Exploring host-commensal-pathogen dynamics in cell line and organotypic human intestinal epithelial models.

Host and microbiome intricately interact in the ecosystem of the human digestive tract, playing a crucial role in our health. These interactions can initiate immune responses in the epithelial cells, which, in turn, activate downstream responses in other immune cells. Here, we used a CaCo-2 and a human intestinal enteroid (HIE) model to explore epithelial responses to both commensal and pathogenic bacteria, individually and combined. CaCo-2 cells were co-cultured with peripheral blood mononuclear cells, revealing downstream activation of immune cells. While both systems showed comparable cytokine profiles, they differed in their responses to the different bacteria, with the organoid system being more representative of responses observed in humans. We provide evidence of the pro-inflammatory responses associated with these bacteria. These models contribute to a deeper understanding of the interactions between the microbiota, intestinal epithelium, and immune cells in the gut, promoting advances in the field of host-microbe interactions.

Adecuación de la prescripción de hierro intravenoso en pacientes hospitalizados / Appropriateness of Intravenous Iron Prescription in Hospitalized Patients

Introducción: La ferropenia y la anemia ferropénica son condiciones clínicas frecuentes en la práctica clínica diaria, requiriendo terapia de reemplazo con hierro. El objetivo de este estudio fue evaluar la adecuación del tratamiento con hierro intravenoso de los pacientes ingresados e identificar las principales causas de prescripción inadecuada. Método: Estudio retrospectivo observacional. Se incluyeron pacientes adultos a los que se les prescribió hierro intravenoso durante el periodo de estudio. Se consideró el tratamiento como adecuado si el paciente presentaba ferropenia, bien por déficit absoluto (ferritina<100 ng/mL) o funcional de hierro (índice de saturación de transferrina sérica < 20 % y ferritina < 300 ng/ml). La variable principal del estudio fue el porcentaje de pacientes con prescripción adecuada de hierro intravenoso. Resultados: El estudio incluyó 250 pacientes. El uso del hierro fue adecuado en el 41,6 % (104) de los 250 pacientes analizados. La causa principal de inadecuación en un 46,4 % de pacientes, fue la falta de datos analíticos que corroboraran la ferropenia. Conclusiones: Nuestro centro presenta un déficit de adecuación de la prescripción de hierro intravenoso pese a la existencia de un protocolo de utilización del mismo. El principal punto de mejora sería la realización de un perfil analítico que permita el diagnóstico adecuado del déficit de hierro previo a la prescripción de hierro intravenoso. En algunas patologías, la inexistencia de guías clínicas con puntos de corte específicos para los marcadores analíticos indicativos de déficit de hierro en la bibliografía disponible puede contribuir a esta situación. (AU)

Introduction: Iron deficiency anemia and iron deficiency are usual clinical conditions in daily clinical practice, requiring iron replacement therapy. The study objective was to evaluate the appropriateness of intravenous iron treatment in hospitalized patients and to identify the main causes of inappropriate prescription. Method: This is a retrospective observational study. Adult patients who received intravenous iron therapy at their hospital during the study period were included. Appropriate treatment was considered when patient presented iron deficiency, either due to absolute (ferritin <100 ng/mL) or functional iron deficiency (transferrin saturation index < 20 % and ferritin < 300 ng/mL). The main variable of the study was patient percentage with adequate prescription or parenteral iron. Results: The study included 250 patients. The use of iron was adequate in 41.6 % (104) of the 250 patients analyzed. The main cause of inadequacy in 46.4 % of patients was the lack of analytical data which corroborated iron defi-ciency. Conclusions: Our center has a deficiency in the appropriateness of the prescription of parenteral iron despite the existence of a protocol for its use. The main point of improvement would be the creation of an analytical profile that allows the adequate diagnosis of iron deficiency prior to the prescription of intravenous iron. In some pathologies, the lack of clinical guidelines with specific cut-off points for analytical markers indicative of iron deficiency in the available literature may contribute to this situation. (AU)

Antioxidant potential of coffee husks in fresh pork sausage.

Coffee husks, a by-product of dry coffee processing, present a disposal problem in coffee-producing countries. Valorization of this residue is necessary to reduce its environmental impact and improve benefits to the producer. This study evaluated the antioxidant effect of coffee husks on physicochemical properties and sensory liking of fresh sausages packaged in aerobic (AEP) or modified atmosphere packaging (MAP) (20% CO2 + 80%N2). Fresh sausages were prepared with different antioxidants: no addition (control C), sodium nitrite (T2), sodium nitrite + sodium erythorbate + BHA/BHT blend (T3), sodium nitrite + coffee husk 1% (T4), sodium nitrite + coffee husk 2% (T5). Physicochemical properties (TBARs, carbonyl content, pH and instrumental color) were analyzed to evaluate the effect of added synthetic and natural antioxidants on fresh sausages. A sensory test (n = 100) was conducted to assess consumer liking of fresh sausages stored in AEP and MAP. The addition of coffee husks reduced lipid oxidation in fresh sausages, especially under MAP packaging, but did not affect carbonyl content. Consumers reported lower liking scores for products packed in MAP. The addition of coffee husks did not affect the degree of liking. Valorization of coffee husks as an antioxidant in fresh meat products is a viable natural option for the meat industry.

Olive Polyphenol Oxidase Gene Family.

The phenolic compounds containing hydroxytyrosol are the minor components of virgin olive oil (VOO) with the greatest impact on its functional properties and health benefits. Olive breeding for improving the phenolic composition of VOO is strongly dependent on the identification of the key genes determining the biosynthesis of these compounds in the olive fruit and also their transformation during the oil extraction process. In this work, olive polyphenol oxidase (PPO) genes have been identified and fully characterized in order to evaluate their specific role in the metabolism of hydroxytyrosol-derived compounds by combining gene expression analysis and metabolomics data. Four PPO genes have been identified, synthesized, cloned and expressed in Escherichia coli, and the functional identity of the recombinant proteins has been verified using olive phenolic substrates. Among the characterized genes, two stand out: (i) OePPO2 with its diphenolase activity, which is very active in the oxidative degradation of phenols during oil extraction and also seems to be highly involved in the natural defense mechanism in response to biotic stress, and (ii) OePPO3, which codes for a tyrosinase protein, having diphenolase but also monophenolase activity, which catalyzes the hydroxylation of tyrosol to form hydroxytyrosol.

Validation of new automated turbidimetric immunoassays for the measurement of haptoglobin and inter-α-trypsin inhibitor heavy chain H4 specific for the bovine species.

BACKGROUND: Good strategical programs are required for the early detection of disease even in the absence of evident clinical signs, which is crucial in satisfying animal welfare. Haptoglobin (Hp) and inter-α-trypsin inhibitor heavy chain H4 (ITIH4) are acute phase proteins and good biomarkers of early inflammation in cattle, with plasma levels that significantly increase after injury or infection. OBJECTIVES: We aimed to develop and validate two new immunoturbidimetric methods for Hp and ITIH4. METHODS: Species-specific antibodies were obtained and used to develop the immunoassays. For the Hp assay, antibodies were fixed to latex microparticles to enhance detection. The immunoassays were set up in an automated analyzer to carry out validation studies. Reference intervals were calculated using Reference Value Advisor. RESULTS: The Hp immunoturbidimetric method had a linear analytical range up to 0.40 mg/mL. The limit of detection (LoD) was 0.005 mg/mL, and the limit of quantification (LoQ) was 0.007 mg/mL. Total imprecision was less than 7%. Comparison with ELISA and single radial immunodiffusion (SRID) showed good correlation, whereas the comparison with the colorimetric method showed constant and proportional differences. The ITIH4 immunoassay showed linearity up to 5 mg/mL, and the LoD was 0.002 mg/mL. Total imprecision was less than 6%. Method comparison showed a good correlation with single radial immunodiffusion, both methods being equivalent. Bilirubin, triglycerides, and hemoglobin presented no interference in any of the assays. Reference intervals were 0.007-0.017 mg/mL for Hp and 0.2-0.7 mg/mL for ITIH4 in dairy cows 10 days before parturition. CONCLUSIONS: Immunoturbidimetric methods developed for Hp and ITIH4 can measure basal and increased levels of these proteins, showing adequate precision, accuracy, and robustness.

Características de pacientes con enfermedades inflamatorias inmunomediadas hospitalizados por infección por SARS-CoV-2 / Characteristics of Patients With Immune-Mediated Inflammatory Diseases Hospitalized for SARS-CoV-2 Infection

Fundamentos: Las enfermedades inflamatorias inmunomediadas (IMID) predisponen a un aumento del riesgo infeccioso al modificar la respuesta inmune del huésped, que resulta crucial para la resolución de la infección por SARS-CoV-2. Las últimas publicaciones indican que los pacientes con IMID y sus tratamientos de base no empeoran el pronóstico de la infección por SARS-CoV-2. Objetivos: Describir las características clínicas y la evolución de pacientes con IMID que requirieron ingreso hospitalario por infección por SARS-CoV-2. En segundo lugar, comparar las características clínicas y la evolución entre pacientes que requirieron ingreso hospitalario por infección por SARS-CoV-2 con IMID y aquellos que no la presentaban. Métodos: Estudio observacional de cohortes retrospectivo, que incluyó pacientes ingresados por sospecha de SARS-CoV-2, tratados según el criterio médico y los protocolos basados en la evidencia científica. La recogida de datos clínicos se realizó por descarga directa o mediante revisión manual de la historia clínica electrónica. El análisis estadístico determinó las diferencias de características y evolución clínica de la infección en pacientes con IMID. Resultados: De los 612 pacientes revisados, 23 padecían IMID y 9 de ellos fueron diagnosticados de infección por SARS-CoV-2. No se observó correlación entre infección por SARS-CoV-2 e IMID. Los pacientes con IMID presentaban mayor prevalencia de enfermedad cardiovascular y obesidad, aunque no significativamente. Asimismo, los pacientes con IMID no presentaron una evolución clínica durante el ingreso hospitalario diferente respecto al resto de pacientes. Conclusión: Las IMID y los tratamientos de las mismas no determinan el pronóstico del ingreso hospitalario de la infección por SARS-CoV-2.(AU)

Background: Immune-mediated inflammatory diseases (IMID) predispose to a higher infection risk by modifying the host's immune response, which acts as a key factor in SARS-CoV-2 infection resolution. Recent publications show that IMID patients and its treatments do not worsen the outcome of SARS-CoV-2 infection. Objectives: To describe the clinical characteristics and outcomes of patients with IMID who required hospital admission due to SARS-CoV-2 infection. Secondly, to compare clinical characteristics and outcomes between patients who required hospital admission due to SARS-CoV-2 infection with IMID and those who were not affected. Methods: We performed an observational retrospective cohort study, including admitted patients with suspected SARS-CoV-2 infection, treated according to medical criteria and local protocols based on the best available scientific evidence. Clinical data were collected from their electronical clinical history. Statistical analysis determined the differences in the characteristics and clinical outcome of the infection in IMID patients. Results: Of a total number of 612 revised patients, 23 had an IMID and 9 of them were positive for the SARS-CoV-2 infection. We did not observe a correlation between these two disorders. There was a higher frequency of obesity and cardiovascular disease among IMID patients, but without statistical significance. The clinical outcomes were no different between hospitalized IMID and non IMID patients. Conclusion: IMID and its treatments do not determine the outcome of patients admitted with SARS-CoV-2 infection.(AU)

Use of Tocilizumab in hospitalized patients during the first wave of the COVID-19 pandemic

Introduction: The aim of this study was to characterize the use of tocilizumab in a hospital. Method: Retrospective cohort study including adult SARS-CoV-2 infected inpatients with moderately severe infection (PaO2/FiO2 < 300). ORs for mortality and treatment success were calculated. Results: The tocilizumab group (n=18), presented 5 days of symptoms vs 7 days in the untreated group (n=71). Minimum PaO2/FiO2 was 147.5 (95%CI: 116.7 to 194.0) vs 255.6 (95%CI: 320.7 to 452.4) (p=0.01).No significant differences were found between the two groups concerning survival (OR=1.22; 95%CI: 0.38 to 3.92), nor treatment success (OR=0.46; 95%CI: 0.16 to 1.33). After adjusting for age, sex and total corticosteroid dose, OR for success was 0.18 (95%CI: 0.03 to 0.96), while mortality was not significant. Conclusions: The use of tocilizumab in moderately severe hospitalized patients could decrease the hyperinflammatory state preventing disease progression (AU)

Introducción: El objetivo del estudio fue estudiar la utilización de tocilizumab en un centro hospitalarioMétodo:Estudio de cohortes retrospectivo que incluyó pacientes ingresados adultos infectados por SARS-CoV-2, con una infección de gravedad moderada (PaO2/FiO2 < 300). Se calcularon las OR de mortalidad y éxito del tratamiento. Resultados: El grupo tocilizumab (n=18), tuvo 5 vs 7 días con síntomas en el grupo no tratado (n=71). El PaO2/FiO2 mínimo fue de 147,5 (IC95%: 116,7 a 194,0) versus 255,6 (IC95%: 320,7 a 452,4) (p=0,01).No se encontraron diferencias significativas entre ambos grupos en supervivencia (OR=1,22; IC95%: 0,38 a 3,92); ni en el éxito del tratamiento (OR=0,46; IC95%: 0,16 a 1,33). Tras ajustar por edad, sexo y dosis total de corticoide, OR de éxitos fue 0,18 (IC95%: 0,03 a 0,96), mientras que mortalidad no fue significativa. Conclusiones: El uso de tocilizumab en pacientes hospitalizados moderadamente graves podría disminuir su estado hiperinflamatorio (AU)

Prevalence and seasonality of viral respiratory infections in a temperate climate region: A 24-year study (1997-2020).

BACKGROUND: Few long-term reports have been published on the epidemiology of respiratory viruses despite their frequent involvement in extremely common infections. The aim here was to determine the frequency and distribution of respiratory viruses in a temperate climate area (Barcelona, Spain) throughout a 24-year period. METHODS: We collected data on all respiratory viruses detected from 1997 to 2020 in our institution. Clinical specimens were analyzed mainly by conventional techniques, and molecular techniques were also used. RESULTS: Of the 59,579 specimens analyzed, 21,382 (35.9%) were positive for at least one virus. The number of positive samples during cold months was significantly higher than in warm months. Respiratory virus infections were detected in patients of all ages, above all in children under 3 years of age, who were most frequently infected with the respiratory syncytial virus, whereas Influenza A virus predominated in the other groups, especially in adults. A clear demographic and seasonal pattern was established for some viruses. Circulation of other respiratory viruses during the FLUAV H1N1pdm09 and SARS-CoV-2 pandemics was observed. CONCLUSIONS: This long-term study provides new knowledge about the prevalence of respiratory viruses in a Mediterranean region. Throughout the study period, the frequency of some viruses remained constant, whereas others varied with the year. A clear demographic and seasonal pattern was established for some viruses. Patients suffering from severe respiratory infections should be examined for a range of respiratory viruses regardless of gender, age, or season.

Characteristics of patients with immune-mediated inflammatory diseases hospitalized for SARS-CoV-2 infection.

BACKGROUND: Immune-mediated inflammatory diseases (IMID) predispose to a higher infection risk by modifying the host's immune response, which acts as a key factor in SARS-CoV-2 infection resolution. Recent publications show that IMID patients and its treatments do not worsen the outcome of SARS-CoV-2 infection. OBJECTIVES: To describe the clinical characteristics and outcomes of patients with IMID who required hospital admission due to SARS-CoV-2 infection. Secondly, to compare clinical characteristics and outcomes between patients who required hospital admission due to SARS-CoV-2 infection with IMID and those who were not affected. METHODS: We performed an observational retrospective cohort study, including admitted patients with suspected SARS-CoV-2 infection, treated according to medical criteria and local protocols based on the best available scientific evidence. Clinical data were collected from their electronical clinical history. Statistical analysis determined the differences in the characteristics and clinical outcome of the infection in IMID patients. RESULTS: Of a total number of 612 revised patients, 23 had an IMID and 9 of them were positive for the SARS-CoV-2 infection. We did not observe a correlation between these two disorders. There was a higher frequency of obesity and cardiovascular disease among IMID patients, but without statistical significance. The clinical outcomes were no different between hospitalized IMID and non IMID patients. CONCLUSION: IMID and its treatments do not determine the outcome of patients admitted with SARS-CoV-2 infection.

Polyphenols and IUGR Pregnancies: Effects of the Antioxidant Hydroxytyrosol on Brain Neurochemistry and Development in a Porcine Model.

Supplementation of a mother's diet with antioxidants, such as hydroxytyrosol (HTX), has been proposed to ameliorate the adverse phenotypes of fetuses at risk of intrauterine growth restriction. In the present study, sows were treated daily with or without 1.5 mg of HTX per kilogram of feed from day 35 of pregnancy (at 30% of total gestational period), and individuals were sampled at three different ages: 100-day-old fetuses and 1-month- and 6-month-old piglets. After euthanasia, the brain was removed and the hippocampus, amygdala, and prefrontal cortex were dissected. The profile of the catecholaminergic and serotoninergic neurotransmitters (NTs) was characterized and an immunohistochemical study of the hippocampus was performed. The results indicated that maternal supplementation with HTX during pregnancy affected the NT profile in a brain-area-dependant mode and it modified the process of neuron differentiation in the hippocampal CA1 and GD areas, indicating that cell differentiation occurred more rapidly in the HTX group. These effects were specific to the fetal period, concomitantly with HTX maternal supplementation, since no major differences remained between the control and treated groups in 1-month- and 6-month-old pigs.

Descriptive study of a cohort of COVID-19 hospitalized patients in Spain.

BACKGROUND: Several descriptive cohort studies of patients affected by COVID-19 have been published. OBJECTIVE: To describe the characteristics of patients with SARS-CoV-2 infection who were admitted to Hospital Universitario la Plana, Castellón, Spain. METHODS: Retrospective, observational cohort study that included 18-year-old or older patients who were consecutively admitted with SARS-CoV2 confirmed infection. Demographic characteristics, comorbidities, clinical symptoms, laboratory results and radiological tests are described. RESULTS: The study included 255 patients with a mean age of 70 years; 54.9 % were males. Most common comorbidities were high blood pressure (58 %), dyslipidemia (42.4 %), diabetes (25.5 %) and obesity (24.3 %). Median number of days from the onset of clinical symptoms prior to hospital admission was seven. Most common manifestations prior to admission were fever (74.5 %), dry cough (61.2 %), malaise (51.8 %) and dyspnea (51.0 %); 19 patients (7.4 %) were admitted to the intensive care unit, where mortality was 50 %; overall mortality was 16.9 %. CONCLUSIONS: Our cohort reflects similar characteristics to those of other European series. Mortality was lower than that in similar studies.

ANTECEDENTES: Se han publicado varios estudios descriptivos de cohortes de pacientes afectados por COVID-19. OBJETIVO: Describir las características de pacientes con infección por SARS-CoV-2 que ingresaron al Hospital Universitario de La Plana, Castellón, España. MÉTODOS: Estudio observacional de cohortes retrospectivo, que incluyó pacientes de 18 años o mayores que ingresaron en forma consecutiva con infección confirmada por SARS-CoV2; se describen características demográficas, comorbilidades, síntomas clínicos, resultados de laboratorio y pruebas radiológicas. RESULTADOS: El estudio incluyó 255 pacientes con edad promedio de 70 años; 54.9 % fue del sexo masculino. Las comorbilidades más frecuentes fueron hipertensión arterial (58 %), dislipemia (42.4 %), diabetes (25.5 %) y obesidad (24.3 %). La mediana de días del inicio de síntomas clínicos antes del ingreso fue de siete. Las manifestaciones más frecuentes previas al ingreso fueron fiebre (74.5 %), tos seca (61.2 %), malestar general (51.8 %) y disnea (51.0 %); 19 pacientes (7.4 %) ingresaron a la unidad de cuidados intensivos, donde la mortalidad fue de 50 %; la mortalidad total fue de 16.9 %. CONCLUSIONES: Nuestra cohorte refleja características similares a las de otras series europeas. La mortalidad fue inferior a la de estudios similares.

Characteristics of Patients With Immune-Mediated Inflammatory Diseases Hospitalized for SARS-CoV-2 Infection. / Características de pacientes con enfermedades inflamatorias inmunomediadas hospitalizados por infección por SARS-CoV-2.

BACKGROUND: Immune-mediated inflammatory diseases (IMID) predispose to a higher infection risk by modifying the host's immune response, which acts as a key factor in SARS-CoV-2 infection resolution. Recent publications show that IMID patients and its treatments do not worsen the outcome of SARS-CoV-2 infection. OBJECTIVES: To describe the clinical characteristics and outcomes of patients with IMID who required hospital admission due to SARS-CoV-2 infection. Secondly, to compare clinical characteristics and outcomes between patients who required hospital admission due to SARS-CoV-2 infection with IMID and those who were not affected. METHODS: We performed an observational retrospective cohort study, including admitted patients with suspected SARS-CoV-2 infection, treated according to medical criteria and local protocols based on the best available scientific evidence. Clinical data were collected from their electronical clinical history. Statistical analysis determined the differences in the characteristics and clinical outcome of the infection in IMID patients. RESULTS: Of a total number of 612 revised patients, 23 had an IMID and 9 of them were positive for the SARS-CoV-2 infection. We did not observe a correlation between these two disorders. There was a higher frequency of obesity and cardiovascular disease among IMID patients, but without statistical significance. The clinical outcomes were no different between hospitalized IMID and non IMID patients. CONCLUSION: IMID and its treatments do not determine the outcome of patients admitted with SARS-CoV-2 infection.

Estudio descriptivo de una cohorte de pacientes con COVID-19 hospitalizados en España / Descriptive study of a cohort of COVID-19 hospitalized patients in Spain

Resumen Antecedentes: Se han publicado varios estudios descriptivos de cohortes de pacientes afectados por COVID-19. Objetivo: Describir las características de pacientes con infección por SARS-CoV-2 que ingresaron al Hospital Universitario de La Plana, Castellón, España. Métodos: Estudio observacional de cohortes retrospectivo, que incluyó pacientes de 18 años o mayores que ingresaron en forma consecutiva con infección confirmada por SARS-CoV2; se describen características demográficas, comorbilidades, síntomas clínicos, resultados de laboratorio y pruebas radiológicas. Resultados: El estudio incluyó 255 pacientes con edad promedio de 70 años; 54.9 % fue del sexo masculino. Las comorbilidades más frecuentes fueron hipertensión arterial (58 %), dislipemia (42.4 %), diabetes (25.5 %) y obesidad (24.3 %). La mediana de días del inicio de síntomas clínicos antes del ingreso fue de siete. Las manifestaciones más frecuentes previas al ingreso fueron fiebre (74.5 %), tos seca (61.2 %), malestar general (51.8 %) y disnea (51.0 %); 19 pacientes (7.4 %) ingresaron a la unidad de cuidados intensivos, donde la mortalidad fue de 50 %; la mortalidad total fue de 16.9 %. Conclusiones: Nuestra cohorte refleja características similares a las de otras series europeas. La mortalidad fue inferior a la de estudios similares.

Abstract Background: Several descriptive cohort studies of patients affected by COVID-19 have been published. Objective: To describe the characteristics of patients with SARS-CoV-2 infection who were admitted to Hospital Universitario la Plana, Castellón, Spain. Methods: Retrospective, observational cohort study that included 18-year-old or older patients who were consecutively admitted with SARS-CoV2 confirmed infection. Demographic characteristics, comorbidities, clinical symptoms, laboratory results and radiological tests are described. Results: The study included 255 patients, with a mean age of 70 years; 54.9 % were males. Most common comorbidities were high blood pressure (58 %), dyslipidemia (42.4 %), diabetes (25.5 %) and obesity (24.3 %). Median number of days from the onset of clinical symptoms prior to hospital admission was seven. Most common manifestations prior to admission were fever (74.5 %), dry cough (61.2 %), malaise (51.8 %) and dyspnea (51.0 %); 19 patients (7.4 %) were admitted to the intensive care unit, where mortality was 50 %; overall mortality was 16.9 %. Conclusions: Our cohort reflects similar characteristics to those of other European series. Mortality was lower than that in similar studies.

Neurobiology of environmental enrichment in pigs: hanges in monoaminergic neurotransmitters in several brain areas and in the hippocampal proteome.

Environmental enrichment in porcine farms improves animal welfare and leads to better public acceptance. To better understand the neurological mechanisms of the response to environmental enrichment, monoaminergic neurotransmitters were quantified in several brain areas from pigs after eight weeks of housing in barren or enriched conditions. Furthermore, iTRAQ labelling combined with LC-MS/MS was used to identify differentially abundant proteins in the hippocampus. Blood biochemical parameters related with stress and welfare were measured. Pigs under enriched conditions showed a decrease in plasma cortisol and lactate. The decrease in noradrenaline in the prefrontal cortex and amygdala, a general decrease in the dopaminergic system and an increase of serotonin in the striatum indicate a lower response to stress in enriched conditions. In the proteomic analysis, 2304 proteins were identified, of which 56 were differential between housing groups (46 upregulated and 10 downregulated). Bioinformatics analysis revealed that they were mainly related to ribosome, translation, microtubules and metabolic mitochondrial processes, indicating that pigs under enriched environments have higher abundance of proteins related to protein synthesis and neuronal activity. Together with previous behavioural studies, our results suggest that environmental enrichment provides a less stressful environment and that pigs cope better with stress conditions like the slaughterhouse. SIGNIFICANCE: Animal welfare has become an important aspect for the sustainability of animal production. The modification of the environment by enriching it with rooting materials and wider space allowance is known to have a positive effect on pigs' welfare. Searching for the underlying neurobiological mechanisms, we found that housing in an enriched environment increased the abundance of proteins related to protein synthesis, microtubule assembly, vesicle-mediated transport and energy metabolism in the hippocampus of pigs. Likewise, changes in the neurotransmitter profile in several brain areas were compatible with a better response to stress. This study expands the knowledge about the biological basis of animal welfare-promoting actions.

Metabolome and proteome changes in skeletal muscle and blood of pre-weaning calves fed leucine and threonine supplemented diets.

In pre-weaning calves, both leucine and threonine play important roles in growth and muscle metabolism. In this study, metabolomics, proteomics and clinical chemistry were used to assess the effects of leucine and threonine supplementation added to milk replacer on 14 newborn Holstein male calves: 7 were fed a control diet (Ctrl) and 7 were fed the Ctrl diet supplemented with 0.3% leucine and 0.3% threonine (LT) from 5.6 days of age to 53.6 days. At this time, blood and semitendinosus muscle biopsies were collected for analysis. Integrated metabolomics and proteomics showed that branched-chain amino acids (BCAA) degradation and mitochondrial oxidative metabolism (citrate cycle and respiratory chain) were the main activated pathways in muscle because of the supplementation. BCAA derivatives and metabolites related to lipid mobilization showed the major changes. The deleterious effects of activated oxidative phosphorylation were balanced by the upregulation of antioxidant proteins. An increase in protein synthesis was indicated by elevated aminoacyl-tRNA biosynthesis and increased S6 ribosomal protein phosphorylation in skeletal muscle. In conclusion, LT group showed greater BCAA availability and mitochondrial oxidative activity; as the muscle cells undergo greater aerobic metabolism, antioxidant defenses were activated to compensate for possible cell damage. Data are available via ProteomeXchange (PXD016098). SIGNIFICANCE: Leucine and threonine are essential amino acids for the pre-weaning calf, being of high importance for growth. In this study, we found that leucine and threonine supplementation of milk replacer to feed pre-weaning calves led to differences in the proteome, metabolome and clinical chemistry analytes in skeletal muscle and plasma, albeit no differences in productive performance were recorded. This study extends our understanding on the metabolism in dairy calves and helps optimizing their nutritional status.

Total and specific activities of superoxide dismutase (SOD) in seminal plasma are related with the cryotolerance of jackass spermatozoa.

This study investigated whether the activities of four antioxidant enzymes present in jackass seminal plasma (SP), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) and glutathione reductase (GSR), are related to the sperm ability to withstand cryopreservation. Eighteen ejaculates from 16 healthy jackasses were collected and split into two aliquots. The first one was centrifuged (3,000×g, 4 °C for 10 min) and used to determine the activities of these four enzymes in SP, whereas the other was diluted in a skim-milk extender and then cryopreserved. Assessment of sperm motility and membrane integrity was performed before and after cryopreservation. Based on the percentages of total motile and viable spermatozoa at post-thaw, samples were classified as good (GFE) or poor (PFE) freezability ejaculates through cluster analyses. Total and specific activities of SOD in seminal plasma were higher (P < 0.05) in GFE than in PFE, whereas no significant differences between GFE and PFE were observed regarding total and specific activities of CAT, GPX and GSR. However, post-thaw sperm parameters were positively correlated with total and specific activities of CAT and negatively correlated with those of GSR. In conclusion, determination of total and specific activities of SOD in the seminal plasma of a given jackass ejaculate may predict the sperm ability to withstand cryopreservation. In addition, our results warrant further research on addressing whether SOD activity in seminal plasma does not only allow predicting the sperm cryotolerance of a given ejaculate but also that of all ejaculates from a given jackass.

Specific Activity of Superoxide Dismutase in Stallion Seminal Plasma Is Related to Sperm Cryotolerance.

While the removal of seminal plasma is a routine practice prior to equine sperm cryopreservation, this fluid contains the main source of antioxidant enzymes able to scavenge these reactive oxygen species. Therefore, stallion seminal plasma components may have an impact on ejaculate freezability. Against this background, this study was designed to investigate whether the activities of the main stallion seminal plasma antioxidant enzymes are related to sperm cryotolerance. With this purpose, 16 ejaculates were collected from 14 healthy stallions, and each ejaculate was split into two aliquots. The first one was used to evaluate the activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), and glutathione reductase (GSR) in seminal plasma. The second aliquot was extended and then processed for cryopreservation. Sperm motility and viability were evaluated before and after cryopreservation, and ejaculates were classified as of good (GFE) or poor freezability (PFE) based on total motile and viable spermatozoa at post-thaw. We observed that, while the specific activities of CAT, GPX, and GSR were similar between GFE and PFE, that of SOD was significantly (p < 0.05) higher in GFE than in PFE. We can thus conclude that, in stallions, the specific activity of SOD in the seminal plasma of a given ejaculate might be related to its freezability.

Activities of antioxidant seminal plasma enzymes (SOD, CAT, GPX and GSR) are higher in jackasses than in stallions and are correlated with sperm motility in jackasses.

This study compared the activities of four antioxidant enzymes (superoxide dismutase, SOD; catalase, CAT; glutathione peroxidase, GPX; and glutathione reductase, GSR) in the seminal plasma of stallions and jackasses. Eighteen stallion ejaculates and 24 jack ejaculates were collected through an artificial vagina. Seminal plasma was obtained by several centrifugations at 3000×g and 4 °C for 10 min, and activities of SOD, CAT, GPX and GSR were subsequently determined. We also evaluated whether the collecting season had any influence on the activities of these four enzymes in both stallions and jackasses. Antioxidant capacity of seminal plasma was significantly higher in jackasses than in stallions (mean ±â€¯SEM, SOD: 1707.7 ±â€¯195.9 U/mL vs. 231.9 ±â€¯29.6 U/mL; CAT: 9094.7 ±â€¯1292.9 U/L vs.1682.7 ±â€¯525.9 U/L; GPX 845.4 ±â€¯106.0 U/L vs. 469.7 ±â€¯60.3 U/L; GSR: 50.3 ±â€¯5.1 U/L vs. 20.7 ±â€¯4.6 U/L). Furthermore, whereas season had no effect on the activity of these four enzymes in stallions, the activities of CAT and GPX in jack seminal plasma were significantly higher in the summer than in the other seasons. In addition, the activities of SOD and CAT were found to be significantly correlated with the percentages of progressively motile spermatozoa, and with the percentages of linearity and straightness, respectively, in jackasses. In contrast, the activities of these four enzymes were not correlated with sperm quality parameters in stallions. Finally, while SOD, CAT, and GPX activities but not those of GSR were correlated in jackasses, the activities of all four enzymes were correlated each other in stallions. We can thus conclude that the activities of SOD, CAT, GPX and GSR differ between the seminal plasma of stallions and donkeys, and vary between seasons in jackasses.

Age-Related Serum Biochemical Reference Intervals Established for Unweaned Calves and Piglets in the Post-weaning Period.

The purpose of the present study is to establish the influence of age on serum biochemistry reference intervals (RIs) for unweaned calves and recently-weaned piglets using large number of animals sampled at different ages from populations under different season trials. Specifically, milk replacer (MR)-fed calves from April-July 2017 (n = 60); from December 2016-March 2017 (n = 76) and from April-August 2018 (n = 57) and one group of healthy weaned piglets (n = 72) were subjected to the study. Serum enzymes and metabolites of calves at age of 24 h (24 h after colostrum intake), 2, 5, and 7 weeks from merged trials and piglets at 0, 7, and 14 days post-weaning (at 21, 28, and 35 days of age) were studied. The main variable is age whereas no major trial- or sex-biased differences were noticed. In calves, ALT, AST, GGT, GPx, SOD, NEFAs, triglycerides, glucose, creatinine, total protein, and urea were greatly elevated (p < 0.001) at 24 h compared with other ages; glucose, creatinine, total protein, and urea constantly decreased through the age; cholesterol's lowest level (p < 0.001) was found in 24 h compared with other ages and the levels of haptoglobin remained unchanged (p > 0.1) during the study. In comparison with the adult RIs, creatinine from 24 h, NEFAs from 2 w, GGT from 5 w, and urea from 7 w are fully comparable with RIs or lie within RIs determined for adult. In piglets, no changes were noticed on glucose (p > 0.1) and haptoglobin (p > 0.1) and there were no major changes on hepatic enzymes (ALT, AST, and GGT), total protein, creatinine and urea even though several statistical differences were noticed on 7 days post-weaning. Cholesterol, triglycerides, NEFAs, cortisol and PigMAP were found increased (p < 0.05) while TNF-alpha was found less concentrated (p < 0.001) at 0 days post-weaning compared with other times. Moreover, the RIs of creatinine and GGT are fully comparable with RIs or lie within RIs determined for adult. In conclusion, clinical biochemistry analytes RIs were established for unweaned calves and recently-weaned piglets and among them some can vary at different ages.

SWATH-MS quantitative proteomic investigation of intrauterine growth restriction in a porcine model reveals sex differences in hippocampus development.

Intrauterine growth restriction (IUGR) is characterized by reduced growth and weight of the foetus, mainly due to the lack of nutrients and oxygen. Animals affected by IUGR show changes in specific brain areas and several neuronal processes. Female offspring affected by IUGR show increased survival and development compared to males. The objective of this study was to analyse changes in the hippocampus proteome in male and female piglets affected by IUGR. Seven pregnant Iberian sows were fed from Day 35 of pregnancy onwards at 50% of their requirements. At Day 100 of pregnancy, foetuses were obtained and classified by sex and weight, as mild IUGR (Normal Body Weight) versus severe IUGR (Low Body Weight). Hippocampi were dissected and the proteomes analysed by SWATH-MS DIA. In this study, 1497 proteins were identified of which 260 were quantitatively analysed. All differential proteins were more abundant in females versus males and were involved in protein synthesis, neuronal development, metabolism, antiapoptotic signalling and vesicular transport. Our findings support that female foetuses tolerate nutrient limitation better than males, especially under mild IUGR. Under severe IUGR, females still seems to maintain normal lipid metabolism and antiapoptotic signalling, which may be related to the increased female survival. SIGNIFICANCE: In the last years, proteomics have been used to evidence differences related to sex in non-reproductive organs. Intrauterine Growth Restriction (IUGR) can affect female and male offspring differently. Female offspring has stronger protective strategies compared to males, enhancing growth and postnatal survival. Most studies regarding this issue have focused on metabolic organs (i.e. liver). However, the predominance of neurodevelopmental disorders in males suggests that the central nervous system in female offspring adapt better to nutritional stress conditions than that of males. Based on the differential protein expression in hippocampal samples, our work demonstrates that female foetuses indeed adapt better to IUGR than males, especially under mild IUGR conditions. In severe IUGR conditions, differences between males and females were not so evident, but even in this case, the remaining differences suggest increased survival in females than in males.