Effect of Acute and Cumulative Stress on Gene Expression in Mammary Tissue and Their Interactions with Physiological Responses and Milk Yield in Saanen Goats.

This study addresses the hypothesis that different acute stressors can cumulatively decrease milk yield. In fact, in a time of global warming, the impact of environmental stress and farm management practices on milk production remains unclear. In this context, our objective was to investigate the effect of acute and cumulative stress on gene expression in mammary tissue and their interactions with physiological responses and milk yield in Saanen goats. Thirty lactating goats were subjected to two treatments: (1) control (CT), in which goats were maintained following a habitual routine under comfort conditions; (2) stress (ST), in which the goats were subjected to different types of environmental stress: heat stress, adrenocorticotropic hormone administration, hoof care management, and exposure to rain. These stressors were performed sequentially, with one stress per day on four consecutive lactation days, to evaluate their effect on milk quality and milk yield. Our results showed that compared to CT goats, cumulative stress increased the gene expression of glucocorticoid receptor (GR), interferon-gamma (IFN-γ), superoxide dismutase (SOD), and catalase (CAT) in mammary tissue, which are indicators of cortisol action, inflammatory response, and antioxidant enzymes. Furthermore, the acute challenges imposed on ST goats changed their rectal temperature and respiratory frequency and increased cortisol, glucose, cholesterol, triglycerides, and high-density lipoprotein release in plasma when compared to CT goats. Although these physiological and metabolic responses restore homeostasis, ST goats showed lower milk yield and higher somatic cell count in milk than CT goats. In conclusion, the results confirmed our initial hypothesis that different acute stressors cumulatively decrease the milk yield in Saanen goats.

Effect of intravenous lipid infusion on biomarkers of insulin resistance and immune functions of dry and nonpregnant dairy cows.

During the transition period, dairy cows often experience negative energy balance, which can induce metabolic and immunological disturbances. Previous work has shown that there is a relationship between the dysfunction of immune cells and the increase in blood nonesterified fatty acid (NEFA) concentration. Nevertheless, it is difficult to determine the exact effect of NEFA on the immune system, as other metabolic and hormonal perturbations occur simultaneously during the transition period. In the present study, we have determined the effect of NEFA on immune functions using an experimental model designed to assess the effects independently of energy balance, as well as hormonal and metabolic changes due to parturition. Six dry and nonpregnant cows were infused with either sterile water (control treatment) or a lipid emulsion (Intralipid 20%, Frenesius Kabi, lipid treatment) at a rate of 1 mL/kg per hour for 6 h according to a crossover design. Blood concentrations of NEFA, ß-hydroxybutyrate (BHB), and glucose were measured every hour throughout the infusion period, and 1 and 18 h after the end of infusion. Proliferation and interferon-γ secretion of lymphocytes, phagocytosis, and oxidative burst of neutrophils and blood insulin concentration were evaluated before, during, and at the end of the infusion. For NEFA, BHB, and glucose, treatment × time interactions were present. When compared with the control condition, NEFA and BHB levels were greater in the plasma of cows infused with lipids from 1 h after the start of infusion until 1 h after the end of infusion. Glucose level also increased in response to lipid infusion from 2 h of infusion until 1 h after the end of treatment. For sterile water and lipid infusions, respectively, maximal concentrations were 0.06 ± 0.10 mM and 1.39 ± 0.10 mM for NEFA, 0.70 ± 0.05 mM and 1.06 ± 0.05 mM for BHB, and 4.56 ± 0.27 mM and 6.90 ± 0.27 mM for glucose. For all blood metabolites, there were no differences between treatments 18 h postinfusion. Lipid infusion significantly increased blood insulin concentration at 3 and 6 h of infusion. However, it returned to its basal concentration 18 h after the end of the infusion. Lymphoproliferation declined as early as 3 h after the start of the lipid infusion. At 3 and 6 h of infusion, lipid treatment significantly reduced INF-γ concentration in the culture cell supernatant. The lipid infusion did not affect neutrophil phagocytosis. Nevertheless, the efficacy of the response was affected by a reduction of neutrophils' oxidative burst. These results confirm that NEFA inhibits immune functions independently of energy balance and other changes that occur during the transition period. They also indicate that high blood lipid concentration causes insulin resistance.

Comparison of hematological parameters between single and twin pregnancies in Dorper ewes during gestation, lambing, and postpartum / Comparação dos parâmetros hematológicos entre ovelhas Dorper de gestação simples e gemelar durante a prenhez, parto e pós-parto

ABSTRACT: The hematopoietic system changes during the pregnancy to carry fetal development and maternal needs. This study compared the hematological parameters between ewes with single and twin pregnancies during gestation, delivery, and postpartum. The experiment was conducted on 60 healthy pregnant Dorper ewes that were divided into two experimental groups: Group 1 (G1), with single pregnancies (n=30), and Group 2 (G2), with twin pregnancies (n=30). Blood samples were collected from all ewes at different times: immediately before fixed-time artificial insemination (AI); on day 30, 90, 120, 130, and 140 of pregnancy; immediately after delivery; and at 24h and 48h postpartum. Statistical analysis compared the two groups at different times (P<0.05). Mild, normocytic, and hypochromic anemia was detected in all ewes from AI time and throughout pregnancy from both groups, but did not prove to be of clinical relevance. In the peripartum stage (from the 140th day of pregnancy to 48h postpartum), the ewes with twin pregnancies (G2) exhibited higher erythrogram values and neutrophil:lymphocyte ratio than did ewes with single pregnancies (G1). This indicated a greater hematopoietic adaptation in the body during the development of two fetuses. Except for the eosinophil numbers, all leukogram parameters were influenced by pregnancy in a similar way in both groups, and was characterized mainly by leukocytosis with neutrophilia during peripartum due to the high presence of endogenous cortisol at delivery. Thus, these findings showed that pregnancy was a stressful physiological event that increased the leukocyte count with a slight alteration in the erythrogram of Dorper ewes.

RESUMO: O sistema hematopoiético sofre mudanças durante a gestação para atender o desenvolvimento fetal e as necessidades maternas. Este estudo comparou os parâmetros hematológicos entre ovelhas com gestação simples e gemelar durante a prenhez, parto e pós-parto. O estudo foi realizado em 60 ovelhas Dorper prenhes, saudáveis, divididas em dois grupos experimentais: Grupo 1 (G1) com gestação simples (n = 30); e Grupo 2 (G2) com gestação gemelar (n = 30). Amostras de sangue foram colhidos de todas as ovelhas em diferentes momentos: imediatamente antes da inseminação artificial em tempo fixo (IA); nos dias 30, 90, 120, 130 e 140 de gestação; imediatamente após o parto; e às 24h e 48h pós-parto. A análise estatística comparou os dois grupos nos diferentes momentos (P < 0,05). Anemia leve, normocítica e hipocrômica foi detectada desde a IA e ao longo da gestação, nas ovelhas de ambos os grupos, mas não houve relevância clínica. Na fase periparto (do 140º dia de gestação às 48h pós-parto), as ovelhas com gestação gemelar (G2) mostraram valores do eritrograma e proporção neutrófilos: linfócitos mais elevados do que as ovelhas com gestação simples (G1). Isso indicou maior adaptação hematopoiética da ovelha para o desenvolvimento de dois fetos. Com exceção do número de eosinófilos, todos os parâmetros do leucograma foram influenciados pela gestação de forma semelhante em ambos os grupos, e foi caracterizada principalmente por leucocitose com neutrofilia no periparto devido ao alto nível de cortisol endógeno no parto. Assim, os achados mostraram que a gestação foi um evento fisiológico estressante que aumentou a contagem de leucócitos com leve alteração no eritrograma de ovelhas Dorper.

Long-term heat stress at final gestation: physiological and heat shock responses of Saanen goats.

The long exposure to heat negatively changes performance and productivity of animals, particularly when heat stress is associated with gestation. Indeed, little is known about the negative effects of long-term heat stress on the final gestation of dairy goats. In this context, the physiological and cellular responses of Saanen goats submitted to heat stress (37°C from 10:00 to 16:00 h) were investigated from day 60th pre-partum to day 60th post-partum. At final gestation, 46 pregnant Saanen goats were randomly assigned to the treatments: control (CT; thermal neutral conditions) and heat stress (HS; climatic chamber). After partum, all experimental goats were maintained in thermal neutral conditions. The rectal, dorsal, mammary temperatures and respiratory frequency, cortisol release, milk yield, milk quality, and the genes HSP60, HSP70, HSP90, Glucocorticoid receptor and ACTHR. Goats subjected to HS showed significantly (P < 0.05) higher rectal, dorsal, and mammary temperatures and significantly mobilized the increase of respiratory frequency to lose heat as compared to CT goats. The HS challenge significantly increased cortisol release from day 15th pre-partum to day 15th post-partum. CT goats produced more milk than HS from weeks 4 to 10 of lactation (P <0.001), with no difference in milk quality. However, on day 15th post-partum, there was a significant effect of HS treatment on the expression of HSP70 and ACTHR genes as compared to CT treatment, confirming the long-term effect of HS on Saanen goats. In conclusion, the physiological parameters studied increased pre-partum in the hottest hour, and cortisol peaked on day 15 pre-partum for heat-stressed goats. Although on the 15th day post-partum, all goats were in thermal comfort, and the physiological parameters were within the normal range, the concentration of cortisol continued to be significantly higher for goats submitted to thermal stress. Indeed, milk yield was greater for goats subjected to pre-partum thermal comfort. Furthermore, the expression of HSP70 and ACTHR genes on peripheral blood mononuclear cells are interesting biomarkers for studying the long-term effect of heat stress on Saanen goats.

Maternal stress in sheep during late pregnancy influences sperm quality in early puberty of the offspring.

Stress at the end of sheep gestation can damage the reproductive development of young males. The aim of the present study was to evaluate the effects of LPS administration in the last third of sheep pregnancy on the reproductive parameters of prepubertal rams. Thirty-six pregnant nulliparous ewes (12 ±â€¯2 months old; 45 ±â€¯6 kg) were assigned to two treatments, LPS (E. coli; 0.8 µg kg-1) and control (placebo/saline) administered in late pregnancy (120 days post-conception). The animals gave birth to 17 male lambs (11 LPS; 8 control). Reproductive development of the young rams was analyzed from 5 to 12 months of age. A completely randomized design in double factorial scheme was used. The data were analyzed by analysis of variance. The model included treatment (LPS; control), age as main effects and their interactions, and the animal as a repeated measure. Means were compared by the PDIFF-SAS (Pr > |t|) at P < 0.05. An effect of age was observed for scrotal circumference, testicular consistency, homogeneity of testicular parenchyma, vascularization, semen quantity and quality, and blood testosterone concentration (P < 0.05). LPS increased sperm defects (P < 0.05) but an interaction with age was not observed (P > 0.05) with higher abnormalities only during months 8 and 9 (P < 0.05) and not thereafter. In summary, LPS did not cause long-term damage to testicular morphology analyzed from the onset of puberty to sexual maturity. However, LPS treatment affected sperm morphology during early puberty of the offspring.

Heat loss efficiency and HSPs gene expression of Nellore cows in tropical climate conditions.

Adaptation is a relevant characteristic to be understood in livestock animals in order to maintain and raise productivity. In Brazil, the Nellore beef cattle are widely disseminated and well-adapted breed that present good thermoregulatory characteristics for tropical environment conditions. Conversely, the physiological and cellular mechanisms required for thermoregulation and thermotolerance in this breed are still limited. The aim of this study was to comprehend the heat loss efficiency at the whole animal level and heat shock response at the cellular level of Nellore cows in tropical climate conditions. Healthy purebred Nellore cows were classified according to their capacity to lose body heat as Efficient or Inefficient based on vaginal temperature which was continuously monitored by data-loggers. Rectal, tail, and ocular temperatures, sweating rate, and respiratory frequency were collected to assess other thermoregulatory responses. Peripheral mononuclear cells were used for gene expression of heat shock proteins 60, 70, and 90 induced by in vitro heat treatments at 38, 40, and 42 °C. In our findings, the Efficient cows presented higher sweating rates compared to Inefficient cows that presented higher rectal temperature with greater amplitude of vaginal temperature profile. Transcription of the HSP genes was stable at 38 and 40 °C and decreased for all HSP genes at 42 °C. In conclusion, the Nellore efficiency to lose heat was mainly associated with their sweating capacity and cellular thermotolerance confirmed by the maintenance of heat shock proteins transcripts under heat stress. Taken together, this knowledge contributes as a future key for genetic selection of adapted animals.

Acute heat stress induces changes in physiological and cellular responses in Saanen goats.

The relationships between rectal temperatures and physiological and cellular responses to heat stress can improve the productivity of Saanen goats in tropical environments. In this context, this study evaluated the physiological responses and gene expression of heat shock proteins (HSP60, 70, and 90) and genes related to apoptosis (Bax, Bcl-2, and p53) of Saanen goats subjected to acute heat stress. Ten health Saanen goats were exposed to solar radiation during 3 consecutive days. The expression of HSP60, HSP70, HSP90, Bax, Bcl-2, and p53 genes in blood leukocytes, rectal and superficial temperatures, respiratory frequency, cortisol, triiodothyronine, and thyroxine was measured at 06:00, 13:00, and 18:00 h. In vitro, blood leukocytes were subjected to 38 °C and 40 °C for 3 h to measure the expression of the same target genes. The temperature humidity index, measured from 12:00 to 15:00, was greater than 80 and black globe temperatures were greater at 40 °C, indicating the intensity of the solar radiation. Although the solar radiation caused acute heat stress, increased cortisol release, and the expression of HSP60 and 70 in dry Saanen goats, the increased respiratory frequency and decreased T4 and T3 restored the homeothermy of the experimental goats. In vitro, the 40 °C increased the expression of p53 (pro-apoptotic protein), Bcl-2 (anti-apoptotic protein), HSP60, HSP70, and HSP90, suggesting that these genes have protective functions. However, further studies are necessary to understand the physiological and cellular responses to heat stress.

Termografia infravermelha da superfície ocular como indicador de estresse em suínos na fase de creche / Infrared thermography of the ocular surface as stress indicator for piglets postweaning

Desmama é uma fase crítica na vida do suíno devido a separação materna e a introdução de uma dieta seca. A termografia infravermelha medida na região ocular se mostra como um indicador confiável para a condição de estresse pontual de suínos. O objetivo deste estudo foi determinar a relação entre medidas de termografia infravermelha da superfície ocular e de cortisol em situações de estresse em leitões na pós desmama. Foram avaliados 66 leitões, uma vez por semana, durante sete semanas, em dois períodos do dia (7h e 15h) com medidas de temperatura superficial ocular, temperatura superficial do dorso e amostras de saliva para determinação de cortisol salivar. A análise estatística contemplou os efeitos fixos de semana e período do dia e sua interação e correlações de Pearson para relação entre termografia ocular, temperatura superficial e cortisol a 5% de significância. Cortisol salivar não diferiu entre os períodos, mas foi superior nas três primeiras semanas após o desmame (P<0,05). Nas duas primeiras semanas após a desmama o cortisol apresentou correlação alta e positiva (P<0,05) com a temperatura ocular máxima (0,89) e a temperatura superficial do dorso (0,80). As duas temperaturas superficiais apresentaram uma associação moderada positiva (r=0,41; P<0,0001) durante todo o período experimental. Este estudo destaca que a temperatura de superfície ocular obtida por meio da termografia infravermelha pode ser um indicador de temperatura de superfície corporal e estado de bem-estar de leitões em fase de creche, além de ser um método não invasivo e de rápida mensuração. Entretanto, mais pesquisas são necessárias para aprofundar a relação entre temperatura ocular e cortisol durante estresse crônico.(AU)

Weaning is a critical phase in pigs' life due to maternal separation and the introduction of a new diet. Infrared thermography measurement taken in the ocular region appears to be a reliable indicator of the stress condition of the pig. The aim of this study was to evaluate the relationship between ocular surface temperature by infrared thermography and cortisol in piglets post weaning. Sixty-six piglets were evaluated once a week, during 7 weeks, in two periods of the day (7am and 15pm) and ocular surface temperature and dorsal surface temperature were collected using a thermographic camera and a laser surface thermometer, respectively. Saliva was also collected to determine salivary cortisol. Statistical analysis included fixed effects of week and period of the day and their interaction, and relationship between thermography, dorsal surface temperature and cortisol were done by Pearson's correlations with 5% significance level. Salivary cortisol did not differ between periods, but it was higher in the first three weeks after weaning (P<0.05). During the first two weeks after weaning cortisol presented high and positive correlation (P<0.05) between ocular surface temperature (0.89) and dorsal surface temperature (0.80). The two superficial temperatures had a moderate and positive association (r=0.41; P<0.0001) during all experiment. This study highlights that the ocular surface temperature obtained through infrared thermography can be a superficial body temperature indicators, besides being a non-invasive and fast method of measurement. However, more research is needed to deepen the relationship between ocular surface temperature and cortisol during chronic stress.(AU)