Inducció d'una proteïna d'estrès per fred en cèl·lules del cúmulus : un nou pas per a la millora de la vitrificació d'oòcits?

La vitrificació preserva les funcions de cèl·lules, com els oòcits, a temperatures properes a -196ºC, però la tolerància a aquestes temperatures extremes es encara reduïda. Per enfortir-los i que puguin resistir, el grup ERPAW/GRI-BCN de la UAB ha estudiat si el tractament previ amb fred (estrès cel·lular) pot servir com a preparació. A més, ha observat que les baixes temperatures indueixen una proteïna d'estrès per fred (CIRBP) en la capa de cèl·lules del cúmulus que envolten l'oòcit, les quals podrien ajudar a incrementar la tolerància dels oòcits al procés de vitrificació.La vitrificación preserva las funciones de células, como los oocitos, a temperaturas cercanas a -196ºC, pero la tolerancia a estas temperaturas extremas es todavía reducida. Para fortalecerlos y que puedan resistir, el grupo ERPAW/GRI-BCN de la UAB ha estudiado si el tratamiento previo con frío (estrés celular) puede servir como preparación. Además, ha observado que las bajas temperaturas inducen una proteína de estrés por frío (CIRBP) en la capa de células del cúmulos que rodean el ovocito, las cuales podrían ayudar a incrementar la tolerancia de los ovocitos al proceso de vitrificación.Vitrification preserves the functions of cells at -196ºC, but the tolerance of these extreme temperatures is still insufficient for the oocytes. To increase the vitrification tolerance, the group ERPAW/GRI-BCN of the UAB has studied if the pre-treatment with cold (cellular stress) could induce and increase the oocyte resistance to the process. Besides, the group has observed that the low temperatures induce a cold-inducible protein (CIRBP) in the layer of cumulus cells that surround the oocyte, which could help to increase the oocyte tolerance towards the vitrification process

Canvis en l'expressió gènica del receptor de glucocorticoides en el tracte reproductiu femení en conill : un nou paper en la fisiologia de la reproducció?

Els glucocorticoides modulen funcions essencials dels animals, entre elles la reproducció. Un nou estudi sobre els seus mecanismes de senyalització, en concret sobre els canvis en l'expressió gènica del receptor NR3C1/GR en conills davant de diferents estímuls, podria tenir aplicacions potencials en l'ús de les tècniques de reproducció assistida en aquesta espècie i també en altres mamífers.Los glucocorticoides modulan funciones esenciales de los animales, entre ellas la reproducción. Un nuevo estudio sobre sus mecanismos de señalización, en concreto sobre los cambios en la expresión génica del receptor NR3C1/GR en conejos ante diferentes estímulos, podría tener aplicaciones potenciales en el uso de las técnicas de reproducción asistida en esta especie y también en otros mamíferos.Glucocorticoids modulate essential animal functions, including reproduction. A new study on their signaling mechanisms, specifically on the changes in gene expression of the NR3C1/GR receptor in rabbits in response to different stimuli, could have potential applications in the use of assisted reproduction techniques in this species and also in other mammals

NR3C1 and glucocorticoid-regulatory genes mRNA and protein expression in the endometrium and ampulla during the bovine estrous cycle

Altres ajuts: acord transformatiu CRUE-CSICThe bovine reproductive tract exhibits changes during the estrous cycle modulated by the interplay of steroid hormones. Glucocorticoids can be detrimental when stress-induced but are relevant at baseline levels for appropriate reproductive function. Here, an analysis of quantitative real-time PCR was performed to study the bovine glucocorticoid-related baseline gene transcription in endometrial and ampullar tissue samples derived from three time points of the estrous cycle, stage I (Days 1-4), stage III (Days 11-17) and stage IV (Days 18-20). Our results revealed expression differences during stages, as expression observed in the ampulla was higher during the post-ovulatory phase (stage I), including the glucocorticoid receptor NR3C1, and some of its regulators, involved in glucocorticoid availability (HSD11B1 and HSD11B2) and transcriptional actions (FKBP4 and FKBP5). In contrast, in the endometrium, higher expression of the steroid receptors was observed during the late luteal phase (stage III), including ESR1, ESR2, PGRMC1 and PGRMC2, and HSD11B1 expression decreased, while HSD11B2 increased. Moreover, at protein level, FKBP4 was higher expressed during the late luteal phase, and NR3C1 during the pre-ovulatory phase (stage IV). These results suggest that tight regulation of the glucocorticoid activity is promoted in the ampulla, when reproductive events are taking place, including oocyte maturation. Moreover, most expression changes in the endometrium were observed during the late luteal phase, and may be related to the embryonic maternal recognition. In conclusion, the glucocorticoid regulation changes across the estrous cycle and may be playing a role on the reproductive events occurring in the bovine ampulla and endometrium

Semen Modulates the Expression of NGF, ABHD2, VCAN, and CTEN in the Reproductive Tract of Female Rabbits

Semen changes the gene expression in endometrial and oviductal tissues modulating important processes for reproduction. We tested the hypothesis that mating and/or sperm-free seminal plasma deposition in the reproductive tract affect the expression of genes associated with sperm-lining epithelium interactions, ovulation, and pre-implantation effects (nerve growth factor, NGF; α/β hydrolase domain-containing protein 2, ABHD2; C-terminal tensin-like protein, CTEN or TNS4; and versican, VCAN) in the period 10-72 h post-mating. In Experiment 1, does (n = 9) were treated with gonadotropin-releasing hormone (GnRH) (control), GnRH-stimulated, and vaginally infused with sperm-free seminal plasma (SP-AI), or GnRH-stimulated and naturally mated (NM). In Experiment 2, does (n = 15) were GnRH-stimulated and naturally mated. Samples were retrieved from the internal reproductive tracts (cervix-to-infundibulum) 20 h post-treatment (Experiment 1) or sequentially collected at 10, 24, 36, 68, or 72 h post-mating (Experiment 2, 3 does/period). All samples were processed for gene expression analysis by quantitative PCR. Data showed an upregulation of endometrial CTEN and NGF by NM, but not by SP-AI. The findings suggest that the NGF gene affects the reproductive tract of the doe during ovulation and beyond, influencing the maternal environment during early embryonic development

Semen Modulates the Expression of NGF, ABHD2, VCAN, and CTEN in the Reproductive Tract of Female Rabbits

Semen changes the gene expression in endometrial and oviductal tissues modulating important processes for reproduction. We tested the hypothesis that mating and/or sperm-free seminal plasma deposition in the reproductive tract affect the expression of genes associated with sperm-lining epithelium interactions, ovulation, and pre-implantation effects (nerve growth factor, NGF; α/β hydrolase domain-containing protein 2, ABHD2; C-terminal tensin-like protein, CTEN or TNS4; and versican, VCAN) in the period 10-72 h post-mating. In Experiment 1, does (n = 9) were treated with gonadotropin-releasing hormone (GnRH) (control), GnRH-stimulated, and vaginally infused with sperm-free seminal plasma (SP-AI), or GnRH-stimulated and naturally mated (NM). In Experiment 2, does (n = 15) were GnRH-stimulated and naturally mated. Samples were retrieved from the internal reproductive tracts (cervix-to-infundibulum) 20 h post-treatment (Experiment 1) or sequentially collected at 10, 24, 36, 68, or 72 h post-mating (Experiment 2, 3 does/period). All samples were processed for gene expression analysis by quantitative PCR. Data showed an upregulation of endometrial CTEN and NGF by NM, but not by SP-AI. The findings suggest that the NGF gene affects the reproductive tract of the doe during ovulation and beyond, influencing the maternal environment during early embryonic development

Semen Modulates the Expression of NGF, ABHD2, VCAN, and CTEN in the Reproductive Tract of Female Rabbits

Semen changes the gene expression in endometrial and oviductal tissues modulating important processes for reproduction. We tested the hypothesis that mating and/or sperm-free seminal plasma deposition in the reproductive tract affect the expression of genes associated with sperm-lining epithelium interactions, ovulation, and pre-implantation effects (nerve growth factor, NGF; α/β hydrolase domain-containing protein 2, ABHD2; C-terminal tensin-like protein, CTEN or TNS4; and versican, VCAN) in the period 10-72 h post-mating. In Experiment 1, does (n = 9) were treated with gonadotropin-releasing hormone (GnRH) (control), GnRH-stimulated, and vaginally infused with sperm-free seminal plasma (SP-AI), or GnRH-stimulated and naturally mated (NM). In Experiment 2, does (n = 15) were GnRH-stimulated and naturally mated. Samples were retrieved from the internal reproductive tracts (cervix-to-infundibulum) 20 h post-treatment (Experiment 1) or sequentially collected at 10, 24, 36, 68, or 72 h post-mating (Experiment 2, 3 does/period). All samples were processed for gene expression analysis by quantitative PCR. Data showed an upregulation of endometrial CTEN and NGF by NM, but not by SP-AI. The findings suggest that the NGF gene affects the reproductive tract of the doe during ovulation and beyond, influencing the maternal environment during early embryonic development

A protein-coding gene expression atlas from the brain of pregnant and non-pregnant goats

Background: The brain is an extraordinarily complex organ with multiple anatomical structures involved in highly specialized functions related with behavior and physiological homeostasis. Our goal was to build an atlas of protein-coding gene expression in the goat brain by sequencing the transcriptomes of 12 brain regions in seven female Murciano-Granadina goats, from which three of them were 1-month pregnant. Results: Between 14,889 (cerebellar hemisphere) and 15,592 (pineal gland) protein-coding genes were expressed in goat brain regions, and most of them displayed ubiquitous or broad patterns of expression across tissues. Principal component analysis and hierarchical clustering based on the patterns of mRNA expression revealed that samples from certain brain regions tend to group according to their position in the anterior-posterior axis of the neural tube, i.e., hindbrain (pons and medulla oblongata), midbrain (rostral colliculus) and forebrain (frontal neocortex, olfactory bulb, hypothalamus, and hippocampus). Exceptions to this observation were cerebellum and glandular tissues (pineal gland and hypophysis), which showed highly divergent mRNA expression profiles. Differential expression analysis between pregnant and non-pregnant goats revealed moderate changes of mRNA expression in the frontal neocortex, hippocampus, adenohypophysis and pons, and very dramatic changes in the olfactory bulb. Many genes showing differential expression in this organ are related to olfactory function and behavior in humans. Conclusion: With the exception of cerebellum and glandular tissues, there is a relationship between the cellular origin of sampled regions along the anterior-posterior axis of the neural tube and their mRNA expression patterns in the goat adult brain. Gestation induces substantial changes in the mRNA expression of the olfactory bulb, a finding consistent with the key role of this anatomical structure on the development of maternal behavior

Anthrax bio-surveillance of livestock in Arua District, Uganda, 2017-2018

Altres ajuts: acords transformatius de la UABThe authors would like to express their gratitude to the local Sub-County chiefs, district veterinary officers, community elders, and kraal leaders for being supportive during data collection. Author JG has received mobility support from Universitat Autònoma de Barcelona (action Erasmus+ KA107 Mobility Fellowship) and was supported by the Generalitat de Catalunya, Agency for Management of University and Research Grants co-financed with the European Social Found (grants for the recruitment of new research staff 2018 FI_B 00236). Authors RAO, ME, MA, MFN, MI, EI, MM, LP, BS, and SAA were funded by Livestock Disease Control project II.Anthrax, caused by Bacillus anthracis, is a widespread zoonotic disease with many human cases, especially in developing countries. Even with its global distribution, anthrax is a neglected disease with scarce information about its actual impact on the community level. Due to the ecological dynamics of anthrax transmission at the wildlife-livestock interface, the Sub-Saharan Africa region becomes a high-risk zone for maintaining and acquiring the disease. In this regard, some subregions of Uganda are endemic to anthrax with regular seasonal trends. However, there is scarce data about anthrax outbreaks in Uganda. Here, we confirmed the presence of B. anthracis in several livestock samples after a suspected anthrax outbreak among livestock and humans in Arua District. Additionally, we explored the potential risk factors of anthrax through a survey within the community kraals. We provide evidence that the most affected livestock species during the Arua outbreak were cattle (86%) compared to the rest of the livestock species present in the area. Moreover, the farmers' education level and the presence of people's anthrax cases were the most critical factors determining the disease's knowledge and awareness. Consequently, the lack of understanding of the ecology of anthrax may contribute to the spread of the infection between livestock and humans, and it is critical to reducing the presence and persistence of the B. anthracis spores in the environment. Finally, we discuss the increasingly recognized necessity to strengthen global capacity using a One Health approach to prevent, detect, control, and respond to public threats in Uganda

Oòcits, hipotèrmia i vitrificació : quin és el paper de les proteïnes induïdes per fred?

Els oòcits poden vitrificar-se per a la conservació de recursos genètics animals i per la preservació de la fertilitat en les dones, però el procés presenta algunes dificultats. Investigadors del Departament de Sanitat i d'Anatomia Animals han investigat el paper de les proteïnes induïdes per fred, que augmenten la seva expressió davant la hipotèrmia, per millorar la tolerància dels oòcits a la vitrificació.Los oocitos pueden vitrificarse para la conservación de recursos genéticos animales y para la preservación de la fertilidad en las mujeres, pero el proceso presenta algunas dificultades. Investigadores del Departamento de Sanidad y Anatomía Animal han investigado el papel de las proteínas inducidas por frío, que aumentan su expresión frente a la hipotermia, para mejorar la tolerancia de los ovocitos a la vitrificación.Oocytes can be vitrified for the conservation of animal genetic resources and for the preservation of fertility in women, but the process presents some difficulties. Researchers from the Department of Health and Animal Anatomy have investigated the role of cold-inducible proteins, which increase their expression in hypothermia conditions, to improve the tolerance of oocytes to vitrification

Cold-inducible RNA-binding protein : an RNA chaperone with potential roles in animal reproductive physiology

La vitrificació ha reemplaçat la congelació convencional per a la criopreservació d'ovòcits en diverses espècies. No obstant això, encara no s'ha establert el protocol òptim per a la criopreservació d'ovòcits bovins, encara que s'han desenvolupat diverses estratègies per a millorar els seus resultats. En aquest sentit, s'ha prestat poca atenció a l'estrès subletal per fred com a inductor de criotolerància en ovòcits bovins. La cold-inducible RNA-binding protein (CIRBP) pertany a les proteïnes induïbles per fred (CIPs), un grup de pèptids induïts per hipotèrmia i altres factors d'estrès. CIRBP és un candidat potencial per a millorar la criotolerància dels ovòcits bovins a causa de la seva funció en la supervivència cel·lular. A més, les CIPs exerceixen funcions crucials en el control del ARNm, la qual cosa planteja interrogants sobre la relació entre aquestes proteïnes i els canvis fisiològics que es produeixen en l'aparell reproductor femení. En conseqüència, la present tesi es va dur a terme amb l'objectiu general d'estudiar CIRBP com a molècula potencial per a millorar la vitrificació d'ovòcits bovins i la seva modulació durant els canvis fisiològics en el tracte reproductiu femení. En primer lloc, es va avaluar els efectes de la hipotèrmia en la maduració dels complexos cumulus-ovòcit. A més, es va determinar l'expressió proteica de CIRBP mitjançant Western blot. Es va demostrar que l'estrès per fred augmenta l'expressió de CIRBP. No obstant això, com s'esperava, es van detectar efectes perjudicials induïts pel fred. Després d'aquesta primera aproximació, es van buscar noves estratègies per a augmentar l'expressió de CIRBP evitant els efectes perjudicials del fred. Per a això, es van testar diferents concentracions de CIRBP exògena i una petita molècula hipotèrmico-mimètica ja establerta (zr17-2) com a suplement dels mitjans de maduració in vitro. Es van trobar pocs canvis en el ARNm de CIRBP i altres CIPs relacionats amb aquesta suplementació, però si es van observar increments d'expressió del ARNm de CIRBP i altres CIPs a causa de la hipotèrmia i vitrificació. Finalment, els estudis en el tracte reproductiu femení van revelar que CIRBP està modulada pels canvis hormonals fisiològics i la interacció mascle-femella. Aquesta tesi ofereix una nova visió de la fisiologia reproductiva de CIRBP, tant per a l'increment de coneixement bàsic com per al seu ús potencial en la millora de la vitrificació d'ovòcits bovins. Una millor comprensió del procés reproductiu pot servir com a nous biomarcadors, potencials eines d'avaluació reproductiva i millora de la producció animal.La vitrificación ha reemplazado la congelación convencional para la criopreservación de ovocitos en varias especies. Sin embargo, aún no se ha establecido el protocolo óptimo para la criopreservación de ovocitos bovinos, aunque se han desarrollado varias estrategias para mejorar sus resultados. En este sentido, se ha prestado poca atención al estrés subletal por frío como inductor de criotolerancia en ovocitos bovinos. La cold-inducible RNA-binding protein (CIRBP) pertenece a las proteínas inducibles por frío (CIPs), un grupo de péptidos inducidos por hipotermia y otros factores de estrés. CIRBP es un candidato potencial para mejorar la criotolerancia de los ovocitos bovinos debido a su función en la supervivencia celular. Además, las CIPs desempeñan funciones cruciales en el control del ARNm, lo que plantea interrogantes sobre la relación entre estas proteínas y los cambios fisiológicos que se producen en el aparato reproductor femenino. En consecuencia, la presente tesis se llevó a cabo con el objetivo general de estudiar CIRBP como molécula potencial para mejorar la vitrificación de ovocitos bovinos y su modulación durante los cambios fisiológicos en el tracto reproductivo femenino. En primer lugar, evaluamos los efectos de la hipotermia en la maduración de los complejos cumulus-ovocito. Además, determinamos la expresión proteica de CIRBP mediante Western blot. Se demostró que el estrés por frío aumenta la expresión de CIRBP. Sin embargo, como se esperaba, se detectaron efectos perjudiciales inducidos por el frío. Tras esta primera aproximación, se buscaron nuevas estrategias para aumentar la expresión de CIRBP evitando los efectos perjudiciales del frío. Para ello, testeamos diferentes concentraciones de CIRBP exógena y una pequeña molécula hipotérmico-mimética ya establecida (zr17-2) como suplemento de los medios de maduración in vitro. Se encontraron pocos cambios en el ARNm de CIRBP y otras CIPs relacionados con esta suplementación, pero si se observaron incrementos de expresión del ARNm de CIRBP y otras CIPs debido a la hipotermia y vitrificación. Finalmente, los estudios en el tracto reproductivo femenino revelaron que CIRBP está modulada por los cambios hormonales fisiológicos y la interacción macho-hembra. Esta tesis ofrece una nueva visión de la fisiología reproductiva de CIRBP, tanto para el incremento de conocimiento básico como para su uso potencial en la mejora de la vitrificación de ovocitos bovinos. Una mejor comprensión del proceso reproductivo puede servir como nuevos biomarcadores, potenciales herramientas de evaluación reproductiva y mejora de la producción animal.Vitrification has replaced the slow freezing method for cryopreservation of oocytes in several species. However, the optimal protocol for bovine oocyte cryopreservation remains to be established. Several strategies for improving bovine oocyte vitrification have been developed. In this regard, little attention has been given to sublethal mild hypothermia as an inductor of cryotolerance in bovine oocytes. The cold-inducible RNA-binding protein (CIRBP) belongs to the cold-inducible proteins (CIPs), a group of peptides induced by mild hypothermia and other stressors. CIRBP is a potential candidate to improve bovine oocyte cryotolerance due to its function in cell survival. In addition, CIPs play crucial roles in RNA transcript control, raising questions about the link between these proteins and the physiological changes in the female reproductive tract. Accordingly, the present thesis was conducted with the general objective to study CIRBP as a potential molecule to improve bovine oocyte vitrification and its modulation during physiological changes in the female reproductive tract. We first evaluated the effects of cold temperatures on the maturation of bovine cumulus-oocyte complexes. Additionally, we determined CIRBP protein expression by Western blot. Sublethal mild hypothermia was demonstrated to increase the expression of CIRBP. However, as expected, we reported detrimental effects induced by the sublethal stress. After this first approach, we aimed to find other strategies to increase the expression of CIRBP, avoiding the damaging effects of sub-physiological temperature variations. For this purpose, we tested different concentrations of exogenous CIRBP and a well-defined hypothermia mimetic small molecule (zr17-2), to supplement the in vitro maturation media of bovine cumulus-oocyte complexes. Minor changes in mRNA CIRBP expression and other CIPs were linked to this supplementation. Additionally, our results revealed that mild hypothermia and vitrification increased the mRNA expression of CIRBP and other CIPs. Finally, the female reproductive tract studies revealed that CIRBP is physiologically modulated by the cyclic hormonal changes and by the male-female interaction. Overall, this thesis offers new insight into CIRBP reproductive physiology, increasing the knowledge between CIRBP and the female reproductive tract and its potential use in improving bovine oocyte vitrification. A better understanding of the reproductive complex process may serve as novel clinical biomarkers, potential diagnostic tools for reproductive evaluation, and the improvement of animal production.Universitat Autònoma de Barcelona. Programa de Doctorat en Medicina i Sanitat Animal