Effect of Prenatal and Neonatal Anti‐Androgen Flutamide Treatment on Aquaporin 5 Expression in the Adult Porcine Ovary

The growth of ovarian follicles is accompanied by fluid‐filled antrum formation. Water movement within the follicular wall is predominantly transcellular via membranous water channels named aquaporins (AQPs). Androgens are important regulators of mammalian folliculogenesis, and their prenatal and/or neonatal deficiency affects female fertility in adulthood. Therefore, this study was performed to determine whether gestational or neonatal exposure to the anti‐androgen flutamide influences androgen‐dependent AQP5 expression in pre‐antral and large antral follicles of adult pigs. Flutamide was injected into pregnant gilts between days 80 and 88 of gestation and into female piglets between days 2 and 10 post‐natally. The ovaries were collected from flutamide‐treated and non‐treated (control) sexually mature pigs. In pre‐antral follicles, AQP5 mRNA and protein levels were both downregulated following maternal (p < 0.01 and p < 0.01, respectively) and neonatal (p < 0.01 and p < 0.01, respectively) flutamide exposure. Likewise, the expression of mRNA (p < 0.01 and p < 0.001, respectively) and protein (p < 0.05 and p < 0.01, respectively) for AQP5 were diminished in large antral follicles in both groups. Immunohistochemistry showed decreased intensity of AQP5 immunoreaction in pre‐antral (p < 0.01) and large antral (p < 0.001) follicles following flutamide treatment. Moreover, radioimmunological analysis revealed that changes observed in AQP5 expression corresponded with diminished follicular androgens production after both maternal (p < 0.05 and p < 0.05, respectively) and neonatal (p < 0.05 and p < 0.01, respectively) flutamide administration. Therefore, AQP5 appears to be a potential regulator of follicular fluid accumulation, under androgen control, and may be a key factor in antral follicle growth.     

Experimental Neospora Caninum Infection in Pregnant Dairy Heifers Raises Concentrations of Pregnancy‐Associated Glycoproteins 1 and 2 in Foetal Fluids

Plasma concentrations of PAG‐1 are used for pregnancy diagnosis and as a marker of placental/foetal well‐being, while those of PAG‐2 may be an indicator of abortion risk in Neospora caninum‐infected cows. Studies have shown that N. caninum infection modifies PAG‐1 and PAG‐2 patterns in maternal blood plasma. However, no prior work has examined the effects of N. caninum infection on concentrations of PAGs in foetal fluids. In this study, PAG‐1, PAG‐2 and pH levels were determined in the amniotic and allantoic fluids of foetuses collected at 152 days of gestation from control uninfected dams and from dams experimentally infected with N. caninum on Day 110 of gestation. Foetal fluids from infected foetuses had significantly higher PAG‐2 concentrations (p = 0.026) and pH values (p = 0.02) than fluids from non‐infected foetuses. In infected foetuses, significantly higher concentrations of PAG‐1 (p < 0.001) and PAG‐2 (p < 0.001) were detected in fluid samples showing antibodies against N. caninum than those without antibodies. Moreover, pH values were significantly higher (p = 0.011) in foetal fluid samples with antibodies than in samples from non‐infected foetuses. In conclusion, this is the first report on the effect of N. caninum infection on PAG levels in foetal fluids. Our results indicate that following the experimental infection of dams with N. caninum on Day 110 of gestation, foetal fluids collected from the infected foetuses of these dams featured higher PAG‐1 and PAG‐2 levels and pH values than fluids from non‐infected controls, provided that the samples tested showed the presence of antibodies. The clinical implications of these findings are that following infection with N. caninum, most cows will experience some level of placental damage and that this injury correlates with foetal fluid PAG levels and pH.     

Altered Expression of 3β‐HSD,CYP17 and 17β‐HSD in the Foetal Porcine Gonads in Response to Anti‐androgen Flutamide Exposure

We investigated whether the limited access to androgens during late prenatal period alters expression of steroidogenic enzymes involved in androgen production: 3β‐hydroxysteroid dehydrogenase/Δ5‐Δ4 isomerase (3β‐HSD), cytochrome P450 17α‐hydroxylase/17,20‐lyase (CYP17) and 17β‐hydroxysteroid dehydrogenase type 1 (17β‐HSD1) or type 3 (17β‐HSD3) in the foetal porcine gonads. Pregnant gilts were injected with anti‐androgen flutamide (for seven days, 50 mg/day/kg bw) or corn oil (control) starting at 83 (GD90) or 101 (GD108) gestational day. To assess 3β‐HSD, CYP17 and 17β‐HSD1 or 17β‐HSD3 expression, real‐time PCR and immunohistochemistry were performed. In testes from flutamide‐treated foetuses, increased 3β‐HSD and CYP17 mRNA expression was observed in the GD90 group, while decreased 3β‐HSD and 17β‐HSD3 mRNA expression and increased CYP17 mRNA expression were found in the GD108 group. CYP17 and 17β‐HSD3 were localized in Leydig cells. Following flutamide administration, the intensity of CYP17 immunostaining was higher in both treated groups, while 17β‐HSD3 intensity was lower in the GD108 group. In ovaries from flutamide‐treated foetuses in the GD90 group, mRNA level for 3β‐HSD was elevated, but it was diminished for CYP17 and 17β‐HSD1. In the GD108 group, flutamide treatment led to lower mRNA level for 3β‐HSD but higher for CYP17. 3β‐HSD was found in granulosa cells, while CYP17 was localized within egg nests and oocytes of forming follicles. Following flutamide treatment, the intensity of 3β‐HSD and CYP17 immunostaining was higher in the GD90 and GD108 groups, respectively. Immunohistochemical staining for 3β‐HSD was restricted to the ovary. Concluding, diminished androgen action in the porcine foetal gonads during late gestation induces changes in steroidogenic enzymes expression, which may led to changes in gonadal function. However, it seems that androgens exert diverse biological effects depending on the gestational period.     

Effects of realimentation after nutrient restriction during mid‐ to late gestation on pancreatic digestive enzymes,serum insulin and glucose levels,and insulin‐containing cell cluster morphology

This study examined effects of stage of gestation and nutrient restriction with subsequent realimentation on maternal and foetal bovine pancreatic function. Dietary treatments were assigned on day 30 of pregnancy and included: control (CON; 100% requirements; n = 18) and restricted (R; 60% requirements; n = 30). On day 85, cows were slaughtered (CON, n = 6; R, n = 6), remained on control (CC; n = 12) and restricted (RR; n = 12), or realimented to control (RC; n = 11). On day 140, cows were slaughtered (CC, n = 6; RR, n = 6; RC, n = 5), remained on control (CCC, n = 6; RCC, n = 5) or realimented to control (RRC, n = 6). On day 254, the remaining cows were slaughtered and serum samples were collected from the maternal jugular vein and umbilical cord to determine insulin and glucose concentrations. Pancreases from cows and foetuses were removed, weighed, and subsampled for enzyme and histological analysis. As gestation progressed, maternal pancreatic α‐amylase activity decreased and serum insulin concentrations increased (p ≤ 0.03). Foetal pancreatic trypsin activity increased (p < 0.001) with advancing gestation. Foetal pancreases subjected to realimentation (CCC vs. RCC and RRC) had increased protein and α‐amylase activity at day 254 (p ≤ 0.02), while trypsin (U/g protein; p = 0.02) demonstrated the opposite effect. No treatment effects were observed for maternal or foetal pancreatic insulin‐containing cell clusters. Foetal serum insulin and glucose levels were reduced with advancing gestation (p ≤ 0.03). The largest maternal insulin‐containing cell cluster was not influenced by advancing gestation, while foetal clusters grew throughout (p = 0.01). These effects indicate that maternal digestive enzymes are influenced by nutrient restriction and there is a potential for programming of increased foetal digestive enzyme production resulting from previous maternal nutrient restriction.     

Metabolic Profile of Broodmares During Late Pregnancy and Early Post‐Partum

Pregnancy represents a specific physiological status characterized by continuous adjustments that affect maternal metabolism of all nutrients. In the last trimester of pregnancy, mare's nutrient requirements greatly increase and most pregnancy‐associated diseases are likely to occur. Therefore, we aimed to assess the metabolic profile of broodmares focusing on the last 3 months of pregnancy and the early post‐partum. Fifteen pregnant mares (Group A) were monitored from 263 ± 3 days of pregnancy until 21 days after foaling. Seven non‐pregnant mares (Group B) were used as the control group. Blood samples were collected weekly by jugular venipuncture throughout the experimental period, and additional blood samples were collected within 24 ± 12 h of foaling. Obtained sera were analyzed for urea (Ur), creatinine (Cre), total protein (TP), total, direct and indirect bilirubin (tB, dB, iB), triglycerides (TG), total cholesterol (tChol) and β‐hydroxybutyrate (BHB). During pregnancy, Ur (p = 0.015) and dB (p = 0.028) were higher in Group A than Group B. Serum Cre, tB, iB and TG (p < 0.001) decreased after foaling; furthermore, Group A exhibited lower tChol (p < 0.001) and higher BHB (p < 0.001) than Group B during the study. There were significant interactions of time × peripartum on Ur (p = 0.007), Cre (p < 0.001), tB (p = 0.018), TG (p < 0.001) and tChol (p < 0.001). These results can be used in the interpretation of biochemical studies in late gestation mares which develop systemic disease. The improved understanding of the metabolic profile during the peripartum might assist in monitoring the health status of the broodmares to promote foetal growth and well‐being.     

Intestinal development of bovine foetuses during gestation is affected by foetal sex and maternal nutrition

We aimed to evaluate the effects of maternal nutrition (MN) and foetal sex on the intestinal development of bovine foetuses throughout different days of gestation (DG). Forty‐four multiparous, dry Holstein × Gyr cows with average initial body weight of 480 ± 10 kg were fed the same diet of either restricted feeding at 1.15% of body weight (CO, n = 24) or fed ad libitum (overnourished, ON, n = 20). Six cows from CO group and five cows from ON group were slaughtered at 139, 199, 241 and 268 DG, and foetuses were necropsied to evaluate the intestinal development. The mass, length and density of foetal intestines were not affected by MN (p ≥ 0.260). An interaction between MN and DG was observed for the villi length of jejunum (p = 0.006) and ileum (p < 0.001). Villi length of jejunum and ileum was higher (p < 0.10) in foetuses from ON‐fed cows than in foetuses from CO‐fed cows at 139 DG. However, at 199 DG, the villi length of jejunum and ileum of foetuses from CO‐fed cows was higher than in foetuses from ON‐fed cows. Despite these differences, MN did not affect the villi length of jejunum and ileum at 268 DG (p > 0.10). Female foetuses had greater small intestine mass (p = 0.093), large intestine mass (p = 0.022), small intestine mass in proportion to body mass (p = 0.017) and large intestine mass in proportion to body mass (p < 0.001) than male foetuses. Female foetuses had also longer small intestine (p = 0.077) and greater small intestine density (p = 0.021) and villi length of jejunum (p = 0.001) and ileum (p = 0.010) than males. We conclude that MN affects the pathway for the development of foetal villi length throughout the gestation in bovine foetuses without changing the final villi length. Female foetuses had higher intestinal mass, density and villi length than males during the foetal phase in bovines.     

The effect of maternal obesity on fatty acid transporter expression and lipid metabolism in the full‐term placenta of lean breed swine

This study was conducted to evaluate the influence of back‐fat thickness (BF), at mating of sows, on the maternal and newborn circulating lipids, expression of placental fatty acids (FA) transporters and lipid accumulation in placenta. Full‐term placentas were obtained by vaginal delivery from BFI (9–14 mm; n = 37), BFII (15–19 mm; n = 43) and BFIII (20–27 mm; n = 38) sows according to BF at mating, and frozen placental sections were analysed for fat accumulation. Blood samples were collected from the sows of day 105 pregnancy and from cord blood at delivery. mRNA and protein expression levels were evaluated with real‐time RT‐PCR and Western blotting. Our results demonstrated that BFII females had significantly increased litter weight and placental efficiency, decreased maternal triglyceride (TG) and non‐esterified fatty acids (NEFA) levels, decreased maternal IL‐6, TNFα and leptin levels compared to BFIII females (p < .05). BFIII sows were associated with significantly decreased newborn TG levels, increased newborn glucose, IL‐6 and TNFα levels compared to BFI or BFII sows (p < .05). BFI and BFII females had significantly decreased placental TG, NEFA and cholesterol (CHOL) contents compared to BFIII females (p < .05). Moreover, decreased CD36, FATP1, FABP4, and FABP1 mRNA and protein and FATP4 protein expression, and increased LPL activity were also observed in BFIII group compared with BFII group (p < .05). PPARγ mRNA and protein and lipogenic genes such as SREBP‐1c, ACSL1, ACCα, FAS and SCD mRNA expression were downregulated or upregulated, respectively, in the placentas of BFIII sows compared to BFI or BFII sows (p < .05). Overall, this study demonstrated that there is no advantage, in terms of litter live size, litter weight and placental FA transport and metabolism, in performing the mating of sows with BF>19 mm.     

Pregnancy Loss in Dairy Cattle: Relationship of Ultrasound,Blood Pregnancy‐Specific Protein B,Progesterone and Production Variables

Objectives were to determine associations between percentage pregnancy loss (PPL) in dairy cattle and: (i) pregnancy diagnosis by ultrasonography; (ii) pregnancy diagnosis by serum pregnancy‐specific protein B (PSPB) concentrations, with or without serum progesterone concentrations; and (iii) production and environmental factors. This study included 149 822 pregnancy diagnoses conducted over 13 years in Holstein‐Friesian cows in Hungarian dairy herds. The following were determined: PPL in cows diagnosed pregnant by transrectal ultrasonography 29–42 days after artificial insemination (AI; n = 11 457); PPL in cows diagnosed pregnant by serum PSPB 29–35 days after AI (n = 138 365); and PPL and its association with serum progesterone concentrations, PSPB and production/environmental variables. The definition of PPL was percentage of cows initially diagnosed pregnant based on ultrasonography or PSPB, but not pregnant when examined by transrectal palpation 60 –70 days after AI. The PPL was lower (p < 0.001) in cows following ultrasonographic vs PSPB diagnosis of pregnancy at 29–35 days (8.1 vs 19.3%, respectively), but was higher in cows following ultrasonographic pregnancy diagnosis on 29–35 vs 36–42 days (8.1 vs 7.1%, respectively, P < 0.05). Furthermore, 72.9% of pregnancies with ultrasound‐detected morphological abnormalities resulted in pregnancy loss. As a subset of PSPB data, a fully quantitative PSPB assay was used for 20 430 samples; PPL in cows with a high PSPB concentration (>1.1 ng/ml) was lowest (15.0%), whereas cows with low concentrations of both PSPB and progesterone (0.6–1.1 and <2 ng/ml, respectively) had the highest PPL (76.3%; p < 0.0001). Furthermore, PPL was higher in cows with advanced parity and with high milk production, when ambient temperatures were high, although body condition score (BCS) had no effect on PPL. Finally, there were no significant associations between serum PSPB and environmental temperatures or number of post‐partum uterine treatments.     

Patterns of protein glycosylation in bovine placentomes as a function of gestational age and in retained versus non‐retained placenta

The formation of placenta at the beginning of pregnancy and its separation at parturition require not only deep remodelling of extracellular matrix, which mainly consists of proteins conjugated with sugar moieties, but also the cooperation with cells from both maternal and foetal parts of placenta. The aim of the study was to compare the patterns of selected conjugated proteins with sugar moieties between pregnant and term placenta as well as between released and retained placenta in cows. Placental samples from healthy pregnant cows (3–5 months of pregnancy) were collected at a slaughterhouse (n = 6), and parturient samples were collected during caesarean section at term and retrospectively divided into retained (n = 6) and released (n = 6). The pattern of selected sugar moieties conjugated with proteins was detected by use of lectin blotting with Phaseolus Vulgaris leucoagglutinin, Maackia Amurensis and Sambucus Nigra (Elderberry). The comparison and analysis of obtained band patterns showed differences between their number, molecular weight and abundance related to the intensity of staining. Samples from 3 to 4 months showed similarities, while at the 5th month, clear differences were visible in all 3 lectins, which were used in this study. Samples from retained/released placenta expressed significant differences in PHA‐L and SNA pattern in the foetal part. Obtained results indicate that the development of placenta related to extracellular matrix and accompanying cells from both sides of placenta shows dynamic changes during pregnancy. Moreover, in the case of animals with the retention of foetal membranes the patterns of proteins conjugated with sugar moieties are altered, suggesting that the changes in extracellular matrix metabolism can be involved in the attachment and detachment of the placenta in cows.     

Differential gene expression of Eph‐ephrin A1 and LEPR‐LEP with high or low number of embryos in pigs during implantation

The objective of this study was to ascertain whether mRNA and protein expressions of implantation‐related genes (erythropoietin‐producing hepatocellular receptor–ligand A1, Eph‐ephrin A1 and leptin receptor–leptin, LEPR‐LEP) differed between pigs with high and low number of embryos, and whether these differences in gene expression might affect embryo implantation. Experimental pig groups (n = 24) for high and low number of embryos were prepared by altering the number of eggs ovulated in pre‐pubertal gilts treated with 1.5 × (High) or 1.0 × (Low) PG600 ([400 IU PMSG + 200 IU hCG]/dose, AKZO‐NOBEL). Gilts expressing oestrus were artificially inseminated twice and maintained in breeding and gestation until the reproductive tract was collected on day 22 of pregnancy. At slaughter, the reproductive tracts from each pregnant gilt from each treatment were immediately processed to collect samples for RNA and protein analysis. Within each gilt, three conceptus points were sampled, one from each horn and then a random conceptus within the tract. At each conceptus point, endometrial attachment site, chorion–allantois and embryo were collected and immediately frozen in liquid nitrogen. Number of corpus luteum (CL) (35.4 vs. 12.6) and total embryo number (18.8 vs. 10.2) were greater in the high‐embryo compared to the low‐embryo group, respectively (p < .05). Real‐time qPCR results showed that Eph‐ephrin A1 mRNA expression was less in the high‐embryo (p < .05) compared to the low‐embryo group. In addition, Western blotting analysis indicated that Eph‐ephrin A1 and LEP protein expression at endometrial attachment site in high‐embryo was less (p < .05) compared to low‐embryo group. It was also noted that mRNA expression of Eph‐ephrin A1 and LEPR‐LEP was greater in pregnant than non‐pregnant gilts (p < .05). Moreover, mRNA expression of Eph‐ephrin A1 (p < .05) and LEPR‐LEP was greatest at endometrial attachment site among all three tissues. There was a positive correlation between expressions of Eph‐ephrin A1, LEPR‐LEP and embryo length with the correlation coefficient 0.31–0.59. For Eph‐ephrin A1, the highest correlation coefficient appeared between Eph A1 expression and normal embryo number, between ephrin A1 expression and embryo length. For LEPR‐LEP, the highest correlation coefficient appeared between LEPR‐LEP expression and ovary weight (0.79 for both, p < .05), followed by embryo length and weight. The results of this study suggest that low expression of Eph‐ephrin A1 and LEPR‐LEP is somehow related to increased embryo number during implantation and that endometrial attachment site might be the main target tissue of these gene products. Yet, the increased expression of Eph‐ephrin A1 and LEPR‐LEP appeared associated with increased embryo growth (length and weight) and ovary weight, Eph‐ephrin A1 and LEPR‐LEP might play roles in the regulation of embryo implantation in pigs.     

Ultrasound during mid‐gestation: Agreement with physical foetal and placental measurements and use in predicting gestational age in sheep

To determine the effects of poor maternal nutrition and litter size on foetal growth during mid‐gestation, pregnant ewes (n = 82) were fed 100%, 60% or 140% of NRC TDN beginning at day 30.2 ± 0.2 of gestation. Transabdominal ultrasound was performed weekly between day 46.0 ± 0.4 and 86.0 ± 0.7 to monitor foetal heart width (HW), umbilical diameter (UMB), rib width (RW) and placentome outer (OD) and inner diameter (ID). Data were analysed with repeated‐measures using the mixed procedure for effects of maternal diet, litter size and gestation, and equations predictive of gestational age were generated using the regression procedure. To determine the agreement of ultrasound measurement and actual size, ewes (n = 20–21) were euthanized at day 45 or 90 to obtain corresponding postmortem measurements for Bland–Altman analysis. The HW, UMB and placentome OD and ID increased with gestation (p < .0001) but were unaffected by maternal diet or litter size (p ≥ .12). Ultrasound underestimated postmortem measurements of HW (14.8%), UMB (7.3%), placentome OD (4.5%) and ID (37.3%) at day 90 of gestation. Ultrasound underestimated RW at day 45 (7.7%) but overestimated RW (23.8%) at day 90, indicating inconsistent bias when reporting RW by ultrasound. Combining the HW, UMB, RW and placentome OD generated the strongest equation predictive of gestational age (R² = .91). These findings indicate that during mid‐gestation, maternal diet or litter size did not affect HW, UMB or placentome diameters and these factors can be used to estimate gestational age.     

The influence of progesterone and prostaglandin F2α on decorin and the adhesion of caruncular epithelial cells of bovine placenta at early-mid pregnancy—Part II

One of the most important processes determining the proper course of gestation and its physiological termination in cows is the adhesion of epithelial cells allowing for direct contact of maternal and foetal parts of the placenta. Throughout pregnancy, placental cells are under strict hormonal control, which among others regulates the concentration and activity of specific proteins participating in the extracellular matrix remodelling of foetal membranes. The aim of the study was to evaluate the influence of progesterone and prostaglandin F_2α on the adhesion of epithelial cells at early-mid pregnancy in cows. Additionally, the impact of selected hormones on anti-adhesive properties of decorin was evaluated. Caruncular epithelial cells were isolated from healthy cows during pregnancy, immediately after slaughter. Primary cell cultures derived from the 2nd and 4th month of gestation were used in the experiments. The viability of cells was assessed by MTT assay. The adhesion of cells to fibronectin was measured spectrophotometrically. The activity of metalloproteinases was confirmed by the metalloproteinase assay. Progesterone (10^–5 and 10^–7 mol/L) and prostaglandin F_2α (10^–4_, 10^–5 and 10^–7 mol/L) increased the viability of bovine caruncular epithelial cells in the 2nd month of pregnancy. The treatment with prostaglandin F_2α significantly reduced the number of adherent cells from the 2nd month of gestation at the doses of 10^–4 and 10^–5 mol/L. Both progesterone and prostaglandin F_2α were shown to have an effect of decorin resulting in both a decrease in metalloproteinase activity and an increase in adhesion of cells to fibronectin.     

Effects of deficiency and surplus dietary threonine on reproductive performance of primiparous pregnant gilts

Dietary threonine imbalance is known to impair reproductive performances of gestating sows, but the underlying mechanisms are largely unknown. In this study, effects of deficiency and surplus dietary threonine during gestation on reproductive performance, serum metabolites and hormones concentration, and colostral nutrient and immunoglobulin contents of primiparous sows were investigated. Ninety primiparous pregnant gilts were assigned to one of the three dietary treatments with different standardized ileal digestible threonine/lysine ratios at 0.59, 0.72 and 0.85, which represented deficient (DT), adequate (AT) and surplus (ST) dietary threonine concentration respectively. Maternal body weight gain from day 80–110 of gestation was highest (p < .05) for gilts fed AT than for gilts fed DT or ST. On days 30 and 110, serum threonine concentration increases in a dose‐dependent manner with the increasing of dietary threonine concentration in (p < .01), serum urea nitrogen concentration was lower (p < .01) in gilts fed AT than DT or ST, and serum insulin‐like growth factor‐I (IGF‐I) was lowest (p < .05) for gilts fed DT. On day 110, gilts fed AT had lower serum progesterone concentration but higher concentrations of serum prolactin (p < .05) compared to DT and ST. Concentration of colostral immunoglobulin A and G from gilts fed DT was lower (p < .05) compared with gilts fed AT or ST. In conclusion, gilts with the adequate threonine intake were more able to conserve dietary amino acids to support foetal and maternal tissue gain. Deficient or ST threonine intake may induce a delay in changes in progesterone and prolactin concentrations during the prepartum period impeding the transition from pregnancy to lactation.     

Expression of Adiponectin and its Receptors in the Porcine Hypothalamus During the Oestrous Cycle

Adiponectin is a hormonal link between obesity and reproduction, and its actions are mediated by two types of receptors: adiponectin receptor 1 (AdipoR1) and adiponectin receptor 2 (AdipoR2). This study compares the expression levels of adiponectin and adiponectin receptor mRNAs and proteins in selected areas of the porcine hypothalamus responsible for GnRH production and secretion: the mediobasal hypothalamus (MBH), pre‐optic area (POA) and stalk median eminence (SME). The tissue samples were harvested on days 2–3, 10–12, 14–16 and 17–19 of the oestrous cycle. Adiponectin mRNA expression in MBH was significantly lower on days 14–16, whereas in SME, the most pronounced gene expression was found on days 2–3 of the cycle (p < 0.05). Adiponectin protein in MBH was most abundant on days 17–19 and in POA on days 2–3 (p < 0.05). Adiponectin protein expression in SME was at similar level throughout the most of the cycle with a statistically significant drop (p < 0.05) on days 14–16. AdipoR1 gene expression in POA was potentiated on days 2–3 and 10–12 of the oestrous cycle (p < 0.05). In SME, the highest AdipoR1 mRNA expression was noted on days 2–3 (p < 0.05). The concentrations of the AdipoR1 protein in POA were similar throughout the luteal phase (days 2–14 of the cycle), and they decreased on days 17–19 (p < 0.05). In SME, AdipoR1 protein expression peak occurred on days 2–3 (p < 0.05). The expression patterns of the AdipoR2 gene in MBH, POA and SME revealed the highest mRNA levels on days 2–3 of the cycle (p < 0.05). The highest content of AdipoR2 protein in MBH was reported on days 2–3 (p < 0.05), while in POA on days 17–19 and in SME on days 10–12 and 14–16 (p < 0.05). This study demonstrated that adiponectin and adiponectin receptor mRNAs and proteins are present in the porcine hypothalamus and that their expression levels are determined by the pig's endocrine status related to the oestrous cycle.     

Pregnancy‐Associated Glycoprotein (PAG) Profile of Holstein–Friesian Cows as Compared to Dual‐Purpose and Beef Cows

Pregnancy‐associated glycoproteins (PAGs) are produced by mono‐ and binucleate trophoblast cells in the ruminant placenta. PAG appears in maternal blood and, from approximately 4 weeks after fertilization onward, may serve as a reliable means of diagnosing pregnancy. A range of factors are said to affect plasma PAG concentrations, such as number and sex of foetus, mass of calf and placenta, level of milk production and genetic constitution. In this study, PAG pregnancy profiles of a dual‐purpose (Simmental) and two beef breeds (Uckermark and Aubrac) are compared with the profile of the specialized dairy breed Holstein–Friesian. Holstein–Friesian cows were sampled weekly; the levels of the other breeds were presented at 3‐week intervals. The overall significant breed difference (p = 0.013) was founded on deviations during the initial 3 weeks of pregnancy and from 23 weeks onward. During the period critical for the detection of pregnancy, between four and 22 weeks, agreement between PAG levels of various breeds was close (p > 0.05). No significant effect of body mass of cow or calf (relative to mass of dam) was detected. These findings imply that the PAG pregnancy test may be executed uniformly irrespective of breed or type of cow, affirming the suitability of the test as a valuable asset for the cattle industry.     

Effects of Dietary Supplementation of β‐hydroxy‐β‐methylbutyrate on Sow Performance and mRNA Expression of Myogenic Markers in Skeletal Muscle of Neonatal Piglets

The effects of dietary β‐hydroxy‐β‐methylbutyrate (HMB) supplementation during gestation on reproductive performance of sows and the mRNA expression of myogenic markers in skeletal muscle of neonatal pigs were determined. At day 35 of gestation, a total of 20 sows (Landrace × Yorkshire, at third parity) were randomly assigned to two groups, with each group receiving either a basal diet or the same diet supplemented with 4 g/day β‐hydroxy‐β‐methylbutyrate calcium (HMB‐Ca) until parturition. At parturition, the total and live litter size were not markedly different between treatments, however, the sows fed HMB diet had a decreased rate of stillborn piglets compared with the sows fed the control (CON) diets (p < 0.05). In addition, piglets from the sows fed HMB diet tended to have an increased birth weight (p = 0.08), and a reduced rate of low birth weight piglets (p = 0.05) compared with piglets from the CON sows. Nevertheless, lower feed intake during lactation was observed in the sows fed the HMB diet compared with those on the CON diet (p < 0.01). The relative weights of the longissimus dorsi (LD) and semitendinosus (ST) muscle were higher (p < 0.05) in neonatal pigs from the HMB than the CON sows. Furthermore, maternal HMB treatment increased the mRNA levels of the myogenic genes, including muscle regulatory factor‐4 (MRF4, p < 0.05), myogenic differentiation factor (MyoD) and insulin‐like growth factor‐1 (IGF‐1, p < 0.01). In conclusion, dietary HMB supplementation to sows at 4 g/day from day 35 of gestation to term significantly improves pregnancy outcomes and increases the expression of myogenic genes in skeletal muscle of neonatal piglets, but reduces feed intake of sows during lactation.     

Early Foetal Loss Correlates Positively with Seroconversion against Mycobacterium avium paratuberculosis in High‐Producing Dairy Cows

This study was designed to examine (i) the seroprevalence of Mycobacterium avium subs paratuberculosis (MAP) in a high‐producing dairy herd with clinical symptoms of bovine paratuberculosis, (ii) MAP seroconversion and seronegativation dynamics in the herd and (iii) possible relationships between MAP infection status and herd reproductive performance. One single blood test per cow was performed early post‐partum on a monthly basis from day 10–40 post‐partum during the first year of the study in 519 cows belonging to a commercial dairy herd. A subset of 111 cows that became pregnant during the study was tested again 60–200 days later during the early foetal period, immediately after the first confirmation of gestation at 58–64 days post‐AI. Logistic regression analysis indicated no effect of any independent variable on MAP seropositivity and conception rate 28–34 days post‐AI. MAP seropositivity was not a factor affecting the anoestrous, subfertility and early foetal loss rates. In the subset of 111 cows, animals that seroconverted had a 3.9 times greater risk of suffering from early foetal loss (30.3%, 10/33) than the remaining pregnant animals (10.3%, 8/78), (95% confidence interval: 1.11–13.4; p = 0.003). In conclusion, early foetal loss was positively correlated with seroconversion to MAP. Reproductive performance was not impaired by MAP infection.     

Endometrial transcript profile of progesterone‐regulated genes during early pregnancy of Water Buffalo (Bubalus bubalis)

Progesterone (P₄) plays a key role in the establishment and maintenance of pregnancy in most mammals. Unravelling the expression of progesterone‐regulated genes can expand the understanding of the embryonic mortality. Accordingly, we studied the relative mRNA expression of the P₄‐regulated genes in the buffalo. Uteri were collected from the abattoir and categorized into nonpregnant late luteal phase, stage I (28–38th days of gestation) and stage II (48–56th days of gestation) of pregnancy (n = 6/group). After extraction of total RNA from the endometrial tissues, we carried out qRT‐PCR for determining the relative mRNA expression of the P₄‐regulated genes using nonpregnant late luteal phase as calibrator group. The expression of LGALS3BP (essential for maternal recognition of pregnancy) gene was found to be significantly upregulated (p < 0.05), while MUC1 (important for embryo attachment) gene was downregulated in stage I and II of pregnancy. We observed no significant change in the expression of LGALS1, LGALS9 and CTSL genes. The SLC5A11 and SLC2A1 genes (involved in the transport of glucose to endometrium) in early pregnancy were upregulated in the pregnancy stage I (p < 0.05) relative to nonpregnant late luteal phase. The CST3 gene was significantly upregulated in pregnancy stage II (p < 0.01). These results provide molecular insights into the specific pathways involved in foeto‐maternal communication during early pregnancy in buffaloes.