Browsing by Subject "ART"

Now showing 1 - 6 of 6
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    Effect of Superovulation Treatment on Oocyte's DNA Methylation
    (MDPI, 2022-12-18) S. Lopes, Jordana; Ivanova, Elena; Ruiz, Salvador; Andrews, Simon; Kelsey, Gavin; Coy, Pilar; Didáctica y Organización Escolar
    Controlled ovarian stimulation is a necessary step in some assisted reproductive procedures allowing a higher collection of female gametes. However, consequences of this stimulation for the gamete or the offspring have been shown in several mammals. Most studies used comparisons between oocytes from different donors, which may contribute to different responses. In this work, we use the bovine model in which each animal serves as its own control. DNA methylation profiles were obtained by single-cell whole-genome bisulfite sequencing of oocytes from pre-ovulatory unstimulated follicles compared to oocytes from stimulated follicles. Results show that the global percentage of methylation was similar between groups, but the percentage of methylation was lower for non-stimulated oocytes in the imprinted genes APEG3, MEG3, and MEG9 and higher in TSSC4 when compared to stimulated oocytes. Differences were also found in CGI of imprinted genes: higher methylation was found among non-stimulated oocytes in MEST (PEG1), IGF2R, GNAS (SCG6), KvDMR1 ICR UMD, and IGF2. In another region around IGF2, the methylation percentage was lower for non-stimulated oocytes when compared to stimulated oocytes. Data drawn from this study might help to understand the molecular reasons for the appearance of certain syndromes in assisted reproductive technologies-derived offspring.
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    Open Access
    Effect of superovulation treatment on oocyte´s DNA methylation.
    (MDPI, 2022-12-18) Lopes, J.S.; Ivanova, E.; Ruiz, S.; Andrews, S.; Kelsey, G.; Coy, P.; Fisiología; Epigenetics Programme. Babraham Institute. Cambridge, U.K.; Bioinformatics Group. Babraham Institute. Cambridge, U.K.; Centre for Trophoblast Research. Cambridge University. U.K.
    Controlled ovarian stimulation is a necessary step in some assisted reproductive procedures allowing a higher collection of female gametes. However, consequences of this stimulation for the gamete or the offspring have been shown in several mammals. Most studies used comparisons between oocytes from different donors, which may contribute to different responses. In this work, we use the bovine model in which each animal serves as its own control. DNA methylation profiles were obtained by single-cell whole-genome bisulfite sequencing of oocytes from pre-ovulatory unstimulated follicles compared to oocytes from stimulated follicles. Results show that the global percentage of methylation was similar between groups, but the percentage of methylation was lower for non-stimulated oocytes in the imprinted genes APEG3, MEG3, and MEG9 and higher in TSSC4 when compared to stimulated oocytes. Differences were also found in CGI of imprinted genes: higher methylation was found among non-stimulated oocytes in MEST (PEG1), IGF2R, GNAS (SCG6), KvDMR1 ICR UMD, and IGF2. In another region around IGF2, the methylation percentage was lower for non-stimulated oocytes when compared to stimulated oocytes. Data drawn from this study might help to understand the molecular reasons for the appearance of certain syndromes in assisted reproductive technologies-derived offspring.
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    Open Access
    Long-term biochemical and haematological analysis, with focus on sex effects, in pigs derived from assisted reproductive technologies
    (2025) Mikita, Małgorzata; Romero Aguirregomezcorta, Jon; Cánovas, Sebastián; Heras, Sonia; Ramis, Guillermo; Gadea, Joaquín; Coy, Pilar; Romar Andrés, Raquel; Anatomía y Anatomía Patológica Comparadas
    Rising livestock demand has increased use of assisted reproductive technologies (ART). Evidence indicates long-term effects of ART on molecular physiology and metabolism. This study explores ART's impact on biochemical and haematological parameters in a colony of pigs from 1 to 2.5 years of age. Animals were born after artificial insemination (AI) and transfer of embryos in vitro produced with (RF-IVP) or without (C-IVP) oviductal and uterine fluids during in vitro fertilization and embryo culture media (París-Oller et al., J Anim Sci and Biotechnol, 2021). Pigs were maintained under identical conditions and sampled every six months. Number of animals in each group decreased throughout life: 33 to 9 (AI); 22 to 10 (C-IVP); and 14 to 4 (RF-IVP). The number ofblood samples was the same as the number of animals and were collected via jugular venipuncture in lithium heparin tubes and immediately transported to laboratory. Plasma was obtained through centrifugation (1200 g, 20 min, 4 ºC, Eppendorf 5810 R) of blood collected via direct jugular venipuncture in lithium heparin tubes. Plasma was assessed using a chemistry analyser (Olympus AU400, Japan) for creatinine (CREA), urea, amylase, creatine kinase (CK), cholesterol, alkaline phosphatase (ALP), gamma-glutamyl transferase (GGT), glucose, aspartate aminotransferase (AST), alanine aminotransferase (ALT), lipase, total protein (TP), albumin (ALB), globulin (GLOB), triglycerides (TRIGL), and total bilirubin (TBIL). A haematology analyser (Siemens ADVIA® 120, USA) assessed red blood cell (RBC) count, haemoglobin concentration (HB), haematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), cell haemoglobin concentration mean (CHCM), red blood cell distribution width (RDW), haemoglobin concentration distribution width (HDW), white blood cell (WBC) count; and the differential count of neutrophils, lymphocytes, monocytes, eosinophils, and basophils. Reticulocyte percentage, mean corpuscular volume of reticulocytes (MCVr), haemoglobin content in reticulocytes (CHr) and platelet count were analysed. Data were analysed by mixed-effects model and likelihood-ratio test to determine effects of age and sex on the variables. Post-hoc test for multiple comparisons Tukey's method was used. Statistical significance was set at P < 0.05. Sex significantly influenced biochemical and haematological profiles. In biochemical parameters, females exhibited higher ALBU and urea concentrations, while males demonstrated elevated GLOB levels. Haematological analysis revealed sex-specific differences: females showed increased CH and PCDW values, whereas males presented higher HDW, PDW, and MPC parameters. Age-sex interactions were observed for TP, GLOB, urea, TRIGLI, amylase, and TBIL. Significant age-related changes occurred in RDW, CH, HDW, WBC, neutrophil percentage and count, lymphocyte percentage, PMDW, CHr, and MCVr. The interaction between sex and group was observed in ALT, HCT, and percentage of reticulocytes. ALT showed differences in males between C-IVP and RF-IVP group (p=0.01); HCT also in males between AI and both C-IVP and RF-IVP groups (p=0.01; p=0.02 respectively). A significant interaction was demonstrated between AI and C-IVP in males, as well as in the percentage of reticulocytes. These physiological findings provide essential reference data for veterinary practitioners and livestock producers, demonstrating minor long-term biochemical and haematological variations between naturally and artificially conceived swine. However, observed differences appear clinically insignificant, suggesting comparable physiological development regardless of conception method.
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    Open Access
    Long-term blood plasma biochemical analysis in pigs derived from assisted reproductive technologies.
    (2024) Mikita, Małgorzata; Romero-Aguirregomezcorta, Jon; Cánovas, Sebastián; Ramis, Guillermo; Gadea, Joaquín; Coy, Pilar; Romar, Raquel; Heras, Sonia; Anatomía y Anatomía Patológica Comparadas
    Rising livestock demand has increased use of assisted reproductive technologies (ART). Evidence indicates long-term effects of ART on molecular physiology and metabolism. This study explores ART’s impact on biochemical parameters in a colony of pigs from 1 to 5 years of age. Animals were born after artificial insemination (AI) and transfer of embryos in vitro produced with (RF-IVP) or without (C-IVP) oviductal and uterine fluids during in vitro fertilization and embryo culture media (París-Oller et al., J Anim Sci and Biotechnol, 2021). Pigs were maintained under identical conditions and sampled every six months. Number of animals in each group decreased throughout life: 33 to 9 (AI); 22 to 10 (C-IVP); and 14 to 4 (RF-IVP). The number of blood samples was the same as the number of animals and were collected via jugular venipuncture in lithium heparin tubes and immediately transported to laboratory. Plasma was obtained through entrifugation (1200 g, 20 min, 4ºC, Eppendorf 5810 R) of blood collected via direct jugular venipuncture in lithium heparin tubes. Plasma was assessed using a chemistry analyzer (Olympus AU400, Japan) for creatinine (CREA), urea, amylase, creatine kinase (CK), cholesterol, alkaline phosphatase (ALP), gamma-glutamyl transferase (GGT), glucose, aspartate aminotransferase (AST), alanine aminotransferase (ALT), lipase, total protein (TP), albumin (ALB), globulin (GLOB), triglycerides (TRIGL), and total bilirubin (TBIL). Data were analyzed by mixed-effects model and likelihood-ratio test to determine effects of age, group and sex on the variables. Post-hoc test for multiple comparisons Tukey’s method was used. Statistical significance was set at P < 0.05. Age significantly influenced all variables. Glucose, cholesterol, lipase, CK, ALP, AST, ALT were not affected by any other variable. TP, GLOB, UREA and TRIGL showed progressive elevation with age, with peak values between 4-5 years. ALB, cholesterol, CK, ALP, AST, GGT exhibited characteristic juvenile decline, with ALP and AST showing reduction during early development (155 to 52 U/L and 121 to 37 U/L respectively) followed by stabilization, while ALT displayed oscillatory changes. Glucose and TBIL showed initial elevation peaking at 2 years, before decreasing. CREA and amylase fluctuated irregularly. Critical developmental transitions occurred at 1.5-2 years and 4-5 years, representing key physiological maturation milestones. ALBU showed the highest mean concentration in the RF-IVP group and lowest in the AI group, while GGT showed the opposite pattern. ALBU, GLOB and urea were influenced by sex, with females higher ALBU and urea levels, while males showed elevated GLOB values. Long-term results confirm slight persisting biochemical differences in naturally and artificially conceived pigs, although the clinical relevance is unnoticeable.
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    Publication
    Open Access
    Long-term haematological analysis in pigs derived from assisted reproductive technologies
    (Brazilian College of Animal Reproduction, 2025) Mikita, Małgorzata; Romero-Aguirregomezcorta, Jon; Cánovas, Sebastián; Heras, Sonia; Ramis, Guillermo; Gadea, Joaquín; Coy, Pilar; Romar, Raquel; Anatomía y Anatomía Patológica Comparadas
    Rising livestock demand has increased use of assisted reproductive technologies (ART). Evidence indicates long-term effects of ART on molecular physiology and metabolism. This study explores ART’s impact on haematological parameters in a colony of pigs from 1 to 5 years of age. Animals were born after artificial insemination (AI) and transfer of embryos in vitro produced with (RF-IVP) or without (C-IVP) oviductal and uterine fluids during in vitro fertilization and embryo culture media (París-Oller et al., J Anim Sci and Biotechnol, 2021). Pigs were maintained under identical conditions and sampled every six months. Number of animals in each group decreased throughout life: 33 to 9 (AI); 22 to 10 (C-IVP); and 14 to 4 (RF-IVP). The number of blood samples was the same as the number of animals and were collected via jugular venipuncture in lithium heparin tubes and immediately transported to laboratory. A haematology analyzer (Siemens ADVIA® 120, USA) assessed red blood cell (RBC) count, haemoglobin concentration (HB), haematocrit (HCT), mean corpuscular volume (MCV), mean corpuscular haemoglobin (MCH), mean corpuscular haemoglobin concentration (MCHC), cell haemoglobin concentration mean (CHCM), red blood cell distribution width (RDW), haemoglobin concentration distribution width (HDW), white blood cell (WBC) count; and the differential count of neutrophils, lymphocytes, monocytes, eosinophils, and basophils. Reticulocyte percentage, mean corpuscular volume of reticulocytes (MCVr), haemoglobin content in reticulocytes (CHr) and platelet count were analyzed. Data were analyzed by mixed-effects model and likelihood-ratio test to determine effects of age, group and sex on the variables. Post-hoc test for multiple comparisons Tukey’s method were used. P<0.05 was considered significant. Age affected all variables. MCH, CH, CHCM, WBC, and platelet indices (MPV, MPC, MPM, PMDW) increased through life. Biphasic trajectories occurred in MCV, HTC, RBC, neutrophil counts, monocyte counts, PLT counts, and lymphocytes (secondyear decline followed by increase). Progressive decline with age was observed in monocyte percentage and reticulocyte parameters. Irregular oscillatory patterns dominated HB, RDW, MCHC, CHDW, HDW, eosinophil and basophil parameters, and advanced platelet indices (large PLT, PCT, PDW, MCVr, PCDW). HCT, RBC, and HB reached lowest values in AI group, highest in RF-IVP. HCM, MCHC, and lymphocyte percentage showed lowest means in AI, highest in C-IVP. RDW, WBC, neutrophil parameters, and lymphocyte count were lowest in C-IVP, highest in AI. Monocyte count exhibited lowest values in C-IVP, highest in RF-IVP group. CH and PCDW were higher in females than males, whereas HDW, PDW, MPC were higher in males. Long-term results confirm slight persisting haematological differences in naturally and artificially conceived pigs, although the clinical relevance is unnoticeable.
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    Open Access
    The impact of in vitro embryo production on placental and umbilical cord vascularization is minimized by the addition of reproductive fluids
    (Elsevier, 2023-09-15) Párraga Ros, Ester; Álvarez Martín, Úrsula; Seva Alcaraz, Juan; Coy, Pilar; Romar, Raquel; Anatomía y Anatomía Patológica Comparadas
    Animals born from in-vitro-produced (IVP) embryos show changes in the placenta and umbilical cord vascularization. This study compares the placental and umbilical vascular morphometry in pigs (n = 19) born through artificial insemination (AI group) or after transfer of IVP embryos cultured with (RF-IVP group) or without (C-IVP group) reproductive fluids. The relationship between vascular parameters and animal growth during the first year of life was also analyzed. Samples were collected at birth, fixed, paraffin-embedded, cut in sections, stained, and photographed for vascular and morphometric analysis with ImageJ® and Slide Viewer®. The average daily weight gain was individually scored from birth to the first year of life. No differences were found in placental vascular morphometry among groups, except for the vascular area of small vessels (arterioles, venules, and small vessels) that was higher in the C-IVP group. Regarding the umbilical cord, the values for perimeter (AI: 26.40 ± 3.93 mm; IVP: 30.51 ± 4.74 mm), diameter (AI: 8.35 ± 1.01 mm; IVP: 10.26 ± 1.85 mm), area (AI: 43.18 ± 12.87; IVP: 56.61 ± 14.89 mm2), and Wharton's jelly area (AI: 36.86 ± 12.04 mm2; IVP 48.88 ± 12.80 mm2) were higher in IVP-derived than AI-derived animals, whereas arterial and venous morphometric data were similar between groups. A correlation study showed that placental and umbilical cord vascular phenotypes affect the further growth of pigs. In conclusion, assisted reproductive technologies impact small caliber vessels in the placenta and morphometric parameters in the umbilical cord. The addition of reproductive fluids in IVP-embryo contributes to reduce the differences with in vivo-derived animals.