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1.
BMC Genomics ; 25(1): 589, 2024 Jun 12.
Artículo en Inglés | MEDLINE | ID: mdl-38867150

RESUMEN

BACKGROUND: Bisphenol S (BPS) is a substitute for bisphenol A in plastic manufacturing and, as a potential endocrine disruptor, may alter the physiology of the oviduct, in which fertilization and early embryo development take place in mammals. The objective of this study was to assess the effect of a daily dietary exposure to BPS combined with a contrasted diet on the oviduct fluid proteome using an ovine model. RESULTS: Eighty adult cyclic ewes were allotted to four groups (20/group): overfed (OF) consuming 50 µg/kg/day of BPS in their diet, underfed (UF) consuming 50 µg/kg/day of BPS, and non-exposed controls in each diet group. After three months, the mean body condition score, plasma levels of glucose and non-esterified fatty acids were significantly higher in OF than in UF females. The proteins in collected OF samples (50 µg) were analyzed by nanoliquid chromatography coupled with tandem mass spectrometry (nanoLC-MS/MS). Overall, 1563 proteins were identified, among which 848 were quantified. Principal component analysis of the data revealed a clear discrimination of samples according to the diet and a segregation between BPS-exposed and non-exposed females in overfed ewes. Hierarchical clustering of differentially abundant proteins (DAPs) identified two clusters of 101 and 78 DAPs according to the diet. Pairwise comparisons between groups revealed a stronger effect of BPS in OF than in UF females (70 vs. 24 DAPs) and a stronger effect of the diet in BPS-exposed than non-exposed females (56 vs. 36 DAPs). Functional analysis of DAPs showed an enrichment in metabolic processes, immune system, cell response to stress, and reproductive processes. CONCLUSIONS: This work highlights for the first time the important impact of BPS on the oviduct proteome, with larger effects seen in OF than UF females. These results, together with previous ones, raise health concerns for everyone and call for a greater regulation of BPS in the food industry.


Asunto(s)
Oviductos , Fenoles , Proteoma , Sulfonas , Animales , Femenino , Ovinos , Fenoles/toxicidad , Proteoma/metabolismo , Oviductos/metabolismo , Oviductos/efectos de los fármacos , Sulfuros/administración & dosificación , Proteómica , Administración Oral , Dieta
2.
Theriogenology ; 217: 113-126, 2024 Mar 15.
Artículo en Inglés | MEDLINE | ID: mdl-38271765

RESUMEN

Most in vitro models of oviduct epithelial cells (OEC) used thus far to gain insights into embryo-maternal communication induce cell dedifferentiation or are technically challenging. Moreover, although the presence of developing embryos has been shown to alter gene expression in OEC, the effect of embryos on OEC physiology remains largely unknown. Here, we propose a model based on bovine oviduct epithelial spheroids (OES) with specific shape and diameter (100-200 µm) criteria. The aims of this study were to i) determine the appropriate culture conditions of bovine OES cultured in suspension by evaluating their morphology, total cell number, viability, and activity of ciliated cells; ii) monitor gene expression in OES at the time of their formation (day 0) and over the 10 days of culture; and iii) test whether the vicinity of developing embryos affects OES quality criteria. On day 10, the proportions of vesicle-shaped OES (V-OES) were higher in M199/500 (500 µl of HEPES-buffered TCM-199) and synthetic oviduct fluid (SOF)/25 (25-µL droplet of SOF medium under mineral oil) than in M199/25 (25-µL droplet of M199 under mineral oil). The proportion of viable cells in V-OES was not affected by culture conditions and remained high (>80%) through day 10. The total number of cells per V-OES decreased over time except in SOF/25, while the proportions of ciliated cells increased over time in M199/500 but decreased in M199/25 and SOF/25. The movement amplitude of OES in suspension decreased over time under all culture conditions. Moreover, the gene expression of ANXA1, ESR1, HSPA8, and HSPA1A in OES remained stable during culture, while that of PGR and OVGP1 decreased from day 0 to day 10. Last, the co-culture of developing embryos with OES in SOF/25 increased the rates of blastocysts on days 7 and 8 compared to embryos cultured alone, and increased the proportion of V-OES compared to OES cultured alone. In conclusion, M199/500 and SOF/25 provided the optimal conditions for the long-time culture of OES. The supporting effect of OES on embryo development and of developing embryos on OES morphology was evidenced for the first time. Altogether, these results point OES as an easy-to-use, standardizable, and physiological model to study embryo-maternal interactions in cattle.


Asunto(s)
Fertilización In Vitro , Aceite Mineral , Femenino , Bovinos , Animales , Fertilización In Vitro/veterinaria , Embrión de Mamíferos , Trompas Uterinas , Oviductos , Blastocisto/fisiología , Medios de Cultivo , Desarrollo Embrionario/fisiología
3.
Sci Rep ; 13(1): 10311, 2023 06 26.
Artículo en Inglés | MEDLINE | ID: mdl-37365288

RESUMEN

When entering the oviduct for fertilisation, spermatozoa come into contact with the oviduct fluid (OF) and can bind to luminal epithelial cells in the isthmus to form a sperm reservoir. The objective of this study was to examine how the OF modulates sperm adhesion to the oviduct reservoir using an in vitro model of oviduct epithelial spheroids (OES). Bovine oviducts from a local slaughterhouse were used to collect OF and isthmic fragments for the in vitro incubation of OES. Compared to a non-capacitating control medium, the pre-ovulatory OF significantly decreased by 80-90% the density of spermatozoa bound to OES without affecting sperm motility, membrane integrity, or sperm-cilia interactions. This effect on sperm binding was reproduced with (1) OF from different cycle stages and anatomical regions of the oviduct; (2) OF fractions of more than 3 kDa; (3) modified OF in which proteins were denatured or digested and (4) heparan sulphate but not hyaluronic acid, two glycosaminoglycans present in the OF. In conclusion, the OF significantly decreased the number of spermatozoa that bind to oviduct epithelial cells without affecting sperm motility and this effect was due to macromolecules, including heparan sulphate.


Asunto(s)
Glicosaminoglicanos , Motilidad Espermática , Femenino , Humanos , Masculino , Animales , Bovinos , Glicosaminoglicanos/metabolismo , Semen/metabolismo , Oviductos/metabolismo , Trompas Uterinas/metabolismo , Espermatozoides/metabolismo , Heparitina Sulfato/metabolismo
4.
J Anim Sci Biotechnol ; 14(1): 30, 2023 Feb 17.
Artículo en Inglés | MEDLINE | ID: mdl-36797800

RESUMEN

BACKGROUND: Spermatozoa interact with oviduct secretions before fertilization in vivo but the molecular players of this dialog and underlying dynamics remain largely unknown. Our objectives were to identify an exhaustive list of sperm-interacting proteins (SIPs) in the bovine oviduct fluid and to evaluate the impact of the oviduct anatomical region (isthmus vs. ampulla) and time relative to ovulation (pre-ovulatory vs. post-ovulatory) on SIPs number and abundance. METHODS: Pools of oviduct fluid (OF) from the pre-ovulatory ampulla, pre-ovulatory isthmus, post-ovulatory ampulla, and post-ovulatory isthmus in the side of ovulation were collected from the slaughterhouse. Frozen-thawed bull sperm were incubated with OF or phosphate-buffered saline (control) for 60 min at 38.5 °C. After protein extraction and digestion, sperm and OF samples were analyzed by nanoLC-MS/MS and label-free protein quantification. RESULTS: A quantitative comparison between proteins identified in sperm and OF samples (2333 and 2471 proteins, respectively) allowed for the identification of 245 SIPs. The highest number (187) were found in the pre-ovulatory isthmus, i.e., time and place of the sperm reservoir. In total, 41 SIPs (17%) were differentially abundant between stages in a given region or between regions at a given stage and 76 SIPs (31%) were identified in only one region × stage condition. Functional analysis of SIPs predicted roles in cell response to stress, regulation of cell motility, fertilization, and early embryo development. CONCLUSION: This study provides a comprehensive list of SIPs in the bovine oviduct and evidences dynamic spatio-temporal changes in sperm-oviduct interactions around ovulation time. Moreover, these data provide protein candidates to improve sperm conservation and in vitro fertilization media.

5.
Theriogenology ; 198: 332-343, 2023 Mar 01.
Artículo en Inglés | MEDLINE | ID: mdl-36640738

RESUMEN

Uterine secretions provide a suitable environment for sperm selective migration during a couple of days preceding ovulation and for early embryo development before implantation. Our goal was to identify and quantify proteins in the bovine uterine fluid during the periovulatory period of the estrous cycle. Genital tracts with normal morphology were collected from adult cyclic Bos taurus females in a local slaughterhouse and classified into pre-ovulatory or post-ovulatory stages of cycle (around days 19-21 and 0-5 of cycle, respectively; n = 8 cows per stage) based on ovarian morphology. Proteins from uterine fluid collected from the utero-tubal junction to the base of each horn (four pools of two cows per condition) were analyzed by nanoLiquid Chromatography coupled with tandem Mass Spectrometry (nanoLC-MS/MS). A total of 1214 proteins were identified, of which 91% were shared between all conditions. Overall, 57% of proteins were predicted to be secreted and 17% were previously reported in uterine extracellular vesicles. Paired comparisons between uterine horns ipsilateral and contralateral to ovulation evidenced 12 differentially abundant proteins, including five at pre-ovulatory stage. Furthermore, 35 proteins differed in abundance between pre- and post-ovulatory stages, including 21 in the ipsilateral side of ovulation. Functional analysis of identified proteins demonstrated roles in binding, metabolism, cellular detoxification and the immune response. This study provides a valuable database of uterine proteins for functional studies on sperm physiology and early embryo development.


Asunto(s)
Ovario , Proteoma , Femenino , Bovinos , Animales , Masculino , Ovario/metabolismo , Proteoma/metabolismo , Espectrometría de Masas en Tándem/veterinaria , Semen/metabolismo , Ciclo Estral/fisiología , Ovulación
6.
BMC Genomics ; 23(1): 839, 2022 Dec 19.
Artículo en Inglés | MEDLINE | ID: mdl-36536309

RESUMEN

BACKGROUND: Despite many improvements with in vitro culture systems, the quality and developmental ability of mammalian embryos produced in vitro are still lower than their in vivo counterparts. Though previous studies have evidenced differences in gene expression between in vivo- and in vitro-derived bovine embryos, there is no comparison at the protein expression level. RESULTS: A total of 38 pools of grade-1 quality bovine embryos at the 4-6 cell, 8-12 cell, morula, compact morula, and blastocyst stages developed either in vivo or in vitro were analyzed by nano-liquid chromatography coupled with label-free quantitative mass spectrometry, allowing for the identification of 3,028 proteins. Multivariate analysis of quantified proteins showed a clear separation of embryo pools according to their in vivo or in vitro origin at all stages. Three clusters of differentially abundant proteins (DAPs) were evidenced according to embryo origin, including 463 proteins more abundant in vivo than in vitro across development and 314 and 222 proteins more abundant in vitro than in vivo before and after the morula stage, respectively. The functional analysis of proteins found more abundant in vivo showed an enrichment in carbohydrate metabolism and cytoplasmic cellular components. Proteins found more abundant in vitro before the morula stage were mostly localized in mitochondrial matrix and involved in ATP-dependent activity, while those overabundant after the morula stage were mostly localized in the ribonucleoprotein complex and involved in protein synthesis. Oviductin and other oviductal proteins, previously shown to interact with early embryos, were among the most overabundant proteins after in vivo development. CONCLUSIONS: The maternal environment led to higher degradation of mitochondrial proteins at early developmental stages, lower abundance of proteins involved in protein synthesis at the time of embryonic genome activation, and a global upregulation of carbohydrate metabolic pathways compared to in vitro production. Furthermore, embryos developed in vivo internalized large amounts of oviductin and other proteins probably originated in the oviduct as soon as the 4-6 cell stage. These data provide new insight into the molecular contribution of the mother to the developmental ability of early embryos and will help design better in vitro culture systems.


Asunto(s)
Embrión de Mamíferos , Proteómica , Bovinos , Animales , Embrión de Mamíferos/metabolismo , Blastocisto , Proteínas/metabolismo , Mórula/metabolismo , Desarrollo Embrionario , Mamíferos
7.
Front Cell Dev Biol ; 10: 863700, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35386205

RESUMEN

Early embryo development is a dynamic process involving important molecular and structural changes leading to the embryonic genome activation (EGA) and early cell lineage differentiation. Our aim was to elucidate proteomic changes in bovine embryos developed in vivo. Eleven females were used as embryo donors and pools of embryos at the 4-6 cell, 8-12 cell, morula, compact morula and blastocyst stages were analyzed by nanoliquid chromatography coupled with label free quantitative mass spectrometry. A total of 2,757 proteins were identified, of which 1,950 were quantitatively analyzed. Principal component analysis of data showed a clear separation of embryo pools according to their developmental stage. The hierarchical clustering of differentially abundant proteins evidenced a first cluster of 626 proteins that increased in abundance during development and a second cluster of 400 proteins that decreased in abundance during development, with most significant changes at the time of EGA and blastocyst formation. The main pathways and processes overrepresented among upregulated proteins were RNA metabolism, protein translation and ribosome biogenesis, whereas Golgi vesicle transport and protein processing in endoplasmic reticulum were overrepresented among downregulated proteins. The pairwise comparison between stages allowed us to identify specific protein interaction networks and metabolic pathways at the time of EGA, morula compaction and blastocyst formation. This is the first comprehensive study of proteome dynamics in non-rodent mammalian embryos developed in vivo. These data provide a number of protein candidates that will be useful for further mechanistic and functional studies.

8.
Sci Rep ; 12(1): 4135, 2022 03 09.
Artículo en Inglés | MEDLINE | ID: mdl-35264682

RESUMEN

Understanding the composition of the oviduct fluid (OF) is crucial to better comprehend the microenvironment in which sperm capacitation, fertilization and early embryo development take place. Therefore, our aim was to determine the spatiotemporal changes in the OF proteome according to the anatomical region of the oviduct (ampulla vs. isthmus), the proximity of the ovulating ovary (ipsilateral vs. contralateral side) and the peri-ovulatory stage (pre-ovulatory or Pre-ov vs. post-ovulatory or Post-ov). Oviducts from adult cyclic cows were collected at a local slaughterhouse and pools of OF were analyzed by nanoLC-MS/MS and label-free protein quantification (n = 32 OF pools for all region × stage × side conditions). A total of 3760 proteins were identified in the OF, of which 65% were predicted to be potentially secreted. The oviduct region was the major source of variation in protein abundance, followed by the proximity of the ovulating ovary and finally the peri-ovulatory stage. Differentially abundant proteins between regions, stages and sides were involved in a broad variety of biological functions, including protein binding, response to stress, cell-to-cell adhesion, calcium homeostasis and the immune system. This work highlights the dynamic regulation of oviduct secretions and provides new protein candidates for interactions between the maternal environment, the gametes and the early embryo.


Asunto(s)
Oviductos , Proteoma , Animales , Bovinos , Trompas Uterinas/metabolismo , Femenino , Oviductos/metabolismo , Ovulación/fisiología , Proteoma/metabolismo , Espectrometría de Masas en Tándem
9.
Biol Reprod ; 105(2): 317-331, 2021 08 03.
Artículo en Inglés | MEDLINE | ID: mdl-34057175

RESUMEN

In vitro fertilization (IVF) gives rise to embryos in a number of mammalian species and is currently widely used for assisted reproduction in humans and for genetic purposes in cattle. However, the rate of polyspermy is generally higher in vitro than in vivo and IVF remains ineffective in some domestic species like pigs and horses, highlighting the importance of the female reproductive tract for gamete quality and fertilization. In this review, the way the female environment modulates sperm selective migration, survival, and acquisition of fertilizing ability in the oviduct is being considered under six aspects: (1) the utero-tubal junction that selects a sperm sub-population entering the oviduct; (2) the presence of sperm binding sites on luminal epithelial cells in the oviduct, which prolong sperm viability and plays a role in limiting polyspermic fertilization; (3) the contractions of the oviduct, which promote sperm migration toward the site of fertilization in the ampulla; (4) the regions of the oviduct, which play different roles in regulating sperm physiology and interactions with oviduct epithelial cells; (5) the time of ovulation, and (6) the steroid hormonal environment which regulates sperm release from the luminal epithelial cells and facilitates capacitation in a finely orchestrated manner.


Asunto(s)
Movimiento Celular , Supervivencia Celular , Fertilización , Oviductos/fisiología , Espermatozoides/fisiología , Animales , Femenino , Humanos , Masculino , Mamíferos
10.
Reprod Biol ; 21(3): 100512, 2021 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-33991764

RESUMEN

The metabolites in the oviduct fluid (OF) of both oviducts were analyzed by proton nuclear magnetic resonance (1H-NMR) in Holstein heifers on day 3 after synchronized estrus. Twenty-six metabolites were quantified, among which lactate, glycine and myoinositol were the most abundant. Six metabolites including glycine and myoinositol varied in amount according to the proximity to the corpus luteum. Glucose and histidine were among the most variable metabolites among heifers while threonine and lactate were among the most stable ones, suggesting different mechanisms of homeostasis in the OF.


Asunto(s)
Líquidos Corporales/química , Líquidos Corporales/fisiología , Bovinos/fisiología , Estro/fisiología , Trompas Uterinas/fisiología , Metabolómica , Animales , Femenino
11.
Mol Cell Endocrinol ; 516: 110956, 2020 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-32712384

RESUMEN

Sperm migration through the female genital tract is not a quiet journey. Uterine contractions quickly operate a drastic selection, leading to a very restrictive number of sperm reaching the top of uterine horns and finally, provided the presence of key molecules on sperm, the oviduct, where fertilization takes place. During hours and sometimes days before fertilization, subpopulations of spermatozoa interact with dynamic and region-specific maternal components, including soluble proteins, extracellular vesicles and epithelial cells lining the lumen of the female tract. Interactions with uterine and oviductal cells play important roles for sperm survival as they modulate the maternal immune response and allow a transient storage before ovulation. The body of work reported here highlights the importance of sperm interactions with proteins originated from both the uterine and oviductal fluids, as well as hormonal signals around the time of ovulation for sperm acquisition of fertilizing competence.


Asunto(s)
Genitales Femeninos/metabolismo , Mamíferos/fisiología , Interacciones Espermatozoide-Óvulo , Espermatozoides/metabolismo , Animales , Femenino , Humanos , Masculino , Embarazo
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