Effect of intrafollicular administration of PGE2 or PGF2α in early estrus on ovulation, hemorrhagic anovulatory follicles formation, progesterone secretion and pregnancy outcome in the mare.

This experiment was performed to evaluate whether intrafollicular treatment of PGE2 or PGF2α administered in early estrus would induce normal ovulation, progesterone production (Experiment 1) and pregnancy (Experiment 2). In Experiment 1, mares in estrus after 2 days of endometrial edema were injected in all largest dominant follicles (28-35 mm in diameter) with 0.5 mL of sterile water containing 500 µg PGE2 (n = 6), 125 µg PGF2α (n = 6) or placebo (n = 7) (Hour 0). Ultrasound examinations were performed daily, until ovulation or anovulation was detected, and daily blood samples were taken for 8 days. In Experiment 2, mares with a dominant follicle ≥35 mm after at least three days of slight-to-moderate endometrial edema, were injected with 500 µg PGE2 diluted in 0.5 mL of sterile water for injection in the follicle (PGE2 group; n = 9 mares and 11 dominant follicles). No puncture was performed in the control group (n = 9 mares and 11 dominant follicles). Mares from both groups were inseminated. In Experiment 1, all mares (6/6) in the PGE2 group ovulated within 24 h of treatment. The mean interval from intrafollicular injection to ovulation was shorter (P < 0.001) in PGE2 mares (24 ± 0 h) than in control mares (77 ± 9 h). Mares from the PGF2α group developed hemorrhagic anovulatory follicles (HAF) more often (7/7) than control mares (2/7); P < 0.05). The progesterone concentration in mares from the PGF2α group was lower (P < 0.004) than control mares in the early post-ovulatory period. The first significant increase in post-ovulatory progesterone concentration occurred earlier (P < 0.05) in mares from the control group than in mares from the PGF2α and PGE2 groups. In Experiment 2, more mares from the control group (7/9, 78 %) became pregnant than from the PGE2 group (2/9, 22 %) (P = 0.015). In conclusion, PGE2 alone induced follicle collapse in all treated mares within 24 h of administrations, while PGF2α blocked ovulation and induced formation of HAFs. However, the post-ovulatory rise in progesterone production was delayed and the fertility reduced in mares with ovulation induced by PGE2 compared to control mares.

Effects of Zinc, Mercury, or Lead on [3H]MK-801 and [3H]Fluorowillardiine Binding to Rat Synaptic Membranes.

Glutamate (Glu) is considered the most important excitatory amino acid neurotransmitter in the mammalian Central Nervous System. Zinc (Zn) is co-released with Glu during synaptic transmission and interacts with Glutamate receptors and transporters. We performed binding experiments using [3H]MK-801 (NMDA), and [3H]Fluorowillardine (AMPA) as ligands to study Zn-Glutamate interactions in rat cortical synaptic membranes. We also examined the effects of mercury and lead on NMDA or AMPA receptors. Zinc at 1 nM, significantly potentiates [3H]MK-801 binding. Lead inhibits [3H]MK-801 binding at micromolar concentrations. At millimolar concentrations, Hg also has a significant inhibitory effect. These effects are not reversed by Zn (1 nM). Zinc displaces the [3H]FW binding curve to the right. Lead (nM) and Hg (µM) inhibit [3H]FW binding. At certain concentrations, Zn reverses the effects of these metals on [3H]FW binding. These specific interactions serve to clarify the role of Zn, Hg, and Pb in physiological and pathological conditions.

New insight on the role of localisation in the electronic structure of the Si(111)(7 × 7) surfaces.

New angle-resolved photoelectron spectroscopy (ARPES) data, recorded at several different photon energies from the Si(111)(7 × 7) surface, show that the well-known S1 and S2 surface states that lie in the bulk band gap are localised at specific (adatom and rest atom) sites on the reconstructed surface. The variations in the photoemission intensity from these states as a function of polar and azimuthal emission angle, and incident photon energy, are not consistent with Fermi surface mapping but are well-described by calculations of the multiple elastic scattering in the final state. This localisation of the most shallowly bound S1 state is consistent with the lack of significant dispersion, with no evidence of Fermi surface crossing, implying that the surface is not, as has been previously proposed, metallic in character. Our findings highlight the importance of final state scattering in interpreting ARPES data, an aspect that is routinely ignored and can lead to misleading conclusions.

Two Synchronous Neonatal Tumors: An Extremely Rare Case.

We report a case of a newborn with two synchronous tumors-sialoblastoma and hepatoblastoma-diagnosed at 20 weeks of gestation by magnetic resonance imaging (MRI) and ultrasonography (US). The aim of this study was to describe the management of this case together with a review of the literature. Our patient had a large facial tumor associated with extremely high alpha-fetoprotein levels. Diagnosis of the tumors was made by surgical biopsy, showing typical features in both. Sialoblastoma is a potentially aggressive tumor. In our case, the Ki67 index in the sialoblastoma was between 20 and 30%, indicating a possibly unfavorable behavior. The infant underwent surgery and chemotherapy in different steps. Complete surgical resection with clean margins is considered to be the best treatment option for sialoblastoma. Only four similar cases were previously reported. Timely management by a multidisciplinary team is essential in these difficult cases. In our patient, outcome was good at the time of this report.

CO2 fluxes in the Northeast Atlantic Ocean based on measurements from a surface ocean observation platform.

The seasonal and spatial variability of the CO2 system parameters and CO2 air-sea exchange were studied in the Northeast Atlantic Ocean between the northwest African coastal upwelling and the oligotrophic open-ocean waters of the North Atlantic subtropical gyre. Data was collected aboard a volunteer observing ship from February 2019 to February 2020. The seasonal and spatial variability of CO2 fugacity in seawater (fCO2,sw) was strongly driven by the seasonal temperature variation, which increased with latitude and was lower throughout the year in coastal regions where the upwelling and offshore transport was more intense. The thermal to biological effect ratio (T/B) was approximately 2, with minimum values along the African coastline related to higher biological activity in the upwelled waters. The fCO2,sw increased from winter to summer by 11.84 ± 0.28 µatm°C-1 on the inter-island routes and by 11.71 ± 0.25 µatm°C-1 along the northwest African continental shelf. The seasonality of total inorganic carbon normalized to constant salinity of 36.7 (NCT) was studied throughout the region. The effect of biological processes and calcification/dissolution on NCT between February and October represented >90% of the reduction of inorganic carbon while air-sea exchange described <6%. The seasonality of air-sea CO2 exchange was controlled by temperature. The surface waters of the entire region acted as a CO2 sink during the cold months and as a CO2 source during the warm months. The Canary basin acted as a net sink of -0.26 ± 0.04 molC m-2 yr-1. The northwest African continental shelf behaved as a stronger sink at -0.48 ± 0.09 molC m-2 yr-1. The calculated average CO2 flux for the entire area was -2.65 ± 0.44 TgCO2 yr-1 (-0.72 ± 0.12 TgC yr-1).

Reaffirmation of the importance of follow-up ultrasound studies in patients with high D-dimers and clinical suspicion of vein thrombosis.

BACKGROUND: Venous thromboembolism is a common disease seen in the emergency department and a cause of high morbidity and mortality, constituting a major health problem. OBJECTIVES: To assess the potential benefit of follow-up ultrasound of patients who attended the emergency department with suspected superficial venous thrombosis or deep venous thrombosis and were found to have an initial negative whole-leg (or arm) ultrasound study. METHODS: This retrospective study included patients aged 18 years or older who were consecutively referred to a thrombosis clinic from the emergency department, with abnormal D-dimer test and moderate to high pre-test probability of deep venous thrombosis (Well's score ≥ 1), but a negative whole-leg (or arm) ultrasound. Demographic characteristics, symptom duration, laboratory and ultrasound data were recorded. At one-week follow-up, an experienced physician repeated ultrasound, and recorded the findings. RESULTS: From January 2017 to April 2018, 54 patients were evaluated. The mean age was 66.8 years (SD 15.0) and 63% were women. The average D-dimer was 2159.9 (SD 3772.0) ng/mL. Ultrasound abnormalities were found in 12 patients (22.2%; 95% confidence interval of 12.5 to 36.0%), with 4 patients having proximal deep venous thrombosis, distal deep venous thrombosis in 2 patients and superficial venous thrombosis in 6 patients. We did not find any significant differences in demographic characteristics, venous thromboembolism risk factors or laboratory parameters between patients with negative and positive follow-up ultrasound. CONCLUSIONS: These preliminary findings suggest that a negative whole-leg (or arm) ultrasound in addition to an abnormal D-dimer in moderate to high deep venous thrombosis pretest probability patients, might be an insufficient diagnostic approach to exclude suspected deep venous thrombosis or superficial venous thrombosis. Confirmation of this higher than expected prevalence would support the need to repeat one-week ultrasound control in this population.

Effect of ration level on growth performance, body composition, intermediary metabolism and serum parameters in juvenile Patagonian blennie Eleginops maclovinus.

Eleginops maclovinus is an endemic species of the southern cone with beneficial physiological characteristics for aquaculture. However, this species has a low growth rate under captive conditions, and the optimal feed ration together with the metabolic process is unknown. This study aimed to determine the optimum feed ration during 90 days based on growth performance, body composition, intermediary metabolism, and serum parameters. For this, fish were randomly assigned to rations of 0.5, 1, 2 and 4% of body weight. No fish mortality was registered, but all fish, developed fatty liver. The results of weight, length, growth performance (WGR, SGR), and body parameters (HSI, VSI and K) followed a similar pattern, with the lowest values observed in the 0.5% and no significant differences between rations of 1, 2 and 4%. The feed intake and feces increased with the feed ration. However, the percentage of food consumed by the fish decreased with the ration size and the feed conversion ratio was lowest in the 1% ration. Total serum proteins and calcium were lowest in the 0.5% ration and presented no differences in the rations 1, 2 and 4%, while triglyceride content was significantly different only between the rations of 0.5 and 4%. Blood cortisol levels were significantly higher in the rations of 0.5 and 1%, and decreased in rations of 2 and 4%. The lipids, fiber, and energy of the total body mass increased with the feed ration, while dry matter, proteins, and ash of the body decreased to higher feed ration. In liver, triglyceride and protein levels decreased with a larger feed ration, amino acids increased in the rations of 0.5 and 4%, while glucose levels increased in rations of 2 and 4%. Liver enzymes Glucose-6-phosphate dehydrogenase and Glutamate dehydrogenase increased their activity at a higher feed ration, while Glycogen Phosphorylase, Aspartate aminotransferase and 3-Hydroxyacyl-CoA dehydrogenase presented their highest enzymatic activity only in the 4% ration. Fructose-1,6-bisphosphate showed low enzymatic activity in rations of 2 and 4% and Glycerol-3-phosphate dehydrogenase was significantly different only between the ration of 0.5 and 1%. Finally, our results suggests that optimal feed rations for E. maclovinus corresponds to 1% since this ration size produces the highest growth and metabolism with a minimum loss of food and feces present in the environment. Additionally, we recommend to reduce the percentage of fat in the diet to avoid the development of fatty liver.

Síndrome De Kinsbourne: Papel De Rituximab / Kinsbourne Syndrome: Role Of Rituximab

Resumen El Síndrome de Kinsbourne también conocido como "síndrome de opsoclonus-mioclonus" (SOM) o "síndrome de los ojos danzantes" se considera una enfermedad rara y poco frecuente. Se caracteriza por una triada clásica: movimientos oculares rápidos, irregulares, multidireccionales (opsoclonus), movimientos mioclónicos en tronco, cara y/o extremidades y ataxia. La etiología es diversa pudiendo presentarse tanto de origen paraneoplásico como no paraneoplásico e idiopática. Fundamentalmente el tratamiento se realiza con inmunosupresores (corticoides), posteriormente inmunoglobulinas inespecifícas endovenosas y rituximab endovenoso. Procedemos a presentar un caso clínico de un niño de 4 años el cuál presentó dicho síndrome debido a una infección por Klebsiella pneumoniae.

Abstract: Kinsbourne syndrome, also known as "Opsoclonus-myoclonus syndrome" or "Dancing eyes syndrome" is a rare condition characterized by rapid, irregular, multi-directional eye movements (opsoclonus), myoclonic movements in the trunk, face and/or limbs and ataxia. It occurs in children aged between 6 and 36 months. Its etiology may be paraneoplasic (neuroblastoma), non-paraneoplasic (infectious processes) or idiopathic. Regardless of its etiology, immunosuppressive drugs have been used in order to reduce the formation of antibodies possibly involved in the pathophysiology. We report the case of a four years-old boy with this syndrome secondary to an infectious urinary illness. He had ataxia, opsoclonus, upper limbs myoclonus, irritability and altered sleep pattern. Neuroblastoma was ruled out. Initial management was done with methylprednisolone pulses, followed by oral prednisolone and intravenous gammaglobulin. He started with rituximab with a good response. He had progressive clinical improvement, and is currently asymptomatic with no sequelae. In these patients a paraneoplasic syndrome should always be ruled out.

[Gastrostomy tubes: indications and infectious complications in a tertiary hospital]. / Sondas de gastrostomía: indicaciones y complicaciones infecciosas en un hospital terciario.

OBJECTIVE: Gastrostomy tube is the best option for long-term enteral nutrition. Among its limitations, infections represent the most frequent minor complication. Our aim is the knowledge of the number and type of gastrostomy tubes and its main indication in our hospital. In addition, prevalence of infectious complications was studied paying attention to the main etiologic agents and their antibiotic susceptibility. METHODS: Observational retrospective study from January 2010 to July 2015 through the electronic clinical history and the clinical microbiology laboratory software. Identification and antibiotic susceptibility of clinically significant isolates from patients with suspicion of gastrostomy tube infection have been analysed. RESULTS: Percutaneous endoscopic gastrostomy was performed in 203 patients (70.5%) and surgical gastrostomy in 85 (29.5%). The main reason identified for starting enteral nutrition through gastrostomy tube was malignant neoplasy, above all, lip, oral cavity and pharynx cancer (11.8%) and that from digestive organs (8.7%). Global prevalence of gastrostomy tube infection was 16.6%. The most common bacterial pathogens isolated were Staphylococcus aureus (21.3%), Pseudomonas aeruginosa (13.1%), and Escherichia coli (9.8%). The percentage of multi resistant isolates was 3.1%. CONCLUSIONS: Gastrostomy tube indications and type, and also, prevalence and microorganisms isolated from culture in infectious complications are similar to those described previously in the literature. The study allows the adaptation of the antibiotic prophylaxis and empirical antibiotic treatment thanks to the knowledge of the etiologic agents and their antibiotic susceptibility.

Identification, characterization and modulation of ferritin-H in the sub-Antarctic Notothenioid Eleginops maclovinus challenged with Piscirickettsia salmonis.

Ferritin is a major iron storage protein essential not only in the infectious process, but also in any circumstance generating oxidative stress. In this study, the cDNA coding sequence of ferritin-H was obtained from the sub-Antarctic Notothenioid fish Eleginops maclovinus through transcriptomic analysis of the head kidney. This sequence contained a 534 bp open reading frame that coded for a 177 amino acid protein with a molecular weight of 20,786.2 Da and a theoretical pI of 5.56. The protein displayed a region of iron putative response elements in the 5'UTR, two putative ferritin iron-binding region signatures, and seven characteristic amino acids with ferroxidase functions. Phylogenetic analysis related this sequence to ferritin-H sequences of other Antarctic Notothenioid fish, sharing 96.61% similarity. Constitutive gene expression analysis in different organs revealed increased ferritin-H gene expression in the gills, spleen, muscle, and liver. After infection with two bacterial strains of Piscirickettsia salmonis (LF-89 and Austral-005), ferritin-H was differentially expressed depending on bacterial strain and tissue. This study provides relevant information towards understanding the iron metabolism of a sub-Antarctic Notothenioid fish.

Birth interval and stunting in children under five years of age in Peru (1996-2014).

BACKGROUND: Examine the evolution of stunting in <5 years of age in Peru and its relationship with birth interval (BI), and associated factors from 1996 to 2014. METHODS: Data were extracted from the National Demographic and Health Survey of Peru (DHS) for the years 1996, 2000, 2005, 2010 and 2014 on children <5 years of age for stunting (height / age ≤ 2DS). The main independent variable of interest was BI dichotomized by interval between date of last and penultimate birth (≤24 months vs. >24 months) and others sociodemographic variables. Crude and adjusted odds ratios with 95% confidence intervals were estimated in logistic regression for the effect of BI and other variables on stunting by each survey year. RESULTS: Between 1996 and 2014, stunting declined in both BI groups: ≤24 months: 35.8% to 21.8%; >24 months: 29.5% to 14.3%. BI was associated with stunting after adjustment for other variables in each survey except 2005. Of the other factors, sex, birth order (fourth child or more), maternal education and poverty were independently associated with stunting in all survey years. Residence in rural areas and in Amazon forest and highland regions was associated with stunting 1996 and 2000. Maternal age was not independently associated with stunting. CONCLUSIONS: While stunting showed a decreasing trend from 1996 to 2014, birth interval exceeding 24 months exerted a protective effect on stunting across the years surveyed. Poverty, low maternal education and high birth order were associated with stunting in all survey years.

Mitochondrial ATP is required for the maintenance of membrane integrity in stallion spermatozoa, whereas motility requires both glycolysis and oxidative phosphorylation.

To investigate the hypothesis that oxidative phosphorylation is a major source of ATP to fuel stallion sperm motility, oxidative phosphorylation was suppressed using the mitochondrial uncouplers CCCP and 2,4,-dinitrophenol (DNP) and by inhibiting mitochondrial respiration at complex IV using sodium cyanide or at the level of ATP synthase using oligomycin-A. As mitochondrial dysfunction may also lead to oxidative stress, production of reactive oxygen species was monitored simultaneously. All inhibitors reduced ATP content, but oligomycin-A did so most profoundly. Oligomycin-A and CCCP also significantly reduced mitochondrial membrane potential. Sperm motility almost completely ceased after the inhibition of mitochondrial respiration and both percentage of motile sperm and sperm velocity were reduced in the presence of mitochondrial uncouplers. Inhibition of ATP synthesis resulted in the loss of sperm membrane integrity and increased the production of reactive oxygen species by degenerating sperm. Inhibition of glycolysis by deoxyglucose led to reduced sperm velocities and reduced ATP content, but not to loss of membrane integrity. These results suggest that, in contrast to many other mammalian species, stallion spermatozoa rely primarily on oxidative phosphorylation to generate the energy required for instance to maintain a functional Na+/K+ gradient, which is dependent on an Na+-K+ antiporter ATPase, which relates directly to the noted membrane integrity loss. Under aerobic conditions, however, glycolysis also provides the energy required for sperm motility.

Quasi-chemical approximation for polyatomic mixtures.

The statistical thermodynamics of binary mixtures of polyatomic species was developed based on a generalization in the spirit of the lattice-gas model and the quasi-chemical approximation (QCA). The new theoretical framework is obtained by combining: (i) the exact analytical expression for the partition function of non-interacting mixtures of linear k-mers and l-mers (species occupying k sites and l sites, respectively) adsorbed in one dimension, and its extension to higher dimensions; and (ii) a generalization of the classical QCA for multicomponent adsorbates and multisite-occupancy adsorption. This process is analyzed using the partial adsorption isotherms corresponding to both species of the mixture. Comparisons with analytical data from Bragg-Williams approximation (BWA) and Monte Carlo simulations are performed in order to test the validity of the theoretical model. Even though a good fitting is obtained from BWA, it is found that QCA provides a more accurate description of the phenomenon of adsorption of interacting polyatomic mixtures.

Safety cost management in construction companies: A proposal classification.

BACKGROUND: Estimating health and safety costs in the construction industry presents various difficulties, including the complexity of cost allocation, the inadequacy of data available to managers and the absence of an accounting model designed specifically for safety cost management. Very often, the costs arising from accidents in the workplace are not fully identifiable due to the hidden costs involved. OBJECTIVE: This paper reviews some studies of occupational health and safety cost management and proposes a means of classifying these costs. METHODS: We conducted an empirical study in which the health and safety costs of 40 construction worksites are estimated. RESULTS: A new classification of the health and safety cost and its categories is proposed: Safety and non-safety costs. CONCLUSIONS: The costs of the company's health and safety policy should be included in the information provided by the accounting system, as a starting point for analysis and control. From this perspective, a classification of health and safety costs and its categories is put forward.

Significant discharge of CO2 from hydrothermalism associated with the submarine volcano of El Hierro Island.

The residual hydrothermalism associated with submarine volcanoes, following an eruption event, plays an important role in the supply of CO2 to the ocean. The emitted CO2 increases the acidity of seawater. The submarine volcano of El Hierro, in its degasification stage, provided an excellent opportunity to study the effect of volcanic CO2 on the seawater carbonate system, the global carbon flux, and local ocean acidification. A detailed survey of the volcanic edifice was carried out using seven CTD-pH-ORP tow-yo studies, localizing the redox and acidic changes, which were used to obtain surface maps of anomalies. In order to investigate the temporal variability of the system, two CTD-pH-ORP yo-yo studies were conducted that included discrete sampling for carbonate system parameters. Meridional tow-yos were used to calculate the amount of volcanic CO2 added to the water column for each surveyed section. The inputs of CO2 along multiple sections combined with measurements of oceanic currents produced an estimated volcanic CO2 flux = 6.0 10(5) ± 1.1 10(5 )kg d(-1) which is ~0.1% of global volcanic CO2 flux. Finally, the CO2 emitted by El Hierro increases the acidity above the volcano by ~20%.

Reduction in hospitalisations and emergency department visits for frail patients with heart failure: Results of the UMIPIC healthcare programme.

OBJECTIVES: Heart failure (HF) is a disease with high morbidity and mortality. We evaluated the usefulness of the Comprehensive Management Units for Patients with HF (Unidades de Manejo Integral para Pacientes con IC [UMIPIC]) programme. PATIENTS AND METHOD: We analysed the patient data from the UMIPIC programme, which was recorded in the HF registry (RICA) of the Spanish Society of Internal Medicine. We compared emergency department visits and hospitalisations for any cause and for HF during the year prior to inclusion in the programme against those that occurred during the subsequent follow-up year, using the chi-squared test. RESULTS: A total of 258 patients (mean age, 80years; 51.9% women) were included in the study. During the previous year, there were 693 hospitalisations for all causes and 174 hospitalisations during the follow-up (75% reduction, P<.001). There were 613 hospitalisations for HF during the previous year and 92 during the follow-up (85% reduction, P<.001); 655 vs. 302 in terms of emergency department visits for any cause (53.9% reduction, P<.001); and 440 vs. 120 for emergency department visits for HF (72% reduction, P<.001). There were no significant differences in the number of hospitalisations or emergency department visits for causes other than HF. CONCLUSIONS: The UMIPIC programme based on the comprehensive care of elderly patients with HF and comorbidity reduces the rate of hospital readmissions and emergency department visits during the first year of follow-up.

Juvenile, adult and late-onset systemic lupus erythematosus: a long term follow-up study from a geographic and ethnically homogeneous population.

OBJECTIVES: This paper aims to identify clinical and serological differences, damage accrual and mortality, in juvenile, adult and late-onset SLE. METHODS: We conducted our study with patients fulfilling SLE classification criteria taken from the Hospital Gregorio Marañon Autoimmune Systemic Rheumatic Diseases' Registry (1986 to 2012). Clinical characteristics, laboratory data and therapies used during the course of the disease were analysed with patients divided into 3 groups: juvenile-onset (≤ 18 years), adult-onset (19-50) and late onset (>50 years). RESULTS: Four hundred and forty-five patients were included. Renal disease and cutaneous manifestations were more frequent in the juvenile-onset group at disease onset. During follow-up, juvenile-onset group presented a higher incidence of renal disease, malar rash, Raynaud's phenomenon, cutaneous vasculitis, and neuropsychiatric manifestations than the other two groups. Arthritis and lymphopoenia were more frequent in the adult-onset group. Arterial hypertension and neoplasm were more frequent in the late-onset group. Low serum complement, anti-dsDNA, anti-U1RNP and anti-Sm antibodies were more common in the juvenile-onset group. Patients with late-onset SLE had more damage accrual. Thirty-seven patients (8.3%) died during the study. All-cause mortality was significantly higher in the late-onset group. Age at disease onset >50 years was an independent risk factor for damage accrual (OR, 2.2; 95%CI, 1.1-4.6; p=0.029) and mortality (OR, 2.6; 95%CI, 1.1-6.3; p=0.03). CONCLUSIONS: We found significant differences in clinical and serological profiles between juvenile, adult and late-onset SLE. The most significant of which was a higher prevalence of neuropsychiatric and renal complications as well as different autoantibody signatures for the juvenile-onset group.

Epigallocatechin-3-Gallate (EGCG) Reduces Rotenone Effect on Stallion Sperm-Zona Pellucida Heterologous Binding.

Stallion spermatozoa are highly dependent on oxidative phosphorylation for ATP production to achieve normal sperm function and to fuel the motility. The aim of this study was to evaluate the response of equine sperm under capacitating conditions to the inhibition of mitochondrial complex I by rotenone and to test whether epigallocatechin-3-gallate (EGCG), a natural polyphenol component of green tea, could counteract this effect. After 2-h incubation of stallion spermatozoa in modified Tyrode's medium, rotenone (100 nm, 500 nm and 5 µm) and EGCG (10, 20 and 60 µm), alone or in combination, did not induce any significant difference on the percentage of viable cells, live sperm with active mitochondria and spermatozoa with intact acrosome. The inhibition of complex I of mitochondrial respiratory chain of stallion sperm with rotenone exerted a negative effect on heterologous ZP binding ability. EGCG at the concentrations of 10 and 20 µm (but not of 60 µm) induced a significant increase in the number of sperm bound to the ZP compared with that for control. Moreover, when stallion sperm were treated with rotenone 100 nm, the presence of EGCG at all the concentrations tested (10, 20 and 60 µm) significantly increased the number of sperm bound to the ZP up to control levels, suggesting that this green tea polyphenol is able to reduce the toxicity of rotenone.

The Impact of Reproductive Technologies on Stallion Mitochondrial Function.

The traditional assessment of stallion sperm comprises evaluation of sperm motility and membrane integrity and identification of abnormal morphology of the spermatozoa. More recently, the progressive introduction of flow cytometry is increasing the number of tests available. However, compared with other sperm structures and functions, the evaluation of mitochondria has received less attention in stallion andrology. Recent research indicates that sperm mitochondria are key structures in sperm function suffering major changes during biotechnological procedures such as cryopreservation. In this paper, mitochondrial structure and function will be reviewed in the stallion, when possible specific stallion studies will be discussed, and general findings on mammalian mitochondrial function will be argued when relevant. Especial emphasis will be put on their role as source of reactive oxygen species and in their role regulating sperm lifespan, a possible target to investigate with the aim to improve the quality of frozen-thawed stallion sperm. Later on, the impact of current sperm technologies, principally cryopreservation, on mitochondrial function will be discussed pointing out novel areas of research interest with high potential to improve current sperm technologies.

Effect of overnight staining on the quality of flow cytometric sorted stallion sperm: comparison with tradtitional protocols.

Flow cytometry is considered the only reliable method for the separation of X and Y chromosome bearing spermatozoa in equines. The MoFlo SX DP sorter is highly efficient, allowing the production of foals of the desired sex. However, to achieve acceptable pregnancy rates the currently used protocol requires working with fresh semen obtained close to, or at, the sorting facility. An alternative protocol was tested during two consecutive breeding seasons. Fresh stallion semen was cooled for 20 h, during which staining with Hoechst 33342 took place. On the following day, this sample was flow sorted and compared with spermatozoa from the same ejaculate that had been sexed on the previous day. All sperm parameters evaluated remained unchanged when fresh sorted and refrigerated sorted semen were compared. Pre-sorting storage at 5°C did not alter sperm velocities nor kinetics, viability or membrane permeability, production of reactive oxygen species, mitochondrial membrane potential or DNA fragmentation index of the sorted sample. The findings open for the possibility of using semen from stallions housed far from the sorting facilities. Processed and stained sperm could be shipped refrigerated on the previous day, sorted and inseminated on the next day.