Invited review: Muscle protein breakdown and its assessment in periparturient dairy cows.

Mobilization of body reserves including fat, protein, and glycogen is necessary to overcome phases of negative nutrient balance typical for high-yielding dairy cows during the periparturient period. Skeletal muscle, the largest internal organ in mammals, plays a crucial role in maintaining metabolic homeostasis. However, unlike in liver and adipose tissue, the metabolic and regulatory role of skeletal muscle in the adaptation of dairy cows to the physiological needs of pregnancy and lactation has not been studied extensively. The functional integrity and quality of skeletal muscle are maintained through a constant turnover of protein, resulting from both protein breakdown and protein synthesis. Thus, muscle protein breakdown (MPB) and synthesis are intimately connected and tightly controlled to ensure proper protein homeostasis. Understanding the regulation of MPB, the catabolic component of muscle turnover, and its assessment are therefore important considerations to provide information about the timing and extent of tissue mobilization in periparturient dairy cows. Based on animal models and human studies, it is now evident that MPB occurs via the integration of 3 main systems: autophagy-lysosomal, calpain Ca2+-dependent cysteine proteases, and the ubiquitin-proteasome system. These 3 main systems are interconnected and do not work separately, and the regulation is complex. The ubiquitin-proteasomal system is the most well-known cellular proteolytic system and plays a fundamental role in muscle physiology. Complete degradation of a protein often requires a combination of the systems, depending on the physiological situation. Determination of MPB in dairy cows is technically challenging, resulting in a relative dearth of information. The methods for assessing MPB can be divided into either direct or indirect measurements, both having their strengths and limitations. Available information on the direct measures of MPB primarily comes from stable isotopic tracer methods and those of indirect measurements from assessing expression and activity measures of the components of the 3 MPB systems in muscle biopsy samples. Other indirect approaches (i.e., potential indicators of MPB), including ultrasound imaging and measuring metabolites from muscle degradation (i.e., 3-methylhistidine and creatinine), seem to be applicable methods and can provide useful information about the extent and timing of MPB. This review presents our current understanding, including methodological considerations, of the process of MPB in periparturient dairy cows.

Intervenções psicológicas necessárias: a prática como residente no serviço de medicina fetal / Necessary psychological interventions: the practice as a resident in the fetal medicine service / Intervenciones psicológicas necesarias: la práctica como residente en el servicio de medicina fetal

Com os avanços tecnológicos e o aprimoramento da prática médica via ultrassonografia, já é possível detectar possíveis problemas no feto desde a gestação. O objetivo deste estudo foi analisar a prática do psicólogo no contexto de gestações que envolvem riscos fetais. Trata-se de um estudo qualitativo sob formato de relato de experiência como psicólogo residente no Serviço de Medicina Fetal da Maternidade Escola da Universidade Federal do Rio de Janeiro (UFRJ). Os registros, feitos por observação participante e diário de campo, foram analisados em dois eixos temáticos: 1) intervenções psicológicas no trabalho em equipe em consulta de pré-natal, exame de ultrassonografia e procedimento de amniocentese; e 2) intervenções psicológicas em casos de bebês incompatíveis com a vida. Os resultados indicaram que o psicólogo nesse serviço é essencial para atuar de forma multiprofissional na assistência pré-natal para gravidezes de alto risco fetal. Ademais, a preceptoria do residente é relevante para sua formação e treinamento para atuação profissional no campo da psicologia perinatal.(AU)

Face to the technological advances and the improvement of medical practice via ultrasound, it is already possible to detect possible problems in the fetus since pregnancy. The objective of this study was to analyze the psychologist's practice in the context of pregnancies which involve fetal risks. It is a qualitative study based on an experience report as a psychologist trainee at the Fetal Medicine Service of the Maternity School of UFRJ. The records, based on the participant observation and field diary, were analyzed in two thematic axes: 1) psychological interventions in the teamwork in the prenatal attendance, ultrasound examination and amniocentesis procedure; and 2) psychological interventions in cases of babies incompatible to the life. The results indicated that the psychologist in this service is essential to work in a multidisciplinary way at the prenatal care for high fetal risk pregnancies. Furthermore, the resident's preceptorship is relevant to their education and training for professional performance in the field of Perinatal Psychology.(AU)

Con los avances tecnológicos y la mejora de la práctica médica a través de la ecografía, ya se puede detectar posibles problemas en el feto desde el embarazo. El objetivo de este estudio fue analizar la práctica del psicólogo en el contexto de embarazos de riesgos fetal. Es un estudio cualitativo basado en un relato de experiencia como residente de psicología en el Servicio de Medicina Fetal de la Escuela de Maternidad de la Universidade Federal do Rio de Janeiro (UFRJ). Los registros, realizados en la observación participante y el diario de campo, se analizaron en dos ejes temáticos: 1) intervenciones psicológicas en el trabajo en equipo, en la consulta prenatal, ecografía y los procedimientos de amniocentesis; y 2) intervenciones psicológicas en casos de bebés incompatibles con la vida. Los resultados señalaron como fundamental la presencia del psicólogo en este servicio trabajando de forma multidisciplinar en la atención prenatal en el contexto de embarazos de alto riesgo fetal. Además, la tutela del residente es relevante para su educación y formación para el desempeño profesional en el campo de la Psicología Perinatal.(AU)

Assessment of Fetal Aortic, Thoracic, and Heartbeat in Healthy Donkey Pregnancies.

Practitioners are frequently requested to diagnose and stage pregnancy in donkeys with unknown breeding dates; however, scant work has been done to stage pregnancy in the species. Therefore, this study aimed to determine the association between measurements of fetal aortic, thoracic, and heartbeat with gestational age in donkeys carrying and delivering healthy foals. Multiparous Dezhou donkeys (n = 50) ranging from 4 to 16 years were enrolled in the study by 150 days of gestation. Transabdominal ultrasonography coupled with a 3.5 MHz sectorial convex transducer was performed at 30 day-intervals until delivery to obtain fetal aortic, thoracic, and fetal heartbeat measurements. Data were tested for normality with Shapiro- Wilk's test and then ANOVA and Tukey's. Statistical significance was set at P < .05. The mean duration of pregnancy was 356.6 ± 10.6 days (339-368). There were significant associations between gestational age and fetal aortic (r = 0.89) and thoracic (r = 0.88) measurements. Fetal heartbeat (r = -0.76) was negatively correlated with gestation length. The fetal aortic and thoracic measurements increased from the seventh month of gestation to the term (P < .001). The fetal heartbeat remained steady from 150 days to 270 days of gestation, then continuously decreased from 270 days until parturition (P < .001). In conclusion, fetal aortic and thoracic measurements are strongly associated with gestational age in donkeys. Fetal heartbeat decreases with advanced pregnancy.

Assessment of peripheral markers and ultrasonographic parameters in pregnant mares receiving intramuscular or intrauterine cloprostenol.

The present study aimed to compare two methods of prostaglandin-induced abortion in mares by determining blood markers (progesterone, estradiol-17ß, alpha-fetoprotein, 13,14-dihydro-15-keto-prostaglandin-F2α (PGFM)), B-mode ultrasonographic parameters, and time until loss of fetal heartbeat. It was hypothesized that intrauterine infusion of cloprostenol results in earlier fetal compromise than intramuscular administration. Ovarian structures (number and sizes of follicles and corpora lutea area), fetal heartbeat, and fetal mobility of thirteen singleton pregnancies were assessed daily by transrectal ultrasonography until induction of pregnancy termination (60 ±â€¯2 days of gestation). Mares received 500 µg of cloprostenol intramuscularly every 12 h (IM, n = 7) or once transcervically (TC, n = 6). After initial cloprostenol administration, ultrasonographic examinations were repeated at 6-h intervals until loss of fetal heartbeat was detected. Plasma progesterone, estradiol-17ß, and alpha-fetoprotein were assessed for five days before and after pregnancy loss. In addition, plasma PGFM concentrations were assessed immediately before cloprostenol administration (0 min), and then 15, 30, and 45 min, and 1, 2, 3, 4, 6, 12 h after administration. Data were analyzed using the MIXED procedure with repeated measures in SAS. Significance was set at P < 0.05. All mares lost their pregnancies within 48 h after initial cloprostenol administration, with no difference in time to pregnancy loss. There were significant effects of time starting by 12 h post-induction of pregnancy termination but there was no time by group interaction for progesterone concentrations. Estradiol-17ß and alpha-fetoprotein concentrations were not altered upon impending abortion. Concentrations of PGFM increased significantly by 2 h after cloprostenol administration, but there were no differences between groups. No time effects or time by group interaction for fetal mobility and heartbeat was detected. Expectedly, the number and area of corpora lutea decreased significantly after cloprostenol administration with no significant differences between groups. In conclusion, intrauterine administration of cloprostenol was not different from repeated systemic administration to terminate the pregnancy. Both models for early fetal loss were equivalent for the endpoints assessed herein. The present study provides evidence that transcervical cloprostenol administration technique is repeatable in different settings and results in negligible side effects. While systemic administration results in colic-like signs and may result in severe reaction.

Assessment of the nitrofen model of congenital diaphragmatic hernia and of the dysregulated factors involved in pulmonary hypoplasia.

PURPOSE: To study pulmonary hypoplasia (PH) associated with congenital diaphragmatic hernia (CDH), investigators have been employing a fetal rat model based on nitrofen administration to dams. Herein, we aimed to: (1) investigate the validity of the model, and (2) synthesize the main biological pathways implicated in the development of PH associated with CDH. METHODS: Using a defined strategy, we conducted a systematic review of the literature searching for studies reporting the incidence of CDH or factors involved in PH development. We also searched for PH factor interactions, relevance to lung development and to human PH. RESULTS: Of 335 full-text articles, 116 reported the incidence of CDH after nitrofen exposure or dysregulated factors in the lungs of nitrofen-exposed rat fetuses. CDH incidence: 54% (27-85%) fetuses developed a diaphragmatic defect, whereas the whole litter had PH in varying degrees. Downregulated signaling pathways included FGF/FGFR, BMP/BMPR, Sonic Hedgehog and retinoid acid signaling pathway, resulting in a delay in early epithelial differentiation, immature distal epithelium and dysfunctional mesenchyme. CONCLUSIONS: The nitrofen model effectively reproduces PH as it disrupts pathways that are critical for lung branching morphogenesis and alveolar differentiation. The low CDH rate confirms that PH is an associated phenomenon rather than the result of mechanical compression alone.

Accuracy of assessment of luteal morphology and luteal blood flow for prediction of early pregnancy in goats.

This study aims to determine the best post-breeding time for an early pregnancy diagnosis in dairy goats, when using luteal morphology and vascularization assessment by B-Mode and color Doppler ultrasonography (US) in association or not with the anechoic uterine content. A total of 131 Saanen goats (2.0 ±â€¯0.5 years old) were used in the two studies. In the first study, pregnancy prediction was daily performed from Day 15-23 post-breeding in 51 does. This was based on a subjective assessment of luteal morphology (B-Mode US), luteal blood flow (color Doppler US), and the presence of anechoic uterine content (B-Mode US). In the second study, pregnancy predictions were performed in 71 does on the best post-breeding day, as determined in the first study (Day 21 using luteal blood-flow assessment and Day 23 using luteal morphological assessment and overall analysis). In both studies, pregnancy diagnosis was confirmed on Day 30 (gold standard method by B-Mode transrectal ultrasonography). The B-Mode and color Doppler US performance in reaching an early pregnancy diagnosis was evaluated by calculating sensitivity (Sens), specificity (Spec), negative and positive predictive values (NPV and PPV), accuracy (Acc), the Kappa index (κ), and the Younden index (J). In Study 1, the pregnancy prediction accuracy progressively increased from Day 17-23 using B-Mode US (D17: 50.98%; D18: 52.94%; D19: 62.75%; D20: 74.51%; D21: 86.27%; D22: 90.20%; D23: 96.08%), and from Day 17-21 using color Doppler US (D17: 49.02%; D18: 54.90%; D19: 70.59%; D20: 86.27%; D21: 96.08%). In the second study, color Doppler, B-Mode, and overall assessments at Day 21 and Day 23, respectively, presented a similar pattern of sensitivity, specificity, and accuracy as was found in the first study. Therefore, both color Doppler and B-Mode assessments can be judged as effective tools for reaching a pregnancy diagnosis in goats as early as on Day 21 and Day 23, respectively, post-breeding.

Quantitative assessment of cervical softening during pregnancy in the Rhesus macaque with shear wave elasticity imaging.

Abnormal parturition, e.g. pre- or post-term birth, is associated with maternal and neonatal morbidity and increased economic burden. This could potentially be prevented by accurate detection of abnormal softening of the uterine cervix. Shear wave elasticity imaging (SWEI) techniques that quantify tissue softness, such as shear wave speed (SWS) measurement, are promising for evaluation of the cervix. Still, interpretation of results can be complicated by biological variability (i.e. spatial variations of cervix stiffness, parity), as well as by experimental factors (i.e. type of transducer, posture during scanning). Here we investigated the ability of SWEI to detect cervical softening, as well as sources of SWS variability that can affect this task, in the pregnant and nonpregnant Rhesus macaque. Specifically, we evaluated SWS differences when imaging the cervix transabdominally with a typical linear array abdominal transducer, and transrectally with a prototype intracavitary linear array transducer. Linear mixed effects (LME) models were used to model SWS as a function of menstrual cycle day (in nonpregnant animals) and gestational age (in pregnant animals). Other variables included parity, shear wave direction, and cervix side (anterior versus posterior). In the nonpregnant cervix, the LME model indicated that SWS increased by 2% (95% confidence interval 0-3%) per day, starting eight days before menstruation. During pregnancy, SWS significantly decreased at a rate of 6% (95% CI 5-7%) per week (intracavitary approach) and 3% (95% CI 2-4%) per week (transabdominal approach), and interactions between the scanning approach and other fixed effects were also significant. These results suggest that, while absolute SWS values are influenced by factors such as scanning approach and SWEI implementation, these sources of variability do not compromise the sensitivity of SWEI to cervical softening. Our results also highlight the importance of standardizing SWEI approaches to improve their accuracy for cervical assessment.

In vivo assessment of the placental anatomy and perfusion in a mouse model of intrauterine inflammation.

BACKGROUND: Magnetic resonance imaging (MRI) provides useful markers to examine placental function. MRI features of placental injury due to intrauterine inflammation-a common risk during pregnancy, are not well known. PURPOSE: To investigate the capability of structural MRI and intravoxel incoherent motion (IVIM) imaging in examining acute placental injury in a mouse model of intrauterine inflammation, as well as gestation-dependent placental changes. STUDY TYPE: Prospective study. ANIMAL MODEL: Pregnant CD1 mice were scanned on embryonic day 15 (E15, n = 40 placentas from six dams) and E17. On E17, mice were subjected to intrauterine injury by exposure to lipopolysaccharide (LPS, n = 25 placentas from three dams) or sham injury (n = 25 placentas from three dams). FIELD STRENGTH/SEQUENCE: In vivo MRI was performed on an 11.7T Bruker scanner, using a fast spin-echo sequence and a diffusion-weighted echo-planar imaging (EPI) sequence. ASSESSMENT: T2 -weighted MRI was acquired to evaluate placental volume. IVIM imaging was performed in a restricted field-of-view using 15 b-values from 10-800 s/mm2 , based on which, the pseudodiffusion fraction (f), pseudodiffusion coefficient (D*), and tissue water coefficient (D) were estimated with a two-step fitting procedure. STATISTICAL TESTS: Two-way analysis of variance (ANOVA) was used to evaluate the group differences. RESULTS: The placental volume increased by ∼21% from E15 to E17 (P < 0.01), and a 15% volume loss was observed at 6 hours after LPS exposure (P < 0.01). IVIM parameters (f, D*, and f·D*) were similar between the E15 and E17 sham groups (P > 0.05), which was significantly reduced in the LPS-exposed placentas compared to the shams (P < 0.001). D values decreased from E15 to E17 (P < 0.05), which were further reduced after LPS exposure (P < 0.05). Changes in placental area and vascular density were histologically identified in the LPS-exposed group, along with gestation-dependent changes. DATA CONCLUSION: Our results suggested structural MRI and IVIM measurements are potential markers for detecting acute placental injury after intrauterine inflammation. LEVEL OF EVIDENCE: 1 Technical Efficacy: Stage 2 J. Magn. Reson. Imaging 2018;47:1260-1267.

Early pregnancy diagnosis in ewes by subjective assessment of luteal vascularisation using colour Doppler ultrasonography.

This study was designed to evaluate early pregnancy diagnosis using colour Doppler ultrasonography (US) for luteal vascularisation assessment. In Study 1, 28 ewes were artificially inseminated (Day 0), and luteal vascularisation was assessed from Day 12 to Day 19 by two evaluators using colour Doppler US, categorising the corpus luteum (CL) on a subjective scale ranging from 1 to 4. Females bearing a CL with score 2 or greater were presumably considered pregnant. Pregnancy was confirmed on Day 30 by B-Mode US. In Study 2, a predictive pregnancy diagnosis was performed on Day 17 in 197 ewes based on the criteria described in Study 1. Pregnancy was confirmed by B-mode US on Day 45. Agreement between evaluators was verified by an intraclass correlation coefficient (ICC) and Kappa index (κ). Performance of colour Doppler US for early pregnancy diagnosis was evaluated calculating sensitivity (Sens), specificity (Spec), negative predictive values (NPV), positive predictive values (PPV) and accuracy (Ac). In Study 1, luteal vascularisation assessment was unable to predict non-pregnant animals between 12 and 14 days after insemination, as all animals still had vascularised CL, and thus were considered pregnant. The colour Doppler US performance improved progressively until Day 17, when it reached maximum values (Sens = 100%, Spec = 76%, PPV = 73%, NPV = 100% and Ac = 86%). The subjective scale for luteal irrigation assessment showed medium to good agreement among evaluators on Day 12 and Day 13 (ICC = 0.66 and 0.68, respectively), and excellent agreement from Day 14 to Day 19 (ICC = 0.90, 0.80, 0.80, 0.84, 0.95 and 0.93, respectively). Agreement was almost perfect for score 1 CLs (κ = 0.87), and moderate for scores 2, 3 and 4 CLs (κ = 0.54, 0.48 and 0.41, respectively). In Study 2, performance of colour Doppler US as a tool to predict pregnancy status in ewes on Day 17 post-insemination was as follows: Sens = 93.5%, Spec = 80.8%, PPV = 85.6%, NPV = 91.1% and Ac = 87.8%. Subjective luteal vascularisation assessment using colour Doppler US to distinguish between pregnant and non-pregnant animals was considered a reliable tool which was highly efficient beginning 17 days after breeding.

Viabilidade econômica de vacas leiteiras F1 Holandês x Zebu com diferentes bases maternas e ordens de parto / Economic viability F1 Holstein x Zebu cows of dairy from different maternal bases and birth orders

Objetivou-se avaliar a viabilidade econômica de vacas F1 de Holandês x Zebu de diferentes bases maternas e ordens de parto. Foram utilizados dados do Campo Experimental da Epamig (Felixlândia-MG). Analisaram-se 406 lactações de vacas F1 H x Z de diferentes bases maternas. Estimaram-se a receita e os custos operacional efetivo, operacional total e custo total. As F1 Holandês x Gir tiveram médias de custos maiores (R$ 3582,30), seguidas pelas F1 Holandês x Nelogir (R$ 3448,06), e o menor foi das F1 Holandês x Nelore (R$ 3145,07). A receita total foi maior para as vacas F1 Holandês x Gir e F1 Holandês x Nelogir, com valores de R$ 4394,96 e R$ 4245,61, respectivamente, e a menor receita foi para F1 Holandês x Nelore, com valor de R$ 3976,12. O lucro foi de R$ 812, 65; R$ 797,54 e R$ 831,04 para as F1 filhas de base materna Gir, Nelogir e Nelore, respectivamente. Todas as bases maternas são viáveis economicamente e podem ser utilizadas no sistema de produção de leite e bezerros para venda. Todas as ordens de parto de Holandês x Zebu estudadas são viáveis economicamente.(AU)

The objective was to evaluate the economic viability of a milk production system of F1 Holstein x Zebu cows of different maternal bases and calving order. Lactations data of F1 Holtein x Zebu cows (n=406) of different maternal bases, from the experimental field of the EPAMIG at Felixlândia county (MG state, Brasil) were analyzed. Revenue, cost/effect of operation, total operating, and total cost have been estimated. F1 cows Holstein x Gir had averages of higher costs R$ 3582.30, followed by F1 cows Holstein x Nelogir R$ 3448.06; and the lowest was from cows F1 Holstein x Nellore with R$ 3145.07. The total revenue was higher for Holstein x Gir and Holstein x Nelogir with recipe values of R$ 4394.96 and R$ 4245.61 respectively, and the lowest value of R$ 3976.12 for Holstein x Nellore. The profits were R$812.65; R$ 797.54 and R$ 831.04 for animals of groups Holstein x Gir, Holstein x Nelogir and Holstein x Nellore respectively. All genetic groups are economically viable and can be used in the production of milk and calves for sale. All calving order of Holstein x Zebu studied are viable economically.(AU)

Dystocia in the cat evaluated using an insurance database.

Objectives The aim of this study was to describe the incidence of feline dystocia with respect to breed. Methods The data used were reimbursed claims for veterinary care insurance and/or life insurance claims in cats registered in a Swedish insurance database from 1999-2006. Results The incidence rates for dystocia were about 22 cats per 10,000 cat-years at risk, 67 per 10,000 for purebred cats and seven per 10,000 for domestic shorthair cats. The median age was 2.5 years. A significant effect of breed was seen. An incidence rate ratio (IRR) that was significantly higher compared with other purebred cats was seen in the British Shorthair (IRR 2.5), the Oriental group (IRR 2.2), Birman (IRR 1.7), Ragdoll (IRR 1.5) and the Abyssinian group (IRR 1.5). A significantly lower IRR was seen in the Norwegian Forest Cat (IRR 0.38), the Maine Coon (IRR 0.48), the Persian/Exotic group (IRR 0.49) and the Cornish Rex (IRR 0.50). No common factor among the high-risk breeds explained their high risk for dystocia. There was no effect of location; that is, the incidence rate did not differ depending on whether the cat lived in an urban or rural area. Caesarean section was performed in 56% of the cats with dystocia, and the case fatality was 2%. Conclusions and relevance The incidence rate for dystocia was of a similar magnitude in purebred cats as in dogs. The IRR varied significantly among breeds, and the main cause for dystocia should be identified separately for each breed. A selection for easy parturitions in breeding programmes is suggested.

Assessment of safety and reproductive performance after vaccination with a modified live-virus PRRS genotype 1 vaccine in pregnant sows at various stages of gestation.

The objective of the present study was to assess safety and efficacy of a new modified live-virus porcine reproductive and respiratory syndrome (PRRS) genotype 1 vaccine in pregnant sows at various stages of gestation under field conditions. A total of 505 sows and gilts were allocated to two treatment groups and maintained in separate facilities. Animals of group 1 were vaccinated with a commercial modified live genotype 1 PRRSV vaccine (control product, CP), while animals of group 2 were immunized with a new modified live genotype 1 PRRSV vaccine (investigational veterinary product, IVP) (ReproCyc® PRRS EU, Boehringer Ingelheim Vetmedica GmbH). Injection site reactions were noted to be significantly less frequent in the IVP group compared to the CP group for pain (p=0.039), redness (p=0.030), heat (p=0.016) and swelling (p=0.002). The mean total number of piglets alive at weaning did not differ significantly between both study groups (10.6 vs. 11.0, p=0.375). However, pre-weaning mortality was significantly higher (p=0.005) in piglets from the CP group (14.1% vs. 10.9%). Analyses of reproductive performance data for both groups did not result in statistically significant differences between CP group and IVP group for number of piglets alive (12.7 and 12.6, respectively), healthy live (11.9 and 11.8), weak (0.7 and 0.5), stillborn (1.0 and 0.8) and mummified piglets (0.3 and 0.2) per litter. No differences were detected between both groups for piglet birth weights, while body weights at weaning (7.2kg vs. 6.6kg, p=0.026) and average daily gain (0.2445kg vs. 0.2211kg, p=0.037) were significantly higher in piglets from the IVP group. In conclusion, the administration of a single dose of ReproCyc® PRRS EU to sows and gilts at various stages of gestation confirmed non-inferiority to a commercial PRRS vaccine regarding safety and efficacy parameters under field conditions.

Interferon stimulated genes as peripheral diagnostic markers of early pregnancy in sheep: a critical assessment.

We investigated the diagnostic reliability of pregnancy detection using changes in interferon stimulated gene (ISG) messenger RNA (mRNA) levels in circulating immune cells in ewes. Two different groups of ewes (an experimental group, experiment 1 and a farm group, experiment 2) were oestrus-synchronized and blood sampled on day 14 (D0=day of insemination in control animals, experiment 1) and day 15 (experiment 2). Real-time PCR were performed to evaluate the abundance of different ISG mRNAs. In the experimental group, peripheral blood mononuclear cells of 29 ewes born and bred in experimental facilities were isolated using a Percoll gradient method. Gene expression for Chemokine (C-X-C motif) ligand 10 (CXCL10), Myxovirus (influenza virus) resistance 1 (MX1) and Signal transducer and activator of transcription 1 (STAT1) mRNA were, respectively, 8.3-fold, 6.1-fold and 2.7-fold higher (P0.10) in CXCL10, STAT1, MX1, Myxovirus (influenza virus) resistance 2 (MX2) and ISG15 ubiquitin-like modifier (ISG15) mRNA expression were found between pregnant and non-pregnant ewes. The ROC curves and the hierarchical classification generated from the real-time PCR data failed to discriminate between pregnant and non-pregnant animals. In this group of animals, our results show a strong variability in ISG expression patterns: 17% of animals identified as non-pregnant by the five tests were in fact pregnant, only 52% of pregnant animals had at least two positive results (two genes above threshold), whereas up to five positive results (five genes above threshold) were needed to avoid misclassification. In conclusion, this study illustrates the high variability in ISG expression levels in immune circulating cells during early pregnancy and, therefore, highlights the limits of using ISG expression levels in blood samples, collected on PAXgene® tubes on farms, for early pregnancy detection in sheep.

Assessment of body composition in Wistar rat offspring by DXA in relation to prenatal and postnatal nutritional manipulation.

BACKGROUND: This experimental study aims to investigate the impact of combinations of prenatal and postnatal food manipulation on body composition in rat offspring. METHODS: On day 12 of gestation, 100 timed pregnant rats were randomized into two nutritional groups: standard laboratory and 50% starved. Pups born to starved mothers were subdivided, based on birthweight (BiW), into fetal growth restricted (FGR) and non-FGR. Pups were born on day 21, cross-fostered, then left undisturbed lactating until the 26th postnatal day when they underwent dual-energy X-ray absorptiometry (DXA) examination. RESULTS: Prenatally control-fed animals had a significantly greater body weight at 26 d postnatally than the prenatally starved groups, irrespective of their postnatal diet (P < 0.001). Postnatal control diet was associated with significantly increased abdominal and total fat in non-FGR compared to FGR rats (P < 0.001). non-FGR/CONTROL rats showed higher values of abdominal fat than prenatally starved animals that were starved postnatally irrespective of their birth weight (P < 0.001). Postnatal control diet significantly increased total bone mineral content (BMC), head BMC, head area, abdominal BMC in non-FGR compared to FGR rats (P < 0.001). CONCLUSION: Interaction between prenatal and postnatal nutrition affects growth, abdominal adiposity, and bone accrual in Wistar rats' offspring at 26 d of life.

Assessment of the accuracy of estimation of gestational age in cattle from placentome size using inverse regression.

AIMS: To assess, in dairy cows between 60-160 days of gestation, whether the agreement between actual gestational age and that predicted by placentome size could be improved by using individual placentome measurements rather than a single average, and to identify how increasing the number of placentomes measured improved the prediction. METHODS: Data were obtained from 58 cows examined using transrectal ultrasonography every 10 days between 60-130 days, and every 15 days between 130-160 days of gestation. For each cow, four to six placentomes in each of the pregnant and non-pregnant horns were examined from the region of the uterus near to the cervix. A mixed-effects model, which included cow as a random effect, and a simple linear regression which ignored within-cow correlation, were fitted to the data. Inverse regression was used to compare the 95% prediction bands obtained for estimating gestational age using the means of three, five or eight placentome measurements. RESULTS: The fit of the mixed effects model was better than a simple linear regression (p<0.001) but the fitted lines from the two models were very similar. Using the simple regression model, for a mean placentome length of 15 mm estimated gestational age was 100 days, with 95% CI of 68-131 days for measurement of three placentomes, and 80-120 days for eight placentomes. CONCLUSIONS: The agreement between placentome size and gestational age was improved by increasing the number of placentomes measured. Direct comparison of these CI with those for other published fetal measures was not possible as similar prediction bands have not previously been calculated for fetal measures; however one study reported a residual SD which had been calculated using size measurements as the predictor variable and age as the response variable. Using these data 95% CI were calculated to be ±9 days for crown-rump length and ±25 days for uterine diameter. These are likely to be an underestimate of the true CI and do not take account of the increase in variance of the difference between predicted and actual gestational age as gestational age increases. CLINICAL RELEVANCE: Placentome measurement may be useful for estimating fetal age in late gestation. Further research is required to better establish the agreement between gestational age and the fetal measurements which are currently used to estimate fetal age.

Assessment of Maternal Vascular Remodeling During Pregnancy in the Mouse Uterus.

The placenta mediates the exchange of factors such as gases and nutrients between mother and fetus and has specific demands for supply of blood from the maternal circulation. The maternal uterine vasculature needs to adapt to this temporary demand and the success of this arterial remodeling process has implications for fetal growth. Cells of the maternal immune system, especially natural killer (NK) cells, play a critical role in this process. Here we describe a method to assess the degree of remodeling of maternal spiral arteries during mouse pregnancy. Hematoxylin and eosin-stained tissue sections are scanned and the size of the vessels analysed. As a complementary validation method, we also present a qualitative assessment for the success of the remodeling process by immunohistochemical detection of smooth muscle actin (SMA), which normally disappears from within the arterial vascular media at mid-gestation. Together, these methods enable determination of an important parameter of the pregnancy phenotype. These results can be combined with other endpoints of mouse pregnancy to provide insight into the mechanisms underlying pregnancy-related complications.

Assessment of lipid and protein peroxidation markers in non-pregnant and pregnant female dogs.

The aim of the study was to investigate oxidative stress during normal pregnancy in female dogs based on an evaluation of plasma markers for lipid and protein peroxidation. Twenty clinically healthy female dogs (10 non-pregnant and 10 pregnant) were used in the study. Blood samples from the pregnant animals were collected at 19-21, 38-40, and 56-58 days of pregnancy. Blood samples from non-pregnant female dogs were obtained between 20 and 35 days after ineffective breeding. As indicators of oxidative stress, we measured the following using spectrophotometric and spectrof- luorimetric methods: thiobarbituric acid reactive substances (TBARS), radical cations of N,N, diethylparaphenylene diamine (RC-DEPPD), sulfhydryl groups (SH groups), bityrosine and formylkynurenine. The mean plasma TBARS concentration in the pregnant dogs (0.486 ± 0.071-0.581 ± 0.191 µmol/g protein) was significantly higher (p < 0.05) than that found in the non-pregnant animals (0.274 ± 0.111 µmol/g protein). A marked, although not significant, decrease in SH group content, as well as an increase in bityrosine and formylkynurenine concentration were concurrently observed in the pregnant dogs. No significant differences were found in terms of the studied markers in the pregnant animals when comparing the values obtained during the investigated periods of pregnancy, although there was a progressive decrease in TBARS concentration and a progressive increase in RC-DEPPD, bityrosine and formylkynurenine contents. Our findings suggest that normal pregnancy in female dogs is associated with oxidative stress. Further studies are necessary to establish the physiological ranges of antioxidative/oxidative profiles in pregnant dogs and to explain if and how the intensity of oxidative stress might contribute to the risk of the complications of pregnancy.

Assessment of the reproductive physiology of the potto (Perodicticus potto) through fecal hormone metabolite analyses and trans-abdominal ultrasonography.

Potto (Perodicticus potto) reproductive biology has been minimally studied. Noninvasive endocrinology and ultrasonography are proven tools for reproductive assessment in other primates. In this study, we used fecal hormone metabolite analysis to monitor one adult male potto and four females at different life stages. Validated testosterone (T), estrone conjugate (EC), and progesterone (P4) enzyme immunoassays (EIA) were used to assess male testicular function and female ovarian and placental activity. The male excreted mean T concentrations of 4.72 (±1.66) µg/g feces, that did not differ (P > 0.05) over time or when paired with alternate females. Baseline concentrations of EC (range: 47.93-78.81 ng/g feces) and P4 (range: 2.29-12.46 µg/g feces) differed among adult females. Follicular phases averaged 9.1 days (±3.43, n = 30 phases), whereas luteal phases averaged 19.89 days (±9.49, n = 19 phases). Gestation length (n = 2 pregnancies) was 170 days. Gestational EC and P4 concentrations were positively correlated (pregnancy A, r (132) = 0.71; pregnancy B, r (145) = 0.76) and returned to non-pregnant luteal phase levels 3-7 days post parturition. Extreme differences between pregnant and non-pregnant EC and P4 concentrations may allow for one-sample pregnancy diagnosis. Trans-abdominal ultrasonography was validated for pregnancy diagnosis with the fetus observed between 100 and 110 days post breeding. To our knowledge, this is the first use of fecal endocrinology and ultrasonography to monitor reproductive function and pregnancy in this species, and the only study in any lorisid to measure progestagens in correlation with reproductive events.

Assessment of immune response in periparturient dairy cows using ex vivo whole blood stimulation assay with lipopolysaccharides and carrageenan skin test.

The transition period is known to be the most critical phase in the life of high yielding dairy cow. Changes in the immune functions have been observed during the transition period which may account for the onset of clinical and subclinical (e.g. inflammatory response) problems at calving or at the beginning of lactation however this relationship has not yet been adequately investigated. Thus, to establish the potential of the periparturient dairy cow's immune system to respond to stimuli, two challenges [an ex vivo whole blood stimulation assay (WBA) with lipopolysaccharides and a carrageenan skin test (CST)] were performed in addition to characterizing the metabolic and inflammatory profile. The WBA was performed using 0, 0.01 and 5 µg LPS/mL on whole blood and CST was administered by subcutaneous injection of 0.7 mL solution containing 4.2mg of carrageenan to the shoulder region of the cows. These tests were performed on 10 Holstein-Friesian cows at -45 ± 2, -20 ± 2, -3, 3, 7, 28 ± 2 days from parturition (DFP). Cows were also monitored for health status, body condition score, milk yield. The results demonstrate a higher production of IL-1ß and IL-6 from leukocytes after LPS stimulation around calving (from -3 to 3 DFP) compared to -45 DFP (P < 0.05). Moreover, IL-6 (but not IL-1ß) was able to reach close to the maximum response at the lower stimulus intensity (0.01 µg LPS/mL), maintaining a higher response over a longer time in early lactation. The release of higher levels of IL-6 in the transition period, with low LPS dose, suggests its crucial role in the regulation of inflammatory response around calving. The response of cows to CST decreased a few days before calving (-3 DFP) compared with response at -45 and 28 DFP (P<0.05), and remained low in the first week of lactation. This result suggests the reduction of the functionality of some vascular factors, which decreases diapedesis. Overall, the WBA and CST tests confirm changes in immunocompetence around calving. These tests are able to better describe the changes of the innate immune response at a local and systemic level, mainly when combined with conventional metabolic and inflammatory indices.

Are we ready to consider transgenerational epigenetic effects in human health risk assessment?

Recently, there has been a growing concern that chemically or nutritionally mediated epigenetic changes might lead to adverse health outcomes. The natural question is whether the existing chemical safety assessment paradigm is or is not protective of epigenetic-mediated effects, and if there is a need to incorporate new endpoints to specifically address epigenetics. Of particular interest are transgenerational epigenetic effects, which can be passed on through multiple generations. To investigate these questions, a comparison was performed between OECD guideline rat toxicology studies versus several rat transgenerational epigenetic studies. This analysis focused on vinclozolin owing to the availability of a comprehensive suite of dose-response data (NOAEL, reference dose, and human exposure estimates) for both conventional and epigenetic endpoints. This analysis revealed that vinclozolin transgenerational effects were demonstrated at a dose level (100 mg/kg/day) that was: (1) ∼40-fold higher than the overall lowest-observed-adverse-effect level (LOAEL) from rat guideline studies, (2) ∼80-fold higher than the lowest NOAEL from rat guideline studies, (3) ∼80,000-fold higher than the reference dose for the molecule, and (4) ∼1.2-million fold above human exposure estimates. Through this analysis, we conclude that additional research across a spectrum of doses is necessary to elucidate the interplay between epigenetics and apical endpoints before considering epigenetics in human health risk assessment. Therefore, we recommend focusing future research toward (1) examining for potential causal relationships between epigenetic alterations and adverse apical endpoints, and (2) understanding the dose-response relationship of these causal epigenetic alterations when compared with those of the apical endpoints.