- Genetic‐Quantitative Study of the First‐Service Pregnancy
Probability of Murrah Heifers
- Abstract: Because of the importance of reproduction in stock breeding systems, it is necessary to find selection criteria that increase reproductive efficiency. The aim of this study was to estimate genetic parameters for the probability of conception on first service (PROB) in Murrah heifers, and its association with other traits of economic interest [age at first calving (AFC), service period, calving interval and milk yield at 270 days], with the purpose of evaluating their use as selection criteria. Reproductive information and first lactation records of 1200 Murrah heifers were used to perform two‐trait analyses between PROB and the other characteristics. Bayesian inference was used to estimate the variance components, considering PROB as threshold and the other as linear factors. The results demonstrate that this trait has heritability of 0.15, indicating the possibility of a genetic gain by using it for selection. With respect to the genetic correlation estimates, the only high‐magnitude association was with AFC (−0.899), which is the current criterion indicating sexual precocity of females. In the light of the parameters estimated, the first‐service pregnancy rate is an alternative for indication of sexual precocity, although presenting a smaller genetic gain than the current standard AFC. Nevertheless, additional research should be conducted regarding this trait to assess the economic importance of its use in dairy buffalo production systems.
- The Effect of Oxidative Stress on Thawed Bulk‐Sorted Red Deer Sperm
- Abstract: The aims of this study were to assess the effects of the sex‐sorting process on post‐thaw sperm quality as well as on induced oxidative stress damage (H2O2 0 mm = H000; H2O2 50 mm = H050; H2O2 100 mm = H100) and the protective action of reduced glutathione (GSH) and Trolox, when comparing sorted (BSS) and non‐sorted (NS) red deer spermatozoa incubated at 37°C. Sperm samples from three stags were collected by electroejaculation and frozen. Immediately after thawing, sperm motility was higher (p
- Effects of Interferon‐Tau and Steroids on Cytochrome P450 Activity
in Bovine Endometrial Epithelial Cells
- Authors: CL Gilfeather; CO Lemley
Abstract: The objective of the current study was to examine cyclooxygenase (COX), cytochrome P450 1A (CYP1A) and 2C (CYP2C) activity in bovine endometrial cell cultures following exposure to oxytocin (OT), interferon‐τ (IFN), estradiol (E2) and/or progesterone (P4). Bovine endometrial epithelial cells were treated with OT, IFN, a combination of OT+IFN or control (CON) media for 24 h. For the second experiment, cells were treated with E2, P4, a combination of E2 + P4 or CON media for 24 h. Treatments were performed in triplicate, and the experiment was repeated four times (n = 12 per treatment). Treatment with OT alone increased (p
- Effects of IGF‐1 on In Vitro Culture of Bovine Preantral Follicles
- Abstract: This study aimed at assessing the effect of different concentrations of the growth factor similar to insulin 1 (IGF‐1) in the development, survival and ultrastructure of the bovine preantral follicles cultured in situ. Fragments of bovine ovarian cortical tissue were cultured during 1 and 7 days in 1 ml of α‐MEM+, supplemented with different concentrations of human recombinant IGF‐1 (0, 30, 70 and 100 ng/ml), in an incubator at 37°C and 5% of CO2 in 24‐well plates with total replacement of the medium every 2 days. Non‐cultured ovarian fragments (control) and ovarian fragments cultured during 1 and 7 days were processed for classic histology, mechanical isolation and electron transmission microscopy (ETM). Parameters such as normality, viability, activation, development, diameter and ultrastructure were evaluated. All statistical analyses were carried out using sas Version 9.2. The results showed that the percentage of follicles morphologically normal in the IGF‐1 30 ng/ml treatment was similar to the fresh control (p > 0.05) both on the day 1 and on the day 7 of in vitro culture. In the viability analysis, the cultured treatments maintained the percentage of viable follicles during the entire culture period (p > 0.05). After 7 days of culture, the IGF‐1 30 ng/ml treatment showed higher percentages of developing follicles (48.33%) than those of the fresh control (22.22%) and the cultured treatments (p
- An In Vitro Evaluation of Biochemical Processes Involved in
Lead‐Induced Changes on Ram Spermatozoa
- Abstract: Lead (Pb2+) is a toxic heavy metal which interferes with several physiological processes regulated by Ca2+, including those characterized by changes of the membrane stability and the motility of spermatozoa necessary for the fertilization of the oocyte. In this study, ejaculated sperm from six rams (Ovis aries) have been incubated in vitro with or without 50 ng Pb2+/ml during 30 min and in the presence or absence of three different potential modulators of the effects of Pb2+ on changes in the sperm membrane before fertilization: charybdotoxin, quinacrine and staurosporine. Sperm samples incubated with Pb2+ have shown significant reductions in acrosome integrity and sperm viability and an increase in progressive movement. None of the studied potential modulators had a protective effect against Pb2+ action. On the contrary, Pb2+‐incubated sperm in the presence of staurosporine had lower acrosome integrity, and lower sperm viability was observed when spermatozoa were incubated with Pb2+ + charybdotoxin. Quinacrine was the only tested substance capable of increasing the concentration of Pb2+ in spermatozoa; thus, the enhancement of Pb2+ effects produced by staurosporine and charybdotoxin was not produced by an increased uptake of Pb2+ by spermatozoa. However, the increase of intracellular Pb2+ in those spermatozoa incubated with quinacrine did not result in an adverse effect on sperm motility or viability although the acrosome integrity was negatively affected.
- Effect of Heat Stress on Concentrations of Faecal Cortisol Metabolites in
- Abstract: The negative impact of heat stress on health and productivity of dairy cows is well known. Heat stress can be quantified with the temperature–humidity index (THI) and is defined as a THI ≥ 72. Additionally, animal welfare is affected in cows living under heat stress conditions. Finding a way to quantify heat stress in dairy cows has been of increasing interest over the past decades. Therefore, the objective of this study was to evaluate concentrations of faecal glucocorticoid metabolites [i.e. 11,17‐dioxoandrostanes (11,17‐DOA)] as an indirect stress parameter in dairy cows without heat stress (DOA 0), with heat stress on a single day (acute heat stress, DOA 1) or with more than a single day of heat stress (chronic heat stress, DOA 2). Cows were housed in five farms under moderate European climates. Two statistical approaches (approach 1 and approach 2) were assessed. Using approach 1, concentrations of faecal 11,17‐DOA were compared among DOA 0, DOA 1 and DOA 2 samples regardless of their origin (i.e. cow, unpaired comparison with a one‐way anova). Using approach 2, a cow was considered as its own control; that is 11,17‐DOA was treated as a cow‐specific factor and only paired samples were included in the analysis for this approach (paired comparison with t‐tests). In approach 1 (p = 0.006) and approach 2 (p = 0.038), 11,17‐DOA values of cows under acute heat stress were higher compared to those of cows without heat stress. Our results also indicate that acute heat stress has to be considered as a confounder in studies measuring faecal glucocorticoid metabolites in cows to evaluate other stressful situations.
- Apoptotic Cell Localization in Preantral and Antral Follicles in Relation
to Non‐cyclic and Cyclic Gilts
- Authors: D Phoophitphong; S Srisuwatanasagul, S Koonjaenak, P Tummaruk
Abstract: The objective of this study was to determine apoptotic cell localization in preantral and antral follicles of porcine ovaries. Additionally, the proportion of cells undergoing apoptosis was also compared between delayed puberty gilts and normal cyclic gilts. Ovarian tissues were obtained from 34 culled gilts with age and weight of 270.1 ± 3.9 days and 143.8 ± 2.4 kg, respectively. The gilts were classified according to their ovarian appearance as ‘non‐cyclic’ (n = 7) and ‘cyclic’ (n = 27) gilts. The terminal deoxynucleotidyl transferase‐mediated dUTP nick end labelling (TUNEL) assay was used to determine apoptotic cell expression in different compartments of the ovarian tissue sections. All apparent preantral (n = 110) and antral (n = 262) follicles were evaluated using image analysis software. It was found that apoptotic cells were expressed in both granulosa (22.2%) and theca cell layers (21.3%) of the follicles in the porcine ovaries. The proportion of apoptotic cells in the granulosa layer in the follicles was positively correlated with that in the theca layer (r = 0.90, p 0.05) or theca cell layers (28.6% and 26.5%, p > 0.05). The proportion of apoptotic cells in non‐cyclic gilts was higher than cyclic gilts in both granulosa (31.7% and 22.6%, p
- Effect of Follicle Size on In Vitro Maturation of Pre‐Pubertal
Porcine Cumulus Oocyte Complexes
- Abstract: Very small follicles (
- Pre‐Selection Test to Identify High Responder Donor Goats
- Abstract: The aim of the study was to evaluate the feasibility of pre‐selection of high or low responder does prior to the superovulatory protocols. Twenty Saanen does received 800 IU of equine chorionic gonadotropin (eCG) at the end of long‐term progestogen treatment. Fourteen days later, a second progestogen protocol associated with a multiple‐dose follicle stimulation hormone (FSH) treatment (5 IU/kg of FSH, in six decreasing doses between days 4 to 6 of the protocol) was administered. Transrectal ultrasound was used to assess the follicular status at the beginning of superovulatory treatments, at the oestrous onset and on the seventh day of the oestrous cycle for counting corpora lutea (CL). A significant lower number of CL was obtained in eCG‐treated in comparision with FSH‐treated does (p
- Mitochondrial DNA Copy Number in Spermatozoa of Fertile Stallions
- Abstract: Predicting male fertility on non‐invasive sperm traits is of big importance to human and animal reproduction strategies. Combining the wide range of parameters monitored by computer‐assisted sperm analysis (CASA) with some molecular traits (e.g. mtDNA content) may help to identify markers of the male fertility. The aim of this study was to characterize variation in the mtDNA copy number in equine sperm and to investigate whether mtDNA content is correlated with quality traits of stallion spermatozoa and the age of the male. Ejaculates collected from 53 fertile stallions were divided into four age groups (3–5, 6–10, 11–14 and >15 years) and were subjected to a complex investigation including conventional analysis, CASA, flow cytometry and mtDNA content (real‐time PCR). The mean (±SD) number of mtDNA copies equalled 14 ± 9 and varied from 3 to 64. Considering the great number of sperm parameters monitored in this study, only few of them were correlated with the mtDNA content: ejaculate volume (a positive correlation), the amplitude of lateral head displacement (ALH; a negative correlation) and the high mitochondrial activity index (a negative correlation). The stallion age was not correlated with the mtDNA copy number. This study provides the first set of data on mtDNA content in equine sperm and confirms phenomena previously described for humans and dog on associations between sperm mtDNA content and selected motility parameters monitored by the CASA. Basing our study on spermatozoa from fertile stallions could however limit the extent of detected associations.
- Morphology and Aquaporin Immunohistochemistry of the Uterine Tube of
- Authors: S Arrighi; G Bosi, S Frattini, B Croizet, D Groppetti, A Pecile
Abstract: The expression of six different aquaporins (AQP1, 2, 3, 4, 5 and 9), integral membrane water channels that facilitate bi‐directional passive movement of water, was investigated by immunohistochemistry in the uterine tube of pre‐pubertal and adult Saanen goats (Capra hircus), comparing the different phases of the oestrous cycle. Regional morphology and secretory processes were markedly different during the goat oestrous cycle. The tested AQP molecules showed different expression patterns in comparison with already studied species. AQP1‐immunoreactivity was evidenced at the endothelium of blood vessels and in nerve fibres, regardless of the tubal tract and cycle period. AQP4‐immunoreactivity was shown on the lateral plasmalemma in the basal third of the epithelial cells at infundibulum and ampulla level in the cycling goats, more evidently during follicular than during luteal phase. No AQP4‐immunoreactivity was noticed at the level of the isthmus region, regardless of the cycle phase. AQP5‐immunoreactivity, localized at the apical surface of epithelial cells, increased from pre‐puberty to adulthood. Thereafter, AQP5‐immunoreactivity was prominent during the follicular phase, when it strongly decorated the apical plasmalemma of all epithelial cells at ampullary level. During luteal phase, immunoreactivity was discontinuous, being weak to strong at the apex of the secretory cells protruding into the lumen. In the isthmus region, the strongest AQP5‐immunoreactivity was seen during follicular phase, with a clear localization in the apical plasmalemma of all the epithelial cells and also on the lateral plasmalemma. AQP2, 3 and 9 were undetectable all along the goat uterine tube. Likely, a collaboration of different AQP molecules sustains the fluid production in the goat uterine tube. AQP1‐mediated transudation from the blood capillaries, together with permeation of the epithelium by AQP4 in the basal rim of the epithelial cells and final intervening of apical AQP5, could be involved in fluid production as well as in secretory processes.
- Fertility Assessment in Sorraia Stallions by Sperm‐Fish and Fkbp6
- Abstract: The Sorraia, a critically endangered indigenous Iberian horse breed, is characterized by low genetic variability, high rate of inbreeding, bad sperm quality and subfertility. Here, we studied 11 phenotypically normal but subfertile Sorraia stallions by karyotyping, sex chromosome sperm‐FISH and molecular analysis of FKBP6 – a susceptibility locus for impaired acrosome reaction (IAR). The stallions had normal sperm concentration (>300 million cells/ml), but the numbers of progressively motile sperm (21%) and morphologically normal sperm (28%) were invariably low. All stallions had a normal 64,XY karyotype. The majority of sperm (89%) had normal haploid sex chromosome content, although 11% of sperm carried various sex chromosome aneuploidies. No correlation was found between the percentage of sperm sex chromosome abnormalities and inbreeding, sperm morphology or stallion age. Direct sequencing of FKBP6 exon 4 for SNPs g.11040315G>A and g.11040379C>A revealed that none of the stallions had the susceptibility genotype (A/A‐A/A) for IAR. Instead, all animals had a G/G‐A/A genotype – a testimony of low genetic variability. The findings ruled out chromosomal abnormalities and genetic predisposition for IAR as contributing factors for subfertility. However, low fertility of the Sorraia stallions could be partly attributed to relatively higher rate of sex chromosome aneuploidies in the sperm.
- Serum Levels of Cardiac Markers NT‐proANP and NT‐proBNP in
Brachycephalic bitches at Different Gestational Stages
- Authors: MAR Feliciano; RR Uscategui, GS Maciel, VT Almeida, MF Silveira, GAC Oliveira, WRR Vicente
Abstract: The aim of this study was to determine serum levels of natriuretic peptide precursors (NT‐proANP and NT‐proBNP) during pregnancy in brachycephalic bitches. Fifteen healthy multiparous bitches were selected for this prospective study. Serum levels of NT‐proANP and NT‐proBNP were measured during anoestrous and at 14, 35, 42, 49 and 56 days (2nd, 5th, 6th, 7th and 8th weeks) of pregnancy. Fourteen animals had normal gestations, and one bitch developed single foetus syndrome. The natriuretic peptide levels of this animal were not included in this study; however, it is important to report that its NT‐proANP levels were four times greater than those of normal patients. There was no significant difference (p = 0.072) in NT‐proBNP levels between anoestrous (0.20 ± 0.10 ng/ml) and the different pregnancy weeks (0.27 ± 0.12 ng/ml). There was a positive correlation (p
- Total Cell Number and its Allocation to Trophectoderm and Inner Cell Mass
in In Vitro Obtained Cats' Blastocysts
- Abstract: The aim of this study was to evaluate the developmental kinetics of cats' blastocysts in connection with their morphology and blastomeres allocation to trophoblast or embryoblast cells. We examined gross blastocyst morphology and the total number of blastomeres together with its allocation to inner cell mass (ICM) or trophectoderm (TE) cells in pre‐implantation feline embryos obtained from 6th to 9th day of in vitro culture. From all the investigated embryos, 61.8% developed to early blastocyst, 37.4% to full and 7.6% to hatching blastocyst stage. The total cell number (TCN) varied form 58 cells in early day 6 to 245 in hatching day 8 blastocyst, with the mean 84.9 cells in early, 156.7 in full, and 204.4 in hatching ones. Day 8 blastocyst had the highest number of total cells, together with the highest mean number of ICM regardless of its morphological assessment. Early blastocyst (apart from day 6) had the highest number of arrested cells, while dead cells were the highest in full day 9 blastocyst. More data about the relationship between blastocyst development and morphology would facilitate the selection of optimal blastocysts for further procedures.
- Cryopreservation of Dog Semen in a Tris Extender with 1% or 2% Soya Bean
Lecithin as a Replacement of Egg Yolk
- Abstract: Egg yolk is usually included in extenders used for preservation of dog semen. Lecithin is an interesting animal‐protein free alternative to egg yolk for semen preservation. The aim of our study was to evaluate soya bean lecithin for cryopreservation of dog semen. Five ejaculate replicates were divided in three equal parts, centrifuged and each pellet diluted with one of the three Tris‐based extenders containing 20% egg yolk, 1% soya bean lecithin or 2% soya bean lecithin. Extended semen was loaded in 0.5‐ml straws, cooled and diluted a second time and frozen in liquid nitrogen vapours. Sperm motility parameters (CASA), acrosome integrity (FITC‐PNA/PI) and sperm membrane integrity (C‐FDA) were evaluated 5 min post‐thaw and after 2 and 4 h of incubation. Total motility was significantly better in the egg yolk extender than in any of the lecithin‐based extender and was better in the 1% lecithin extender than in the 2% lecithin extender. Sperm membrane integrity was significantly better in the egg yolk extender than in any of the lecithin‐based extenders but did not differ significantly between the 1% and 2% lecithin extenders. Acrosome integrity was significantly better in the egg yolk extender than in the 2% lecithin extender but did not differ between the egg yolk extender and the 1% lecithin extender or between the two lecithin extenders. In conclusion, egg yolk was superior to lecithin in our study. The extender with 1% lecithin preserved sperm motility better than the extender with 2% lecithin.
- The Anti‐Müllerian Hormone Profile is Linked with the In Vitro
Embryo Production Capacity and Embryo Viability after Transfer but Cannot
Predict Pregnancy Outcome
- Authors: N Ghanem; JI Jin, SS Kim, BH Choi, KL Lee, AN Ha, SH Song, IK Kong
Abstract: The current study investigated the possibility of using the AMH concentration as a predictor of the ability of Korean Hanwoo cows to produce cumulus‐oocyte complexes, embryos that survive after transfer as well as the pregnancy outcome of surrogates. Eight sessions of ovum pick‐up (OPU) were performed with 19 donor cows at an interval of 3–4 days. Antral follicle count (AFC), oocyte quality and in vitro embryo development were recorded for each cow. Embryos produced from cows with different AMH profiles were transferred into recipients (n = 96). Cows in the high (≥0.25 ng/ml) and intermediate (0.1≥ to
- Effect of Bovine Sperm‐Bound Antisperm Antibodies on Oviductal
- Authors: MS Ferrer; DE Anderson, LMJ Miller, A George, M Miesner, M Wilkerson
Abstract: This study was designed to test the hypothesis that sperm‐bound IgG and IgA decrease binding of bull spermatozoa to oviductal epithelial cells in vitro. Three ejaculates were cryopreserved from each of four antisperm antibody (ASA)‐negative satisfactory breeder bulls. Bulls were then immunized with autologous spermatozoa, and three ASA‐positive ejaculates were cryopreserved from each bull post‐immunization. First, microscopy methods were compared to select the most appropriate assay for evaluation of oviductal binding index (BI). The BI did not differ when the evaluation was performed under fluorescence microscopy (131.1 sperm/mm2; 62.5–251.1 sperm/mm2), phase‐contrast microscopy (160.5 sperm/mm2; 56.8–397.4 mm2) or their combination (116.4 sperm/mm2; 56.8–249.6 sperm/mm2) (Median; IQR). The combination of microscopy methods was selected as it allowed better visualization of cells. Then, BI was compared between ASA‐negative and ASA‐positive ejaculates, and the association between BI and ASA binding was evaluated. The BI was less in ASA‐positive (114.9; 0 to 201.8 sperm/0.1 mm2) than ASA‐negative samples (218.9; 24.7 to 276.8 sperm/0.1 mm2) (P = 0.0002). This reduction in BI was significant in three of the four bulls. Regression analysis identified a negative association between BI and the percentage of IgG‐bound (p = 0.013) but not IgA‐bound spermatozoa. In conclusion, sperm‐bound IgG decreased the ability of bovine spermatozoa to bind to oviductal epithelial cells in vitro.
- Experimental Neospora Caninum Infection in Pregnant Dairy Heifers Raises
Concentrations of Pregnancy‐Associated Glycoproteins 1 and 2 in
- Abstract: 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
- l‐carnitine Mediated Reduction in Oxidative Stress and Alteration in
Transcript Level of Antioxidant Enzymes in Sheep Embryos Produced In Vitro
- Authors: A Mishra; IJ Reddy, PSP Gupta, S Mondal
Abstract: The objective of this study was to find out the effect of l‐carnitine on oocyte maturation and subsequent embryo development, with l‐carnitine‐mediated alteration if any in transcript level of antioxidant enzymes (GPx, Cu/Zn‐SOD (SOD1) and Mn‐SOD (SOD2) in oocytes and developing sheep embryos produced in vitro. Different concentrations of l‐carnitine (0 mm, 2.5 mm, 5 mm, 7.5 mm and 10 mm) were used in maturation medium. Oocytes matured with 10 mm l‐carnitine showed significantly (p
- Can the Genetic Origin Affect Rabbit Seminal Plasma Protein Profile along
- Abstract: The study was designed to evaluate the influence of genetic origin on rabbit seminal plasma protein profile variation along the year. Seminal plasma of rabbits from line A (maternal line) and R (paternal line) collected during a natural year was subjected to polyacrylamide gel electrophoresis (SDS‐PAGE). The electrophoretic profile of rabbit seminal plasma resulted in multiple protein bands of different intensity ranging from 9 to 240 kDa. Results showed that seven protein bands were significantly different between genetic lines, and among these, three protein bands were significantly different between seasons. The differentially expressed proteins were identified by MALDI‐TOF/TOF or LC‐MS/MS analysis and were the following ones: FAM115E‐like (220, 113 and 59 kDa), ectonucleoside triphosphate diphosphohydrolase 3 isoform X2 (72 kDa), annexin A5 (32 kDa), lipocalin allergen Ory c 4 precursor (19 kDa), and haemoglobin subunit zetalike (13 kDa) between genetic lines and FAM115E‐like (113 kDa), haemoglobin subunit zetalike (13 kDa) and β‐nerve growth factor (12 kDa) between seasons. These results indicate that proteins from rabbit seminal plasma are under both seasonal control and genetic control. Furthermore, the differential presence of these proteins could be one of the causes explaining the differences observed in fertility and seminal parameters between these two lines in earlier studies.
- Follicle Diameter and Systemic Hormone Interrelationships during Induction
of Follicle Collapse with Intrafollicular Prostaglandin E2 and F2α in
- Abstract: The objectives were to determine: (i) whether intrafollicular administration of PGE2 and PGF2α to mares would hasten follicle collapse and (ii) the differences in reproductive hormone characteristics in mares with spontaneous and prostaglandin‐induced follicle collapses. Six mares were followed for two oestrous cycles each: when the mares reached a follicle diameter of 30–35 mm and showed mild‐to‐moderate endometrial oedema, mares were administered a single 0.5 ml dose containing 500 μg PGE2 and 125 μg PGF2α (treatment cycle) or a placebo (0.5 ml of water for injection; control cycle) into the preovulatory follicle (Hour 0). Blood samples were collected, and serial ultrasound examinations were performed until follicle collapse. Treated mares showed follicle collapse significantly earlier (20.0 ± 5.9 h) than the control mares (72.0 ± 10.7 h). The LH, progesterone, total oestrogens and oestradiol concentrations did not differ between groups; however, the progesterone concentration increased more between 48 and 72 h after follicle injection in the treatment compared to the control cycles (P
- Profile of Steroid Receptors and Increased Aromatase Immunoexpression in
Canine Inflammatory Mammary Cancer as a Potential Therapeutic Target
- Abstract: Canine inflammatory mammary cancer (IMC) has been proposed as a model for the study of human inflammatory breast cancer (IBC). The aims of this study were to compare the immunohistochemical expression of aromatase (Arom) and several hormone receptors [estrogen receptor α (ERα), estrogen receptor β (ERβ), progesterone receptor (PR) and androgen receptor (AR)], in 21 IMC cases vs 19 non‐IMC; and to study the possible effect of letrozole on canine IMC and human inflammatory breast cancer (IBC) in vitro using IPC‐366 and SUM‐149 cell lines. Significant elevations of the means of Arom Total Score (TS), ERβ TS and PR TS were found in the IMC group (p = 0.025, p = 0.038 and p = 0.037, respectively). Secondary IMC tumours expressed higher levels of Arom than primary IMC (p = 0.029). Non‐IMC PR‐ tumours contained higher levels of Arom than non‐IMC PR+ tumours (p = 0.007). After the addition of letrozole, the number of IMC and IBC cells dropped drastically. The overexpression of Arom found and the results obtained in vitro further support canine IMC as a model for the study of IBC and future approaches to the treatment of dogs with mammary cancer, and especially IMC, using Arom inhibitors.
- In vitro Developmental Competence of Adult Sheep Oocytes Treated with
- Authors: LF Crocomo; F Ariu, L Bogliolo, D Bebbere, S Ledda, SD Bicudo
Abstract: The efficiency of in vitro sheep embryo production is still low compared to that observed in vivo and in other species. In this context, meiotic inhibition strategies emerged as a promising alternative to improve this biotechnology. So, this study aimed to evaluate, for the first time, the effects of roscovitine on in vitro maturation of sheep oocytes and their subsequent embryo development. For this, cumulus–oocyte complexes (COCs) were cultured for 6 h in the presence (Rosco) or absence (Control) of 75 μm roscovitine and, subsequently, in vitro matured (IVM) for 18 h with gonadotropins. At 0 (Immature), 6 and 24 h of culture, the nuclear status of oocytes was evaluated by Hoechst staining. Embryo cleavage and blastocyst formation were recorded 30 h after in vitro fertilization and on day 7 of culture, respectively. Blastocyst quality was evaluated by differential staining. At 6 h, the GV rate in the Rosco treatment (93.8%) was similar to that observed in the Immature oocytes (94.9%) and significantly higher compared to Control (41.3%). After IVM for 18 h, a high and similar proportion of oocytes from Rosco (93.6%) and Control (88.4%) reached the MII stage. In both treatments, approximately 70% of oocytes cleaved and 50% of them developed up to blastocyst. The mean percentage of blastocyst cells, embryoblast, trophoblast and pyknosis did also not differ between Control and Rosco. In conclusion, roscovitine, at the studied experimental conditions, was efficient to reversibly inhibit the meiosis of adult sheep oocytes without detrimental effect on development and quality of the in vitro produced embryos.
- Effects of Supplementing Holstein Heifers with Dietary Melatonin during
Late Gestation on Growth and Cardiovascular Measurements of their
- Authors: KE Brockus; CG Hart, BO Fleming, T Smith, SH Ward, CO Lemley
Abstract: The objective was to examine the effects of supplementing dams with dietary melatonin during late gestation on offspring growth and cardiovascular measurements. On day 190 of gestation, heifers (n = 20) were blocked by body weight and randomly assigned to one of two dietary treatments consisting of 20 mg of dietary melatonin per day [melatonin (MEL)] or no melatonin supplementation [control (CON)]. Dietary treatments were terminated on day 262 of gestation. At birth, calves were separated from their dams with no further treatments. Calf (n = 18) blood pressure, cortisol, nitrites and total antioxidant capacity were collected on weeks 0, 1, 2, 3 and 4 of age. Calf hepatic portal blood flow and concentrations of insulin‐like growth factor 1 were determined on weeks 0 and 4 of age. Calf body weight, abdominal girth, hip height and wither height increased (p
- Successful Vitrification of In vivo Embryos Collected from Superovulated
Japanese Black Cattle (Wagyu)
- Authors: L An; PP Ling, X Zhu, Y Liu, F Zhang, X Ma, B Xu, Y Wang, Z Du, L Yang, F Xue, A Bella, GA Presicce, F Du
Abstract: The aim of this study was to determine whether vitrification is an effective method when used for Japanese Black Cattle (Wagyu) in vivo‐derived embryos, collected following a superovulation treatment and embryo transfer (MOET) programme. In vivo‐derived morula and blastocysts collected on day 7 after artificial insemination, were vitrified using a modified droplet vitrification (MDV) procedure and subsequently warmed for transfer (ET) into synchronized recipients. Fresh embryos, and embryos cryopreserved using a standardized slow freezing procedure (direct thaw/direct transfer, DT) served as ET controls. Two different follicle‐stimulating hormone (FSH) sources, Folltropin® Canada (FSH BAH, 24 donors) and a brand prepared by the Chinese Academy of Science (FSH CAS, 16 donors), were compared in a series of superovulation outcomes following well‐established FSH administration protocols. Following data analysis, the total number of ovulations recorded at the time of embryo flushing (10.5 vs 8.5; p = 0.28) and the total number of transferable embryos (6.2 vs 5.1; p = 0.52) were similar between the two FSH sources. ET for MDV (39.7%, n = 78), DT (35.2%, n = 71) and fresh controls (47.1%, n = 34) resulted in similar pregnancy rates (p > 0.05). When MDV was used, a higher pregnancy rate (42.6%) resulted from the transfer of vitrified morulae, when compared to the DT counterparts (24.3%), (p = 0.05). Transfer of vitrified morulae resulted also in higher pregnancy rate, when compared to the transfer of vitrified blastocysts (42.6% vs. 29.4%; p
- Characteristics of Antioxidant Systems of Yellow Fraction of Red Deer's
(Cervus elaphus L.) Semen During the Rutting Period
- Abstract: The objective of this study was to make the preliminary characterization of the antioxidant defence systems of the yellow fraction (YF) of red deer's (Cervus elaphus L.) semen during the rutting period. The semen was collected using artificial vagina (AV). The studies included spectrophotometric determination of antioxidant enzymes activities such as superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx). We also analysed the contents of low‐molecular antioxidants such as L‐glutathione (GSH + GSSG), L‐ascorbate (ASC) and total antioxidant status (TAS). Additionally, the samples were subjected to PAGE and stained for SOD and GPx activities. It was demonstrated that the yellow fraction exhibited activities of SOD and GPx, with the highest activities in September and October. CAT activity was not detected. Staining for the SOD and GPx activities confirmed three protein bands with SOD activity and one protein band with GPx activity. The content of GSH + GSSG was similar in trials dating from October to December contrary to the content of ASC which was high in samples from September and October. The stable rate of TAS was observed during the whole rutting period. The results of this study showed that the YF of red deer semen is equipped with basic battery of antioxidant enzymes comprising SOD and GPx, with the supporting role of GSH + GSSG and ASC. Moreover, the samples obtained at the peak of the rutting period occurring from September to October had the highest enzymatic activity in comparison with remaining months of the rutting period, which contributed to the high quality of the semen by preventing it from the formation of oxidative stress during the short period of intense sexual activity of male red deer. The better understanding of the mechanisms of antioxidant defence systems in the YF of deer's semen may contribute to the potential use of this fraction in technology of wild ruminant semen preservation.
- Seminal Plasma Characteristics and Expression of ATP‐binding
Cassette Transporter A1 (ABCA1) in Canine Spermatozoa from Ejaculates with
Good and Bad Freezability
- Abstract: The composition of seminal plasma and the localization of the ATP‐binding cassette transporter A1 (ABCA1) in spermatozoa from good and bad freezers were compared to frozen–thawed spermatozoa from the same dog. Ejaculates were obtained from 31 stud dogs, and the sperm‐rich fraction (SRF) was kept for analysis. One aliquot was used for the analysis of concentration, progressive motility (P; CASA), viability (V; CASA) and leucocyte count, and the analysis was performed by flow cytometry (FITC‐PNA/PI), SCSA and HOST. In seminal plasma, concentration of albumin, cholesterol, calcium, inorganic phosphate, sodium, potassium, zinc and copper was measured. Semen smears were prepared and evaluated for the expression of ABCA1. The remainder of each ejaculate was frozen. After thawing, the quality assessment was repeated and further smears were prepared. According to post‐thaw semen quality, dogs were assigned to good freezers (n = 20) or bad freezers (n = 11), the latter were defined as 40% morphologically abnormal sperm and/or
- Early Foetal Loss Correlates Positively with Seroconversion against
Mycobacterium avium paratuberculosis in High‐Producing Dairy Cows
- Abstract: 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.
- Immunolocalization of the Anti‐Müllerian Hormone (AMH) in
Caprine Follicles and the Effects of AMH on In Vitro Culture of Caprine
Pre‐antral Follicles Enclosed in Ovarian Tissue
- Abstract: The aims of this study were to evaluate the localization, by immunohistochemistry, of the anti‐Müllerian hormone (AMH) in goat ovaries and to investigate its effects on the in vitro survival and development of caprine pre‐antral follicles enclosed in fragments of ovarian tissue. Pre‐antral follicles were cultured in vitro for 1 or 7 days in α‐MEM+ in the absence or presence of kit ligand (KL; 50 ng/ml, positive control) or AMH (50 or 150 ng/ml). The results showed that AMH was localized in oocytes and granulosa cells from the primordial follicle to antral follicle stages. Addition of AMH maintained the percentage of developing follicles, similar to that in the uncultured control; however, the percentage of developing follicles was significantly lower than that in the cultured control and KL. Nonetheless, addition of AMH to the culture medium did not affect survival rates and follicular growth. In conclusion, this study demonstrated that the expression of AMH varies according to the compartment and stage of follicular development. Furthermore, AMH inhibits the activation of caprine primordial follicles.
- Cilostazol Improves Developmental Competence of Pig Oocytes by Increasing
Intraoocyte Cyclic Adenosine Monophosphate Level and Delaying Meiotic
- Abstract: Cilostazol (CLZ) is a cyclic adenosine monophosphate (cAMP) modulator that influences the steady state of the meiotic stage. This study was conducted to determine the effects of CLZ treatment during in vitro maturation (IVM) on developmental competence of pig oocytes. Immature oocytes were exposed to 0 (control), 0.5, 2 and 4 μm CLZ during the first 22 h of IVM. Nuclear maturation, intraoocyte glutathione content and embryo cleavage after parthenogenesis (PA) and somatic cell nuclear transfer (SCNT) were not influenced by CLZ at any concentrations. However, 4 μm CLZ significantly (p
- Campylorrhinus lateralis, Bilateral microphthalmia and odontoma temporalis
in an Oldenburg Foal
- Abstract: An Oldenburg colt with wry nose was autopsied after having lived for only 30 min. It presented cyanotic oral mucosae, underdeveloped eyes and a right‐sided temporal osseous mass. The applicable nomenclature for the defects is discussed, and the potential etiopathogenesis is explored by describing the normal embryonic development of the affected body parts.
- Successful Cryopreservation of Domestic Cat (Felis catus) Epididymal Sperm
after Slow Equilibration to 15 or 10°C
- Abstract: To support conservation strategies in wild species, simple but highly reproducible procedures of sperm cryopreservation are required for an application under field conditions. We used epididymal sperm of the domestic cat to optimize a sperm freezing procedure for felid species, particularly questioning the demand for sperm cooling to 4°C. We equilibrated sperm during slow cooling to only 15 or 10°C in a Tes–Tris–fructose extender with final concentrations of 4.7% (v/v) glycerol and 10% (v/v) of the water‐soluble fraction of hen's egg yolk (low‐density lipoproteins). Subsequently, sperm were frozen over liquid nitrogen. Total and progressive motility (mean ± SD) after thawing was 60.7 ± 8.6% and 53.9 ± 9.6% in samples cooled to 15°C or 61.6 ± 9.5% and 55.3 ± 9.9% in samples cooled to 10°C. Therefore, a one‐step addition of glycerol to sperm at room temperature together with the freezing extender, the use of cryovials (loaded with diluted sperm aliquots of 300 μl), an equilibration period of 40 min comprising slow cooling to 15°C at a rate of approximately −0.14 K/min before rapid freezing over liquid nitrogen, yielded satisfying results. Cooling, freezing and thawing rates were exactly characterized as a prerequisite for further optimization and to provide a repeatable protocol to other practitioners.
- Effect of Temporary Meiotic Attenuation of Oocytes with Butyrolactone I
and Roscovitine in Resistance to Bovine Embryos on Vitrification
- Abstract: This study aimed to produce in vitro bovine embryos by the addition of two drugs, which is responsible for oocyte meiosis inhibition: roscovitine (ROS) and butyrolactone I (BL‐I). Oocytes were recovered from slaughtered cows and matured in a commercial medium and maintained in a 5% CO2 atmosphere. Oocytes were maintained for 6 h in an in vitro maturation (IVM) medium containing ROS (12.5 μm), BL‐I (50 μm) and association of drugs (ROS 6.25 μm and BL‐I 25 μm). Oocytes were cultured for 18 h in an agent‐free medium for the resumption of meiosis. After 24 h of maturation, oocytes were inseminated in the commercial in vitro fertilization (IVF) medium. Presumptive zygotes were cultured in SOFaa medium in a 5% CO2 atmosphere. On day 3, rate of cleavage was evaluated and on days 6 and 7, rate of blastocyst formation. BL‐I and its association with the ROS increased the rates of cleavage and blastocyst formation (p
- Santa Maria, Brazil: Regulation of Anti‐Müllerian Hormone and
Its Receptor Expression around Follicle Deviation in Cattle
- Abstract: The anti‐Müllerian hormone (AMH) is an important marker of ovarian reserve and for predicting the response to superovulatory treatments in several species. The objective of this study was to investigate whether AMH and its receptor (AMHR2) are regulated in bovine granulosa cells during follicular development. In the first experiment, granulosa cells were retrieved from the two largest follicles on days 2 (before), 3 (at the expected time) or 4 (after deviation) of follicular wave. In the second experiment, four doses of FSH (30, 30, 20 and 20 mg) or saline were administered twice a day starting on Day 2 of the first follicular wave of the cycle. Granulosa cells and follicular fluid were collected from the two largest follicles 12 h after the last injection of FSH or saline. AMH mRNA abundance was similar in granulosa cells of the two largest follicles (F1 and F2) before deviation (Day 2), but greater in dominant (DF) than subordinate follicles (SF) at the expected time (Day 3) and after (Day 4) deviation (p 0.05), but they tended to be greater in DFs than SFs (p 0.05) between both co‐dominant follicles collected from the FSH‐treated cows. These findings indicate the followings: AMH mRNA levels decrease in both DFs and SFs during follicular deviation; granulosa cells from heathy follicles express more AMH mRNA compared to subordinate follicles undergoing atresia and FSH stimulates AMH and AMHR2 mRNA expression in granulosa cells of co‐dominant follicles.
- Key Factors Affecting Reproductive Success of Thoroughbred Mares and
Stallions on a Commercial Stud Farm
- Authors: EA Lane; MLJ Bijnen, M Osborne, SJ More, ISF Henderson, P Duffy, MA Crowe
Abstract: To evaluate factors contributing to fertility of thoroughbred mares, data from 3743 oestrous periods of 2385 mares were collected on a large thoroughbred farm in Ireland. Fourteen stallions (mean age 8.3 years; range 4–15 years) had bred 2385 mares (mean age 9.4 years; range 3–24 years). Maiden mares accounted for 12%, mares with a foal at foot for 64%, and barren, slipped or rested mares for 24% of the total. The mean pregnancy rate per cycle was 67.8% (68.6% in year 1 and 66.9% in year 2). Backward stepwise multivariable logistic regression analysis was utilized to develop two models to evaluate mare factors, including mare age, reproductive status, month of foaling, dystocia, month of cover, foal heat, cycle number, treatments, walk‐in status and stallion factors including stallion identity, stallion age, shuttle status, time elapsed between covers and high stallion usage on the per cycle pregnancy rate and pregnancy loss. Old age (p
- Issue Information
- Pages: 179 - 180
- Letter to the Editor
- Authors: Firdous Ahmad Khan
Pages: 335 - 335
- Authors: Mats HT Troedsson
Pages: 336 - 336