity, apoptotic-like changes, and DNA fragmentation were improved in the groups fed camphor levels compared to the control group (P < 0.05). Testes weight (left, right, and combined weights) and gonadosomatic index were increased linearly by the camphor supplementation (P < 0.05). The serum activity of glutathione peroxidase (GPX) was not affected by treatments, however, superoxide dismutase (SOD) activity, ferric ion reducing antioxidant power (FRAP), and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity were significantly higher in C50, C250, and C750, respectively (P < 0.05). The lower malondialdehyde (MDA) content in the testes and liver samples was observed in C750 (P < 0.05). Excluding the number of Sertoli cells and blood vessels, other histomorphological traits of testes showed one of the linear or quadratic responses to the camphor levels (P < 0.05). It can be concluded that camphor as an antioxidant source may improve reproduction performance in roosters.

Astaxanthin improves preserved semen quality at 17°C by enhancing sperm antioxidant and motility properties.

Porcine sperm is rich in polyunsaturated fatty acids; therefore, it is highly susceptible to oxidative damage during storage. Inhibition of oxidative stress during preservation is essential for maintaining sperm motility. Astaxanthin is a potent antioxidant used in the cosmetic and pharmaceutical industries. This study aimed to explore the effect of supplementing astaxanthin as an extender of porcine semen preservation dilutions at 17°C. Various concentrations of astaxanthin were added to diluted porcine semen at 17°C. We performed computer-assisted semen analysis, evaluation of plasma membrane integrity and acrosome integrity, and measurement of total antioxidant activity, malondialdehyde (MDA) content, reactive oxygen species levels, superoxide dismutase (SOD) activity, catalase (CAT) activity, glutathione peroxidase (GSH-PX) activity and sperm motility parameters. Compared with the control group, the addition of 0.25 µg/ml astaxanthin group significantly improved sperm motility parameters stored on the fifth day; these were increased levels of sperm SOD, GSH-PX and CAT (p < .05), increased sperm adenosine trisphosphate and lactate dehydrogenase levels and decreased sperm MDA levels (p < .05). These findings suggest that adding 0.25 µg/ml of astaxanthin improves the quality of porcine semen stored at 17°C. Our findings provide theoretical support for developing new protective agents critical for preserving pig semen at 17°C.

Dietary carob fruit (Ceratonia siliqua L.) supplementation improves spermatogenesis, semen quality and embryonic death via antioxidant effect in aging broiler breeder roosters.

Rooster semen contains high content of long-chain polyunsaturated fatty acids; therefore, spermatozoa are very susceptible to lipid peroxidation. Carob fruit (CF) contains abundant amounts of flavonoid known as natural antioxidants and semen enhancers. The purpose of the study was to investigate the effect of dietary supplementation of CF on semen characters, blood parameters, immune response, testicular histology, and reproductive performance of aging broiler breeder roosters. Thirty-two 47-week-old Ross-308 breeder roosters were randomly assigned to either control or supplemented (CF, 0 or 1 g/kg diet) dietary treatment for 14 continuous weeks. Semen, blood and testis samples were collected at the end of trial. Furthermore, the rate of fertility and hatchability were evaluated using 440 natural mated hens. The sperm kinematic values and progression ratios, seminal antioxidant enzymes (superoxide dismutase and glutathione peroxidase) activity, and total antioxidant capacity of semen and blood were increased, and malondialdehyde concentration in semen and blood were decreased by dietary CF supplementation (P < 0.01). The treatment had no effect on the seminal sperm concentration, blood parameters and immune response to Newcastle disease virus vaccine. The testes weight, seminal tubes spermatozoids number, spermiogenesis index and repopulation index, were improved by dietary supplementation (P < 0.05). Fertility and hatchability rate of roosters in the CF group were insignificant. In conclusion, CF supplementation in aging roosters' diet enhanced the sperm motility, seminal and blood antioxidant capacity, and testicular spermatogenenic indexes.

Utility of carboxylated poly L-lysine for the liquid storage of bottlenose dolphin semen.

Assisted reproduction techniques are required to maintain a genetically diverse captive population of bottlenose dolphins. These techniques include semen preservation, and liquid storage has been proposed as a suitable alternative to cryopreservation, but the optimum conditions, in terms of temperature, duration, and media, have yet to be fully established. The aim of this study, therefore, was to determine the optimum temperature for the liquid storage of dolphin semen during a 14-day period and the usefulness of carboxylated poly-L-lysine (CPLL) as an additive to the semen extender used for the liquid storage. The semen was collected from a mature male dolphin housed at the Kagoshima Aquarium, Japan, transferred into a Beltsville (BF5F) extender, and analyzed for motility and characteristics after five-fold dilution. The optimum temperature was determined by evaluating sperm viability after liquid storage at 4, 17, or 36 °C, and the usefulness of CPLL was evaluated at concentrations of 0%, 0.5%, 1.0%, 2.0%, and 3.0% (v/v) at the optimum temperature. Sperm stored at 4 â„ƒ had a greater motility maintenance compared with samples stored at 17 or 36 â„ƒ. The most efficacious storage regimen at various time points occurred when there was addition of CPLL at 1.0% (v/v) in terms of sperm motility and other relevant determinations, with this storage approach having greater efficacy that samples stored without CPLL. The most efficacious processes for preserving bottlenose dolphin sperm functions is storage at 4 °C and with there being semen extender supplementation of 1% CPLL.

Factors affecting the production of quality ejaculates from boars.

Production of acceptable quality ejaculates in boars is dependent upon the Sertoli cell population established before puberty and how effectively these cells function after sexual maturity. In general, factors affecting Sertoli cell mitosis tend to have a two-fold greater effect on sperm production compared with those affecting spermatogenesis. Birthweight is a reliable indicator of in utero testicular development and prepubertal growth rates are positively correlated with testis size and sperm production after sexual maturity. Colostrum intake and pre-weaning nutrition account for much of the variation associated with quality and quantity of ejaculates and represent opportunities to further enhance lifetime sperm production. Interactions between young boars and humans, shortly after weaning, have important effects on spermatogenesis after sexual maturity and need to be studied further. The seasonal effect on depression in semen quality is the most significant factor affecting production of acceptable quality ejaculates after puberty. Ambient temperatures, greater than those of the thermoneutral zone, have both acute and chronic effects that compromise all aspects of the male reproductive axis. Identification of genes associated with heat-tolerant phenotypes holds promise for addressing this challenge, especially in light of the current trend in global warming. Supplementation of vitamins, minerals and other compounds have positive effects on sperm production during periods in which other stressors, especially heat stress, are present and is an important mitigation strategy. Recent information on housing conditions and boar usage patterns indicate these cause relatively minor changes in sperm production, overall, but for some males can have significant, long-term effects.

Improving the quality and in vitro fertilization rate of frozen-thawed semen of buffalo (Bubalus bubalis) bulls with the inclusion of vitamin B12 in the cryopreservation medium.

This study was designed to evaluate the effects of vitamin B12 in cryopreservation medium on frozen-thawed semen of buffalo (Bubalus Bubalis) bulls. Semen from five bulls (fertility-proven) were diluted in five aliquots not supplemented (control), or supplemented with 1, 2, 4, or 5 mg/mL of vitamin B12 and evaluated using the Computer Assisted Sperm motion Analysis, antioxidant enzymes, lipid peroxidation (LPO), reactive oxygen species (ROS), ATP concentrations, and in vitro fertilization rate (%). Sperm progressive motility, rapid velocity (%), mitochondrial potential, and acrosome integrity were greater (P < 0.05) with supplementation of 4, and 5 mg/mL vitamin B12 than the control sample. Similarly, compared with the control, samples with 5 mg/mL vitamin B12 supplementation had markedly greater average-path, straight-line, and curved-line velocities (µm/sec). Semen samples supplemented with 2, 4 and 5 mg/mL vitamin B12 had greater concentrations of GPx (U/mL) and SOD (U/mL), whereas LPO (µM/mL) was less (P < 0.05) compared with the control sample. Seminal plasma ROS concentrations (104/25 × 106) were less in the 5 mg/mL vitamin B12 supplemented than control sample. Semen samples supplemented with 5 mg/mL of vitamin B12 had greater concentrations of ATP than control and the 1 mg/mL vitamin B12 supplemented sample. Semen samples supplemented with 5 mg/mL of vitamin B12 had greater plasmalemma and DNA integrities (%) than the control sample. In summary, vitamin B12 supplementation augments semen quality, as evidenced by values for CASA variables, antioxidant enzymes, and ATP concentrations, which may occur as a consequence of inhibition in LPO and ROS production by buffalo spermatozoa.

Effects of dietary alpha-lipoic acid supplementation on the seminal parameters and fertility potential in aging broiler breeder roosters.

High levels of polyunsaturated fatty acids in avian sperm cause more susceptibility to lipid peroxidation. Aging in roosters reduces the antioxidant capacity of sperm and thus fertility. The purpose of this study was to investigate the effects of different levels of alpha-lipoic acid (ALA) as a feed supplement to improve the semen quality and fertility parameters of aged broiler breeder roosters and identification of its most effective level. A total of forty-two roosters at 45 wk of age were randomly assigned to 7 treatments (0, 15, 40, 70, 95, 120, and 145 mg ALA/bird per day) for 8 wk. Semen parameters and body weight were assessed biweekly, and testosterone plasma levels were determined in the 8th wk of the experimental period. Artificial insemination was performed at the end of the experiment to evaluate the fertility potential. The dietary administration of ALA had no significant effects on body weight, semen volume, average path velocity, linearity, straightness, wobble, the amplitude of lateral head displacement, beat-cross frequency, sperm concentration, morphology, plasma testosterone level, fertility, or hatchability (P > 0.05). Alpha-lipoic acid supplementations resulted in a significant decrease in seminal malondialdehyde concentration and immotile (type D) sperms (P < 0.05). The total motility, progressive motility (types A + type B sperms), curvilinear velocity, straight-line velocity, viability, and membrane integrity of sperm improved with ALA dietary supplementations (P < 0.05). With increasing ALA levels, improvement in semen parameters had an incremental trend until the level of 95 mg ALA. Thus, 95 mg dietary ALA as an antioxidant supplement can improve semen quality of aging breeder roosters while higher doses resulted in no further improvement.

Cryopreservation of dog semen in a Tris extender with two different 1% soybean preparations compared with a Tris egg yolk extender.

Egg yolk is widely used as a cryoprotectant in dog semen extenders, but there is a risk of contamination with animal pathogens. In addition, egg yolk may vary in composition, making it difficult to standardize the extender. Lecithin is an animal protein-free alternative to egg yolk for semen cryopreservation. Recently, it was shown that 1% of soybean lecithin type II-S was better than 2% for freezing canine semen. The aim of the study was to compare two different types of soybean lecithin, with egg yolk as a control. Ejaculates from eight dogs were divided into three equal parts and diluted with a Tris-based extender, containing either 20% egg yolk, 1% soybean lecithin Type II-S or 1% soybean lecithin Type IV-S. The samples were then frozen. Sperm motility was evaluated by computer-assisted sperm analysis (CASA), acrosome integrity (FITC-PNA/PI) and sperm membrane integrity (SYBR-14/PI) post-thaw, as well as after 2 and 4 hr incubation at 37°C. Post-thaw sperm chromatin structure assay and plasma membrane integrity were evaluated by flow cytometry. Total motility, sperm plasma membrane integrity and acrosome integrity were significantly better in the egg yolk extender than in the two soybean lecithin-based extenders. Individual motility post-thaw differed more than in the fresh samples, illustrating individual differences in tolerance to the cryostress. The DNA Fragmentation Index (% DFI) was significantly lower in the Tris egg yolk (TEY) extender compared to any of the soybean-based extenders. The number of high green stained spermatozoa were significantly higher in Type IV-S compared to the control TEY extender. In conclusion, egg yolk was superior to the two lecithin-based extenders to cryopreserve canine semen.

Efecto de la suplementación del medio de incubación con extracto de Berberis microphylla (calafate) sobre la funcionalidad espermática en semen bovino descongelado / Effect of supplementation of incubation medium with Berberis microphylla (calafate) extract on sperm functionality in thawed bovine semen

RESUMEN: En el semen criopreservado, los procesos de congelación/descongelación y posterior manipulación, dañan las células espermáticas provocando disminución de la capacidad fecundante de los espermatozoides descongelados. Estos procesos han sido asociados con el estado de estrés oxidativo (EO) inducido por altos niveles de especies reactivas de oxígeno (EROS), causando daño a la función y estructura espermática. Los espermatozoides descongelados pueden ser protegidos de este daño, con la adición de antioxidantes (AO) al medio de incubación. El fruto de Calafate (Berberis microphylla G. Forst.) posee una alta capacidad antioxidante, lo que hace interesante investigar el efecto de sus componentes antioxidantes en estos procesos biotecnológicos especialmente postdescongelación. El objetivo de este estudio fue determinar el efecto de la suplementación de extracto liofilizado de fruto de Calafate (ELC), sobre la calidad espermática post-descongelación. Previamente se caracterizó el ELC, determinando la actividad antioxidante y metabolitos como fenoles y antocianinas; posteriormente, espermatozoides de bovino descongelados fueron incubados en un medio base suplementado con diferentes concentraciones de ELC. Post-incubación se evaluó la motilidad progresiva; la viabilidad e integridad de la membrana plasmática (SYBR14- PI) y acrosomal (FITC-PNA/PI) y la peroxidación lipídica (BODIPY) por citometría de flujo. La caracterización de ELC demostró que tanto la actividad antioxidante como los fenoles y antocianinas incrementan concomitante con el aumento de la concentración de ELC. La adición de ELC al medio de incubación, dependiendo de la concentración y tiempo de incubación, sería eficaz en proteger la motilidad, viabilidad e integridad de la membrana plasmática y disminuir la lipoperoxidación en los espermatozoides de bovino descongelados.

SUMMARY: In cryopreserved semen, the freezing/thawing process following of manipulation, damage the sperm cell, decreasing the fertilizing capacity of the thawed sperm; being one of the main factors of this damage the oxidative stress. The sperm once thawed can be protected from this damage, with the addition of antioxidants to the incubation medium. The Calafate fruit (Berberis microphylla G. Forst.) has a high antioxidant capacity, making it an interesting resource for investigating the effect of its antioxidant components on biotechnological processes. The objective of this study was to determine the effect of supplementation of Calafate fruit lyophilized extract (ELC) on sperm quality. The lyophilized extract of the Calafate fruit was characterized, determining the antioxidant activity and metabolites such as phenols and anthocyanins; subsequently, thawed bovine sperm were incubated in a medium supplemented with different concentrations of ELC. Post-incubation, progressive motility was evaluated. By flow cytometry, the viability and integrity of the plasma (SYBR14-PI), and acrosomal (FITC-PNA / PI), as well as lipid peroxidation (BODIPY), was determined. The characterization of Calafate fruits lyophilized extract indicated that antioxidant activity, phenols and anthocyanins increased concomitantly with the increase of dose extract used. The addition of ELC to the incubation medium, depending on the concentration and incubation time, would be effective to protect motility, viability and integrity of the plasma membrane and decreased lipid peroxidation in thawed bovine sperm.

Efecto de la borra de café sobre la biológica espermática: aproximación in vitro / Effect of spent coffee grounds on sperm biology: in vitro approach

OBJETIVO: El objetivo de este trabajo fue establecer el efecto de la borra de café sobre la movilidad y los parámetros funcionales de los espermatozoides humanos in vitro. MATERIALES Y MÉTODOS: La borra de café, un subproducto obtenido en establecimientos especializados en la preparación de café soluble a base de grano, se diluyo en tampón fosfato salino y se mezcló en proporciones iguales con las muestras de semen de 16 voluntarios aparentemente sanos. A cada muestra se le determinó el efecto sobre la movilidad espermática en función del tiempo (30, 60, 90 y 120 minutos, n=16) y sobre los parámetros funcionales (n=6) por medio de citometría de flujo: potencial de membrana mitocondrial, producción de especies reactivas de oxígeno y lipoperoxidación de la membrana espermática. RESULTADOS: La incubación de los espermatozoides con la borra de café evidencio un cambio positivo en la movilidad espermática. Adicionalmente, la incubación con la borra de café incremento significativamente el potencial de membrana mitocondrial en los espermatozoides. CONCLUSIÓN: La borra de café, seguramente debido a los compuestos antioxidantes, afecta positivamente la movilidad espermática aumentando el potencial de membrana mitocondrial. Por lo tanto, esto es un paso inicial en la búsqueda de un suplemento de origen natural que aumente la calidad seminal.

OBJECTIVE: The objective of this work is to establish the effect of spent coffee grounds on the motility and functional parameters of human spermatozoa, in vitro. MATERIALS AND METHODS: Spent coffee grounds, a by-product obtained in specialized establishments in the preparation of soluble coffee based on grain, was diluted in saline phosphate buffer and mixed in equal proportions with semen samples from 16 apparently healthy volunteers. Each sample was determined the effect on sperm motility as a function of time (30, 60, 90 and 120 minutes, n=16) and on functional parameters (n=6) by means of flow cytometry: mitochondrial membrane potential, reactive oxygen species production and membrane lipoperoxidation. RESULTS: The incubation of the spermatozoa with the spent coffee grounds showed a positive change in sperm motility. Additionally, incubation with spent coffee grounds significantly increased the mitochondrial membrane potential in human sperm cells. CONCLUSION: Spent coffee grounds, probably due to antioxidant compounds, positively affects sperm motility by increasing mitochondrial membrane potential. Therefore, this is an initial step in the search for a supplement of natural origin that increases seminal quality.

Economic opportunities of using crossbreeding and sexing in Holstein dairy herds.

With the increasing availability of sexed semen, farms have the opportunity to select genetically superior dams to produce their replacement animals and to produce crossbred calves for beef production of higher economic value than the remainder of the herd. However, higher costs and reduced fertility of sexed semen complicate the decision of when and to what extent sexed semen should be applied in a herd. The objective of this study was to explore the economically optimal utilization of sexed semen and crossbreeding among North Rhine-Westphalian dairy farms in a holistic single-farm model. For the analysis, we derived a representative sample of farms from Latin Hypercube sampling based on the observed distribution of farm characteristics from official North Rhine-Westphalian Farm Structure Survey data. Market- and technology-related input parameters such as output prices and sexed semen accuracy and fertility were included in the sampling procedure. Modeling results of the systematic sensitivity analysis were evaluated in a statistical meta-model. We found that the profit-maximizing utilization of sexed semen and crossbreeding was highly heterogeneous among the farms. Farms with lower stocking densities, <2 livestock units (LU)/ha, were generally found to produce excess heifers for sale, whereas farms with stocking densities >2 LU/ha were producing crossbred calves and using sexed semen only to produce replacement animals. On average, female-sexed dairy semen was used on 25.3% of all inseminations. Beef semen (both sexed and conventional) for producing crossbred calves was used in an average of 21.5% of the inseminations. The combination of sexed semen and crossbreeding increased profits from €0 to €568 per cow per year, with an average of €79.42 per cow per year. Farms characterized by low stocking densities (<2 LU/ha) and above-average replacement rates (>40%) were found to have higher profit increases as a result of selling more heifers from the use of sexed semen. Overall, sexed semen and crossbreeding adoption were most sensitive to stocking density and average cow longevity, as well as to additional costs for sexed semen and sexed semen accuracy. Our results show the potential of modern breeding technologies to improve dairy farm profits and the need to judge their profitability in the light of farm-specific production settings.

The effect of modifiable lifestyle factors on semen quality.

INTRODUCTION AND OBJECTIVES: To examine the association between lifestyle factors (body mass index, smoking, alcohol consumption, coffee intake, physical activity, sauna and cell phone usage, wearing tight-fitting underwear), and conventional semen parameters. MATERIALS AND METHODS: 1311 participants who attended the Andrology Clinic were included in the study. All participants were separated into two groups as men with normozoospermia and dysspermia. All participants answered a questionnaire which contains questions about the modifiable lifestyle factors. The total risk scores were calculated after all the positive lifestyle factors had been counted. RESULTS: Men with normozoospermia and dysspermia consisted of 852 (65.0%) and 459 (35.0%) participants respectively. A negative relationship between the wearing of tight underwear and having normal semen parameters was detected between the two groups (p=0.004). While going to a sauna regularly was negatively related to semen concentration, wearing tight underwear was also related to both lower motility, normal morphology as well as semen concentration (p<0.05). While the total score of all participants was 5.22±1.34 point, there were no statistical differences between the two groups (p=0.332). It was found that having 3 more or fewer points was not related to any type of semen parameters and results of a spermiogram. CONCLUSION: The clinicians should give advice to infertile male patients about changing their risky lifestyle, for infertility, to a healthy lifestyle for fertility. Better designed studies, with larger sample sizes using conventional semen analysis with sperm DNA analysis methods, should be planned to identify the possible effects of lifestyle factors on semen quality.

Effects of zinc deficiency on impaired spermatogenesis and male infertility: the role of oxidative stress, inflammation and apoptosis.

Zinc (Zn) is necessary for the normal function of the male reproductive system and spermatozoa. Although influences of zinc deficiency on impaired spermatogenesis and male infertility have been widely considered, the molecular and cellular mechanisms of these abnormalities are not well understood. General abnormalities, including hypogonadism, Leydig cells damage, deficiency of sex hormone production and impaired spermatogenesis, as well as inflammation, antioxidant depletion, sperm death and male infertility can be observed during zinc deficiency. However, it is not obvious which pathways are relevant to the pathogenesis of zinc deficiency. Oxidative stress (OS) induced by reactive oxygen species is likely as the main mechanism of zinc deficiency which is associated with sperm DNA fragmentation, decrease in sperm membrane integrity, apoptosis, depletion of antioxidants, and consequently poor sperm quality and male infertility. Therefore, identification of these pathways will give valuable information regarding the mechanisms of zinc deficiency on the male reproductive system and the potential way for developing a better clinical approach. In this review, we aim to discuss the proposed cellular and molecular mechanisms of zinc deficiency on the male reproductive system, the importance of OS and mechanisms by which zinc deficiency induces OS and depletion of other antioxidants.

Effects of supplementation of tris-egg yolk extender with different sugars and antioxidants on freezability of ram semen.

This study was designed to determine the effects of the combined addition of different levels of certain sugars (trehalose, sucrose and raffinose) and antioxidants (vitamin E, C and taurine), in Tris-egg yolk extender on frozen-thawed ram semen parameters. Semen samples were collected from five healthy, mature and fertile Iranian Afshari rams, twice a week for 8 weeks. Selected samples were pooled and diluted with a Tris-egg yolk extender containing different levels of sugars and antioxidants. In Experiment 1, different levels of trehalose (0, 50 and 100 mM) were tested with different levels of taurine (0, 25 and 50 mM), vitamin E and C (0, 1 and 2 mM). In Experiment 2, different levels of sucrose (0, 60 and 80 mM) were tested with different levels of taurine (0, 25 and 50 mM), vitamin E and C (0, 1 and 2 mM). In Experiment 3, different levels of raffinose (0, 5, 10 mM) were tested with different levels of taurine (0, 25 and 50 mM), and vitamin E and C (0, 1 and 2 mM). In Experiment 4, the selected extenders of experiments 1, 2 and 3 were compared statistically with control (no selected sugar and antioxidant) extender. The results of experiments 1, 2 and 3 revealed that the highest frozen-thawed sperm parameters were recorded for the selected extenders containing 100 mM trehalose +2 mM vitamin E (T100E2), 60 mM sucrose + 2 mM vitamin E (S60E2) and 10 mM raffinose + 2 mM vitamin E (R10E2), respectively. The results of experiment 4 revealed that the post-thaw sperm total motility in T100E2 (62.41 ± 2.41%), S60E2 (59.52 ± 1.91%) and R10E2 (58.33 ± 2.00%) was higher than that of the control extender (46.00 ± 1.79%; P ≤ 0.05). Similarly, the progressive sperm motility in T100E2 (57.18 ± 1.96%), S60E2 (57.49 ± 1.94%) and R10E2 (55.03 ± 2.99%) was also higher than that of the control extender (41.20 ± 1.70%; P ≤ 0.05). Post-thaw sperm viability in selected extenders of T100E2 (65.20 ± 2.67%), S60E2 (62.00 ± 2.07%) and R10E2 (61.80 ± 2.46%) was higher than that of control extender (51.00 ± 1.88%; P ≤ 0.05). In conclusion, the addition of 100 mM trehalose, 60 mM sucrose and 10 mM raffinose combined with 2 mM vitamin E in Tris-egg yolk extender significantly improved frozen-thawed ram semen parameters.