Academic Research on cattle’s acclimatisation process.

ACADEMIC RESEARCH ON CATTLE’S ACCLIMATISATION PROCESS Flávio Silva1, Cristina Conceição1,2, Liliana Cachucho3, Catarina Matos1, Ana Lúcia Garrido1, Ana Geraldo1,2, Elsa Lamy2, Fernando Capela e Silva2,4, Paulo Infante5 & Alfredo Pereira1,2, 1Departamento de Zootecnia, Universidade de Évora, Portugal; 2ICAAM, Universidade de Évora, Portugal; 3CEBAL, Beja, Portugal; 4Departamento de Biologia, Universidade de Évora, Portugal; 5Deparatamento de Matemática, Universidade de Évora, Portugal. ABSTRACT Animal’s welfare and performance is compromised by environmental heat stress, present during summer in the Mediterranean climate. Heat stress effects are well known and widely studied, although the mechanisms of season acclimatisation are less well understood. Throughout the last 20 years, we have been studying this process in the Mediterranean and tropical regions, aiming to understand it better, developing and improving methodologies and search for reliable biomarkers of thermal stress. The climatic changes, the more frequent extreme events and the increase of ambient temperature at the surface of the earth were identified as a problem to the animals in general and particularly under production systems conditions. Then, the main question is how those factors can affect the farm animals, especially those with higher genetic merit for production and lack of environmental adaptation. With these premises, were developed research projects on cattle’s acclimatisation process. These projects outcomes were: 2 PhD thesis and 5 Master dissertations, alongside ten articles peer-review and index journals and 20 publications in the book of abstracts and also several communications in national and international symposiums. Keywords: acclimatisation; heat stress; cattle

Variações fisiológicas face à aclimatação sazonal – Estudo em vacas leiteiras com diferente potencial leiteiro

As alterações climáticas estão identificadas como uma das maiores ameaças ambientais, sociais e económicas para o planeta. O aumento de eventos climáticos extremos, como secas prolongadas, temperaturas extremas ou períodos com precipitação intensa e prolongada têm efeitos nos sistemas de produção animal (Avendaño-Reyes, 2012). Sabese que temperaturas elevadas afetam o bem-estar e o desempenho dos animais, pois provocam sérias alterações nas suas funções biológicas, como a diminuição na ingestão de matéria seca, na eficiência de utilização dos nutrientes e alterações metabólicas e hormonais (Marai et al., 2007). Nos bovinos leiteiros, estas mudanças culminam na redução da fertilidade, na diminuição da produção leiteira e da qualidade do leite, bem como na diminuição da resposta do sistema imunológico (Avendaño-Reyes, 2012). Segundo Kadzere et al. (2002), o progresso genético refletir-se-á em animais cada vez mais produtivos, uma vez que a seleção, em grande parte, tem vindo a ser efetuada pela quantidade de leite e não pela adaptabilidade às condições ambientais. Assim, as vacas de alta produção converteram-se em animais extremamente sensíveis a ambientes quentes (Cerqueira, 2013; Bernabucci et al., 2014). Quando expostos a um stresse térmico por calor prolongado, as vacas tendem a aclimatarem-se por forma a reajustarem a sua fisiologia para a menor aquisição de calor e à maior perda de calor (Horowitz, 2001), com efeitos negativos na produtividade do animal (Bernabucci et al., 2010). O objetivo deste estudo foi estudar o processo de aclimatação sazonal em vacas leiteira com distintos potenciais leiteiros. Pretendeu-se avaliar as respostas de vacas leiteiras de alto e baixo potencial leiteiro, aclimatadas ao Verão e ao Inverno, relativamente aos parâmetros composição do leite, temperatura retal, frequência respiratória, hemograma e triiodotironina

Effect of acclimatisation in physiological parameters of high-yielding dairy cows

Heat stress alter the physiological status and the energetic balance in high producing animals. Acclimatisation is a thermoregulatory adaptation to heat stress with detrimental effects on productivity. We hypothesised that high-yielding dairy cows (HP; ≥9,000 kg – 305 days in milk; n=7) suffer a more significant influence of elevated environmental temperatures than low-yielding cows (LP; <9,000 kg – 305 DIM; n=6). Physiological and milk composition data was collected in summer (5 days with mean environmental temperatures of 23.5 °C – heat stress) and winter (5 days with mean environmental temperature of 6.6 °C – thermoneutrality). Respiratory rate (HP: 63.95±12.35; LP: 64.34±13.67 movements/minute), sweat rate (HP: 77.70±48,90; LP: 75.86±45.02 g/m2/h) rectal temperature (HP: 38,87±0,72; LP: 38,76±0,63 °C) were significantly higher in summer than in winter across both groups, indicating a response to mild heat stress. Plasma triiodothyronine levels were lower in HP than LP in summer, indicating a higher degree of acclimatisation in HP cows. Haematocrit and hemoglobulin were significantly higher in summer but not different between groups. Regarding milk production, HP produced more milk than LP, but the difference between groups was shorter in summer than winter (17.90% and 22,30%, respectively). There were no differences in milk parameters within groups, except urea in the summer period (293.62 mg/kg and 253.69 mg/kg for HP and LP, respectively). Milk fat and protein were significantly lower in summer than winter. These results showed that elevated environmental temperatures alter the physiological status in both groups. Cows with different milk yield had similar first responses to heat stress. However, during the acclimatisation process, HP decreased metabolism rate while alterations in nitrogen pathways were observed

Characterization of the mechanism of action of two compounds with bone formation properties

Tese de mestrado, Ciências Biofarmacêuticas, Universidade de Lisboa, Faculdade de Farmácia, 2016Introduction: Osteoporosis is an asymptomatic disease characterized by decreased bone mass density and microarchitectural changes in the bone tissue. These changes lead to increased bone fragility and consequently a higher risk of fractures. Bone fragility occurs due to alterations in the bone remodeling process, when bone resorption exceeds formation. For the treatment of osteoporosis are used anti-catabolic drugs that inhibit bone degradation, anabolic agents that induce or facilitate bone formation, and dual effect compounds which at the same time promote reduction of bone resorption, as well as, maintenance or increasing of bone formation. However, current treatments have serious adverse effects and some of them are not very effective. Aim: In order to reduce fracture risk and improve patients’ quality of life, it is important to continue to search for the ideal therapeutic treatment for osteoporosis. TechnoPhage found two chemical compounds that increase bone formation, however, it is not known their mechanism of action. In order to understand the effect of these compounds we evaluated their toxic, proliferative and anabolic effect in osteoblastic cell lines and their toxic and anabolic effect in vivo in zebrafish larvae. Conclusions: In Saos-2 cell line, both compounds did not affect cell proliferation but one compound had an anabolic effect on cell mineralization. In zebrafish larvae, one compound was toxic in concentrations higher than 10 μM and did not have an anabolic effect; other compound was not toxic and did not have an anabolic effect. Drawbacks of the work: During this work we had problems with stability of compounds what lead to the loss of their initial anabolic effect on cells mineralization. Therefore, it was necessary to do new batches of compounds to understand their real effect. Using the zebrafish larvae, we realize that one compound had altered toxicity in different batches (old and new) but for the other compound we did not observe significant differences between two batches.Introdução: Osteoporose é uma doença assintomática caracterizada pela diminuição da densidade da massa óssea e por alterações na microarquitectura do tecido ósseo. Estas alterações levam a um aumento da fragilidade do osso e consequentemente um maior risco de fraturas. A fragilidade óssea ocorre devido a alterações no processo de remodelação óssea, quando a reabsorção óssea é superior à formação. Para o tratamento da osteoporose são utilizados fármacos anti-catabólicos que inibem a degradação do osso, compostos anabólicos que induzem ou facilitam a formação óssea e compostos com dupla ação que ao mesmo tempo promovem a redução da reabsorção óssea, assim como a manutenção ou o aumento da formação de osso. No entanto, os tratamentos atuais têm graves efeitos adversos e a maioria deles são pouco eficazes. Objetivo: De forma a diminuir o risco de fraturas e melhorar a qualidade de vida dos doentes é necessária mais investigação para estabelecer o tratamento ideal para a osteoporose. A TechnoPhage descobriu dois compostos químicos que aumentam a quantidade de osso formada, no entanto, desconhece-se o seu mecanismo de ação. A fim de compreender o efeito destes compostos, avaliámos o seu efeito tóxico, proliferativo e anabólico em linhas celulares de osteoblastos, e o seu efeito tóxico e anabólico em larvas de peixe-zebra. Conclusões: Na linha celular Saos-2, estes compostos não afetaram a proliferação celular mas um destes compostos tem um efeito anabólico na mineralização celular. Em larvas de peixe-zebra, um composto foi tóxico em todas as concentrações superiores a 10 μM e não teve um efeito anabólico; o outro composto não foi tóxico e não teve um efeito anabólico. Imprevistos: Durante este trabalho tivemos alguns problemas com a estabilidade dos compostos o que levou à perda do seu efeito anabólico inicial na mineralização celular. Portanto, foi necessário fazer novas soluções dos compostos para compreender o seu verdadeiro efeito. Usando larvas de peixe-zebra como modelo, percebemos que um composto tem uma toxicidade distinta nas diferentes soluções (antiga e nova) mas para o outro composto não observámos nenhuma diferença significativa entre as duas soluções.Trabalho realizado na TechnoPhage SA

Caracterização histofisiológica das glândulas sudoríparas e das características da pelagem em vacas leiteiras face à aclimatação sazonal.

SEASONAL ACCLIMATISATION PROCESS IN DAIRY COWS: STUDYING HISTOFISIOLOGICAL CHARACTERISTICS OF SWEAT GLANDS AND HAIR COAT ABSTRACT: This trial aimed to study the seasonal acclimatisation process in dairy cows in a Mediterranean climate, focusing the changes in the histophysiology of sweat glands and hair coat. From 10 cows we collected data in two periods (P1 – summer; P2 – winter). Ambient temperature with a black globe thermometer (sun and shadow) was registered. From each cow, sweat rate (Ts), skin temperature (Tpl) and cutaneous tissue samples were taken to perform histomophormetical evaluation. Hair coat samples were collected for measurements and for medula expression. Conclusion: in summer, animals had higher Ts and Tpl, even though our results were not as higher as many other results in some other papers. In our trial, animals were always in shadow and ambient temperature decreased significantly at night time. Significantly differences in histomorphometric were found between periods. Hair coat was similar in summer and winter (measurements and medulation). So, the physiological mechanisms that regulate sweat rate and skin temperature changed with the acclimatisation process, confirmed by modifications in the morphometric of sweat glands and epidermis. Keywords: acclimatisation, cattle, hair, sweat gland

Salivary biomarkers of acclimatization in dairy cows with different milk yield potential

Environmental heat stress affects the productive performance of animals, especially those of high genetic merit. The Mediterranean region is characterized by sudden and prolonged heat periods. Although animals can adapt to warm environmental conditions, in these conditions they are less likely to acclimate. Saliva testing is a non-invasive and inexpensive test that can be a source of biomarkers. Results from our team suggested that the salivary levels of Hsp70 could function as a putative biomarker of thermal stress. The main objective of this work was to study the acclimatization process in dairy cows with high milk yield potential (HP; > 8000 kg of milk at 305 days in lactation) and low milk yield potential (LP; ≤ 8000 kg of milk) and to relate physiological parameters with salivary protein profiles. 6 HP and 6 LP animals were followed during four days in two periods: Summer - high environmental temperatures (animals under heat stress) and Winter – low environmental temperature (animals in thermoneutrality). Environment temperature was evaluated using a black globe thermometer (BGT) placed outside (BGTsun) and inside the facility (BGTshade). Rectal temperatures (RT) and respiratory rate (RR) were measured. On each period samples of saliva were collected through cotton rolls (Salivettes ®) and samples of blood from the coccygeal vein. BGT, RT and RR were significantly higher (P<0.05) in Summer (BGTsun 35.71±4.71°C; BGTshade 23.88±2.05°C; 38.80±0.10°C; 64.13±3.69 mov.min.) comparatively to Winter (BGTsun 14.21±2.61°C; BGTshade 6.15±1.01°C; 38.07±0.02; 36.13±2.21 mov.min.), but no differences were observed between HP and LP. Regarding T3, in Summer, HP had significatively lower values than LP, which indicated a more intense acclimatization. Higher values of salivary pH were observed in Summer in both HP and LP group. No significant differences were observed among periods in SC, however there was a significative negative correlation between SC and RT (r=-0.615, P=0.03). In Winter, HP had higher levels of SC than LP cows, which could be associated with metabolic heat production. Salivary albumin was higher in HP, during Summer. The results reinforce the concept that saliva can be useful in monitoring temperature adaptation in milk-producing animals

MECANISMOS HOMEOSTÁTICOS E HOMEORRÉTICOS DECORRENTES DA ACLIMATIZAÇÃO SAZONAL EM BOVINOS LEITEIROS DE DIFERENTE CAPACIDADE PRODUTIVA

O objetivo deste estudo consistiu em avaliar a resposta fisiológica de bovinos leiteiros, de raça Holstein-Frísia, com diferente potencial para a produção de leite, em duas fases de aclimatação sazonal. Utilizando a produção de leite corrigida para os 305 dias de lactação, foram selecionadas 6 vacas multíparas, com uma produção de leite inferior a 9000 kg (BP) e 7 vacas multíparas, com produção de leite superior a 9000 kg (AP). Os animais foram acompanhados em dois períodos durante 5 dias (P1: animais aclimatados às condições de Verão submetidos a temperaturas que indiciam stress térmico; P2: animais aclimatados às condições de inverno submetidos a temperaturas dentro da faixa de termoneutralidade). Mediu-se a temperatura de globo negro (TGN), à sombra e ao sol, a frequência respiratória (FR), a taxa de sudação (TS) e a temperatura retal (TR) em ambos os períodos. Diariamente registou-se as produções de leite de cada vaca e recolheu-se amostras de leite para a determinação da sua composição química e células somáticas. Em cada período foi recolhida uma amostra de sangue para a determinação de parâmetros bioquímicos e da triiodotironina (T3). Observou-se a influência do fator período na TGN, FR, TS, TR, na composição leiteira, nos parâmetros hematológicos e no metabolismo (T3). Foram observados valores superiores de ureia do leite e valores inferiores de T3 nas AP durante o P1. Estes resultados mostraram que vacas com diferente potencial leiteiro apresentaram uma resposta ao calor semelhante, no entanto, com a aclimatação as AP demonstraram modificações fisiológicas mais pronunciadas, com uma redução da taxa metabólica e com alterações no metabolismo do azoto

Overexpression of wild-type IL-7Rα promotes T-cell acute lymphoblastic leukemia/lymphoma

© 2021 by The American Society of HematologyTight regulation of IL-7Rα expression is essential for normal T-cell development. IL-7Rα gain-of-function mutations are known drivers of T-cell acute lymphoblastic leukemia (T-ALL). Although a subset of patients with T-ALL display high IL7R messenger RNA levels and cases with IL7R gains have been reported, the impact of IL-7Rα overexpression, rather than mutational activation, during leukemogenesis remains unclear. In this study, overexpressed IL-7Rα in tetracycline-inducible Il7r transgenic and Rosa26 IL7R knockin mice drove potential thymocyte self-renewal, and thymus hyperplasia related to increased proliferation of T-cell precursors, which subsequently infiltrated lymph nodes, spleen, and bone marrow, ultimately leading to fatal leukemia. The tumors mimicked key features of human T-ALL, including heterogeneity in immunophenotype and genetic subtype between cases, frequent hyperactivation of the PI3K/Akt pathway paralleled by downregulation of p27Kip1 and upregulation of Bcl-2, and gene expression signatures evidencing activation of JAK/STAT, PI3K/Akt/mTOR and Notch signaling. Notably, we also found that established tumors may no longer require high levels of IL-7R expression upon secondary transplantation and progressed in the absence of IL-7, but remain sensitive to inhibitors of IL-7R-mediated signaling ruxolitinib (Jak1), AZD1208 (Pim), dactolisib (PI3K/mTOR), palbociclib (Cdk4/6), and venetoclax (Bcl-2). The relevance of these findings for human disease are highlighted by the fact that samples from patients with T-ALL with high wild-type IL7R expression display a transcriptional signature resembling that of IL-7-stimulated pro-T cells and, critically, of IL7R-mutant cases of T-ALL. Overall, our study demonstrates that high expression of IL-7Rα can promote T-cell tumorigenesis, even in the absence of IL-7Rα mutational activation.The work was supported by the MRC under Programme Codes U117573801 and MR/P011225/1, to and by the consolidator grant ERC CoG-648455 from the European Research Council, under the European Union's Horizon 2020 research and innovation programme, and the FAPESP/20015/2014 grant from Fundação para a Ciência e a Tecnologia (FCT) (J.T.B.). A.S. received a fellowship from FCT (SFRH/BD/18388/2004). J.T.B. was an FCT Investigator (consolidator) and A.R.G. was the recipient of an FCT Investigator grant (IF/00510/2014). This work was also supported by a grant from “Fonds Kinderen Kankervrij” (KiKa 2010-082) (Y.L.).info:eu-repo/semantics/publishedVersio

Detection of 70 kDa heat shock protein in the saliva of dairy cows

This Research Communication describes, for the first time, the detection of HSP70 in saliva of dairy cows. Thermal stress is a major environmental stress that limits animal growth, metabolism, and productivity. The cellular response to heat stress involves the synthesis of heat shock proteins (HSPs), presumably to protect the functional stability of cells at increasing temperatures. HSP70 has been found to be present in cattle blood serum and may also be present in other secretory fluids, such as saliva, as already observed in humans. The aim of this study was to detect heat shock protein HSP70 in bovine saliva. Saliva samples were taken from higher- (n = 5) and lower milk producing (n = 5) Holstein-Friesian cows in summer and in winter for the detection of HSP70. HSP70 concentrations were assayed using the ELISA technique. Salivary HSP70 concentrations ranged from 0·524 to 12·174 ng/ml in cows. Higher salivary HSP70 concentrations were significantly associated with higher milk production and higher environmental temperature, but not with rectal temperature