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Quantitative reverse transcription PCR (RT-qPCR) remains as an accurate approach for gene expression analysis but requires labor-intensive validation of reference genes using species-specific primers. To ease such demand, the aim was to design and test a multi-species primer set to validate reference genes for inter-genus RT-qPCR gene expression analysis. Primers were designed for ten housekeeping genes using transcript sequences of various livestock species. All ten gene transcripts were detected by RT-PCR in Bos taurus (cattle), Bubalus bubalis (buffaloes), Capra hircus (goats), and Ovis aries (sheep) cDNA. Primer efficiency was attained for eight reference genes using B. taurus-O. aries fibroblast cDNA (95.54-98.39%). The RT-qPCR data normalization was carried out for B. taurus vs. O. aries relative gene expression using Bestkeeper, GeNorm, Norm-finder, Delta CT method, and RefFinder algorithms. Validation of inter-genus RT-qPCR showed up-regulation of TLR4 and ZFX gene transcripts in B. taurus fibroblasts, irrespectively of normalization conditions (two, three, or four reference genes). In silico search in mammalian transcriptomes showed that the multi-species primer set is expected to amplify transcripts of at least two distinct loci in 114 species, and 79 species would be covered by six or more primers. Hence, a multi-species primer set allows for inter-genus gene expression analysis between O. aries and B. taurus fibroblasts and further reveals species-specific gene transcript abundance of key transcription factors.

The extent to which mammalian cells share similar transcriptomes remains unclear. Notwithstanding, such cross-species gene expression inquiries have been scarce for defined cell types and most lack the dissection of gene regulatory landscapes. Therefore, the work was aimed to determine C-MYC relative expression across mammalian fibroblasts ( Ovis aries and Bos taurus ) via cross-species RT-qPCR and comprehensively explore its regulatory landscape by in silico tools. The prediction of transcription factor binding sites in C-MYC and its 2.5 kb upstream sequence revealed substantial variation, thus indicating evolutionary-driven re-wiring of cis-regulatory elements. C-MYC and its downstream target TBX3 were up-regulated in Bos taurus fibroblasts. The relative expression of C-MYC regulators [ RONIN (also known as THAP11 ), RXRβ , and TCF3 ] and the C-MYC -associated transcript elongation factor CDK9 did not differ between species. Additional in silico analyses suggested Bos taurus -specific C-MYC exonization, alternative splicing, and binding sites for non-coding RNAs. C-MYC protein orthologs were highly conserved, while variation was in the transactivation domain and the leucine zipper motif. Altogether, mammalian fibroblasts display evolutionary-driven C-MYC relative expression that should be instructive for understanding cellular physiology, cellular reprogramming, and C-MYC -related diseases.

The addition of growth factors and vitamins enhances goat embryonic development in vitro. However, few attempts have been reported trying to identify supplementation regimens for oocyte maturation or embryo culture with additive properties. The present report was aimed to evaluate if retinoids [0.3 μM retinyl acetate (RAc) and 0.5 μM 9-cis-retinoic acid (RA)] supplementation during goat oocyte maturation and retinoids and/or 50 ng mL–1 IGF-I during embryo culture synergically enhanced embryonic development while diminishing the incidence of apoptosis. All combinations of RAc and RA treatment produced blastocysts with similar efficiencies, while IGF-I enhanced embryos yields irrespectively of retinoid addition. Moreover, retinoids and IGF-I supplementation showed similar caspase activity or DNA fragmentation indexes in blastocysts. In conclusion, supplementation with retinoids and IGF-I during goat embryo culture enhances blastocysts development without synergic reduction of apoptosis.

Background: Cryopreservation preserves cellular viability under low temperatures, resulting in diminished intracellular enzymatic activity and reduced cellular metabolism that ultimately allows preserving genetic material for indefinite periods of time. Embryos submitted to cryopreservation suffer from considerable morphological and functional damage, resulting in reduced survival and development rates. Objective: To evaluate pregnancy and delivery rates of in vitro-produced (IVP) Nellore (Bos indicus) embryos after vitrification under field conditions. Methods: The IVP embryos at blastocyst (Bl) and expanded blastocyst (Bx) were transferred fresh (n= 137) or after vitrification (n= 127). Results: Pregnancy rates at 35 d for fresh embryos were lower in Bl (41.6) than in Bx (60.9) (p0.05). Pregnancy loss at 60 d were similar (p>0.05) for both fresh (Bl: 3.1 and Bx: 4.8) and vitrified embryos (Bl: 1.9 and Bx: 4.7). Delivery rates were similar between groups (p>0.05). Conclusion: Both pregnancy and delivery rates of Bos indicus IVP embryos vitrified under field conditions are indistinguishable from fresh embryos.Key words: cattle reproduction, cryobiology, cryoinjury, cryotolerance, embryo surplus, embryo transfer. Resumen Antecedentes: La criopreservación se caracteriza por el mantenimiento de la viabilidad celular a bajas temperaturas, resultando en reducido metabolismo y actividad enzimática intracelular, lo que permite la preservación del material genético por períodos de tiempo indefinidos. Los embriones sometidos a ésta técnica sufren daños morfológicos y funcionales considerables, dando como resultado una sobrevivencia y tasas de desarrollo reducidas. Objetivo: Evaluar la tasa de preñez a partir de embriones Nelore (Bos indicus) producidos in vitro (IVP) después de la vitrificación bajo condiciones de campo. Métodos: Embriones IVP en los estadios de blastocisto (Bl) y blastocisto expandido (Bx) se transfirieron en fresco (n= 137) o después de la vitrificación (n= 127). Resultados: La tasa de preñez a los 35 d fue menor para los embriones transferidos en fresco en fase Bl (41,6) en relación con los Bx (60,9) (p0,05). Las pérdidas de preñez a los 60 d fueron similares (p>0,05) tanto para embriones en fresco en Bl (3,1) y Bx (4,8) como para los vitrificados (Bl: 1,9 y Bx: 4,7). Las tasas de nacimiento fueron similares entre los grupos (p>0,05). Conclusión: Las tasas de preñez y nacimiento de embriones IVP vitrificados de Nelore (Bos indicus) bajo condiciones de campo son semejantes a las de embriones en fresco.Palabras clave: criobiología, crioinjuria, criotolerancia, embriones sobrantes, reproducción de ganado, transferencia de embriones. Resumo Antecedentes: A criopreservação é caracterizada pela manutenção da viabilidade celular em baixas temperaturas, resultando em atividade enzimática intracelular e metabolismo celular reduzido, que permite a preservação do material genético por períodos indefinidos de tempo. Embriões submetidos à criopreservação sofrem danos morfológicos e funcionais consideráveis, resultando em sobrevivência reduzida e menores taxas de desenvolvimento. Objetivo: Avaliar a taxa de prenhez a partir de embriões Nelore (Bos indicus) produzidos in vitro (IVP) após a vitrificação sob condições de campo. Métodos: Embriões IVP nos estádios de blastocisto (Bl) e blastocisto expandido (Bx) foram transferidos a fresco (n= 137) ou depois da vitrificação (n= 127). Resultados: A taxa de prenhez aos 35 d foi menor para os embriões transferidos a fresco na fase de Bl (41,6), em relação aos Bx (60,9) (p0,05). As perdas de prenhez aos 60 d foram semelhantes (p>0,05) tanto para embriões a fresco nos estádios de Bl (3,1) e Bx (4,8), e vitrificados em Bl (1,9) e Bx (4,7). As taxas de nascimentos foram semelhantes entre os grupos (p>0,05). Conclusão: As taxas de prenhez e nascimentos dos embriões IVP vitrificados de Nelore (Bos indicus) sob condições de campo é semelhante àquela dos embriões a fresco.Palavras-chave: criobiologia, crioinjúria, criotolerância, embriões excedentes, reprodução de gado, transferência de embriões.

Abstract Background: Proper timing for embryo collection and transfer in horses -which is critical for the success of this biotechnology- is still debated. Additionally, there is little information on this technology under tropical conditions. Objective: To determine the best day for collection and transfer of embryos in Mangalarga Marchador mares under Brazilian northeast’s conditions. Methods: Donors (n= 30) and recipients (n= 76) in diestrus phase were selected based on both clinical and gynecology examinations. Estrus was induced on both donor and recipient mares by intramuscular injection of 5 mg Dinoprost, aiming to obtain an ovulation interval of -1 to +3 between recipient and donor. Ovulation was induced with buserelin acetate when the largest follicle reached at least 35 mm in diameter. At this time, mares were subjected to artificial insemination at 48-hour intervals until ovulation. The embryos were collected on days 7, 8, and 9 after ovulation. Results: The embryo collection on day 8 was more efficient (p<0.05) than on day 7, but it was not more effective (p>0.05) than day 9, which presented the same efficiency (p>0.05) as day 7. From a total of 76 embryos transferred to the recipients, that were between days 4 and 9 after ovulation, there was no influence (p>0.05) of the day of transfer on pregnancy rate. Conclusions: The embryo collection must be performed on day 8 after ovulation, and transfer can be performed on any day of that interval (4-9) without affecting the pregnancy rate. Resumo Antecedentes: A importância do momentoda colheita e da transferência do embrião equino para o sucesso dessa biotécnica em equino continua sem ser completamente entendida. Adicionalmente, existe pouca informação sobre essa tecnologia em condições tropicais. Objetivo: Determinar o melhor dia para colheita e para transferência de embriões em eguas manga larga marchador nas condições do nordeste brasileiro. Métodos: Doadoras (n = 30) e receptoras (n = 76) na fase de diestro foram selecionadas com base nos exames clínico e ginecológicos. O estro das éguas doadoras e receptoras foi induzido com 5 mg de Dinoprost administrado por via intramuscular, buscando obter um intervalo de ovulação de -1 a +3 entre a receptora e a doadora. A ovulação foi induzida com acetato de buserelina quando o foliculo maior alcançou o tamanho de 35 mm de diâmetro. Nesse momento, as éguas foram submetidas a inseminação artificial em intervalos de 48 horas até a ovulação. Os embriões foram colhidos nos dias 7, 8 e 9 depois da ovulação. Resultados: A colheita de embriões no dia 8 foi mais eficiente (p<0,05) do que no dia 7, porem não foi mais efetivo (p>0,05) do que o dia 9, o qual apresentou a mesma eficiência (p>0,05) que o dia 7. De um total de 76 embriões transferidos para as receptoras que se encontravam entre os dias 4 e 9 depois da ovulação, não se registrou influência (p>0,05) do dia da transferência sobre a taxa de prenhez. Conclusões: A colheita embrionária deve ser realizada no dia 8 depois da ovulação, e a transferência pode ser realizada em qualquer dia desse intervalo (4-9) sem que a taxa de prenhez seja afetada. Resumen Antecedentes: El momento mas apropiado para la recolección y transferencia de embriones en equinos -que es fundamental para el éxito de esta biotecnología- continua siendo sujeto de estudio. Además, es escasa la información sobre esta tecnología en condiciones tropicales. Objetivo: Determinar el momento mas adecuado para la recolecta y transferencia de embriones en yeguas Mangalarga Marchador, en las condiciones del nordeste Brasileño. Métodos: Donadoras (n= 30) y receptoras (n= 76) en la fase de diestro se seleccionaron con base en los exámenes clínicos y ginecológicos. El estro de las yeguas donadoras y receptoras fue inducido con 5 mg de Dinoprost, vía intramuscular, intentando obtener un intervalo de ovulación de -1 a +3 entre la receptora y la donadora. La ovulación fue inducida con acetato de buserelina cuando el folículo mayor alcanzó 35 mm de diámetro. En ese momento, las yeguas fueron sometidas a inseminación artificial en intervalos de 48 horas hasta la ovulación. Los embriones fueron recolectados en los días 7, 8 y 9 después de la ovulación. Resultados: La recolecta de embriones en el día 8 fue más eficiente (p<0,05) que en el día 7, pero no fue más efectivo (p>0,05) que en el día 9, el cuál presentó la misma eficiencia (p>0,05) que en el día 7. De un total de 76 embriones transferidos a las receptoras, que se encontraban entre el día 4 y 9 después de la ovulación, no se registró influencia (p>0,05) del día de la transferencia en la tasa de preñez. Conclusiones: La recolecta embrionaria debe ser realizada el día 8 después de la ovulación, y la transferencia puede ser realizada en cualquier día de este intervalo (4 a 9) sin que se afecte la tasa de preñez.

Background: Cryopreservation preserves cellular viability under low temperatures, resulting in diminished intracellular enzymatic activity and reduced cellular metabolism that ultimately allows preserving genetic material for indefinite periods of time. Embryos submitted to cryopreservation suffer from considerable morphological and functional damage, resulting in reduced survival and development rates. Objective: To evaluate pregnancy and delivery rates of in vitro-produced (IVP) Nellore (Bos indicus) embryos after vitrification under field conditions. Methods: The IVP embryos at blastocyst (Bl) and expanded blastocyst (Bx) were transferred fresh (n= 137) or after vitrification (n= 127). Results: Pregnancy rates at 35 d for fresh embryos were lower in Bl (41.6) than in Bx (60.9) (p0.05). Pregnancy loss at 60 d were similar (p>0.05) for both fresh (Bl: 3.1 and Bx: 4.8) and vitrified embryos (Bl: 1.9 and Bx: 4.7). Delivery rates were similar between groups (p>0.05). Conclusion: Both pregnancy and delivery rates of Bos indicus IVP embryos vitrified under field conditions are indistinguishable from fresh embryos.

Background: Proper timing for embryo collection and transfer in horses -which is critical for the success of this biotechnology- is still debated. Additionally, there is little information on this technology under tropical conditions. Objective: To determine the best day for collection and transfer of embryos in Mangalarga Marchador mares under Brazilian northeast’s conditions. Methods: Donors (n= 30) and recipients (n= 76) in diestrus phase were selected based on both clinical and gynecology examinations. Estrus was induced on both donor and recipient mares by intramuscular injection of 5 mg Dinoprost, aiming to obtain an ovulation interval of -1 to +3 between recipient and donor. Ovulation was induced with buserelin acetate when the largest follicle reached at least 35 mm in diameter. At this time, mares were subjected to artificial insemination at 48-hour intervals until ovulation. The embryos were collected on days 7, 8, and 9 after ovulation. Results: The embryo collection on day 8 was more efficient (p<0.05) than on day 7, but it was not more effective (p>0.05) than day 9, which presented the same efficiency (p>0.05) as day 7. From a total of 76 embryos transferred to the recipients, that were between days 4 and 9 after ovulation, there was no influence (p>0.05) of the day of transfer on pregnancy rate. Conclusions: The embryo collection must be performed on day 8 after ovulation, and transfer can be performed on any day of that interval (4-9) without affecting the pregnancy rate.

The current work reports the functional characterization of human induced pluripotent stem cells (iPSCs)- arterial and venous-like endothelial cells (ECs), derived in chemically defined conditions, either in monoculture or seeded in a scaffold with mechanical properties similar to blood vessels. iPSC-derived arterial- and venous-like endothelial cells were obtained in two steps: differentiation of iPSCs into endothelial precursor cells (CD31 pos /KDR pos /VE-Cad med /EphB2 neg /COUP-TF neg ) followed by their differentiation into arterial and venous-like ECs using a high and low vascular endothelial growth factor (VEGF) concentration. Cells were characterized at gene, protein and functional levels. Functionally, both arterial and venous-like iPSC-derived ECs responded to vasoactive agonists such as thrombin and prostaglandin E2 (PGE 2 ), similar to somatic ECs; however, arterial-like iPSC-derived ECs produced higher nitric oxide (NO) and elongation to shear stress than venous-like iPSC-derived ECs. Both cells adhered, proliferated and prevented platelet activation when seeded in poly(caprolactone) scaffolds. Interestingly, both iPSC-derived ECs cultured in monoculture or in a scaffold showed a different inflammatory profile than somatic ECs. Although both somatic and iPSC-derived ECs responded to tumor necrosis factor-α (TNF-α) by an increase in the expression of intercellular adhesion molecule 1 (ICAM-1), only somatic ECs showed an upregulation in the expression of E-selectin or vascular cell adhesion molecule 1 (VCAM-1).

We present a suitable approach to address the electric vehicle charging roaming problem (e-roaming). Blockchain technologies are applied to support the identity management process of users charging their vehicles and to record energy transactions securely. At the same time, off-chain cloud-based storage is used to record the transaction details. A user wallet settled on a mobile application stores user verified credentials; a backend application in the vehicle charging station validates the user credentials to authorize the energy transaction. The current model can be applied to similar contexts where the user may be required to keep several credentials from different providers to authenticate digital transactions.

This study investigated the use of distributed optical fiber sensing to measure temperature and strain during thermomechanical processes in printed circuit board (PCB) manufacturing. An optical fiber (OF) was bonded to a PCB for simultaneous measurement of temperature and strain. Optical frequency-domain reflectometry was used to interrogate the fiber optic sensor. As the optical fiber is sensitive to both temperature and strain, a demodulation technique is required to separate both effects. Several demodulation techniques were compared to find the best one, highlighting their main limitations. The importance of good estimations of the temperature sensitivity coefficient of the OF and the coefficient of thermal expansion of the PCB was highlighted for accurate results. Furthermore, the temperature sensitivity of the bonded OF should not be neglected for accurate estimations of strains. The two-sensor combination model provided the best results, with a 2.3% error of temperature values and expected strain values. Based on this decoupling model, a methodology for measuring strain and temperature variations in PCB thermomechanical processes using a single and simple OF was developed and tested, and then applied to a trial in an industrial environment using a dynamic oven with similar characteristics to those of a reflow oven. This approach allows the measurement of the temperature profile on the PCB during oven travel and its strain state (warpage).