RESUMEN
Among factors like hormonal imbalance and uterine condition, oocyte quality is regarded as one of the key factors involved in age-related decline in the reproductive capacity. Here, are discussions about the functions played by organelles within the oocyte in forming the next generation that is more suitable for survival. Many insights on the adaptation to aging and maintenance of quality can be obtained from: interactions between mitochondria and other organelles that enable the long life of primordial oocytes; characteristics of organelle interactions after breaking dormancy from primary oocytes to mature oocytes; and characteristics of interactions between mitochondria and other organelles of aged oocytes collected during the ovulatory cycle from elderly individuals and animals. This information would potentially be beneficial to the development of future therapeutic methods or agents.
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Mitocondrias , Oocitos , Oocitos/metabolismo , Oocitos/fisiología , Humanos , Mitocondrias/metabolismo , Mitocondrias/fisiología , Animales , Femenino , Orgánulos/metabolismo , Orgánulos/fisiología , Envejecimiento/fisiologíaRESUMEN
Vitrification of oocyte has become an important component of assisted reproductive technology and has important implications for animal reproduction and the preservation of biodiversity. However, vitrification adversely affects mitochondrial function and oocyte developmental potential, mainly because of oxidative damage. Rutin is a highly effective antioxidant, but no information is available to the effect of rutin on the mitochondrial function and development in vitrified oocytes. Therefore, we studied the effects of rutin supplementation of vitrification solution on mitochondrial function and developmental competence of ovine germinal vesicle (GV) stage oocytes post vitrification. The results showed that supplementation of vitrification solution with 0.6 mM rutin significantly increased the cleavage rate (71.6 % vs. 59.3 %) and blastocyst rate (18.9 % vs. 6.8 %) compared to GV-stage oocytes in the vitrified group. Then, we analyzed the reactive oxygen species (ROS), glutathione (GSH), mitochondrial activity and membrane potential (ΔΨm), endoplasmic reticulum (ER) Ca2+, and annexin V (AV) of vitrified sheep GV-stage oocytes. Vitrified sheep oocytes exhibited increased levels of ROS and Ca2+, higher rate of AV-positive oocytes, and decreased mitochondrial activity, GSH and ΔΨm levels. However, rutin supplementation in vitrification solution decreased the levels of ROS, Ca2+ and AV-positive oocytes rate, and increased the GSH and ΔΨm levels in vitrified oocytes. Results revealed that rutin restored mitochondrial function, regulated Ca2+ homeostasis and decreased apoptosis potentially caused by mitophagy in oocytes. To understand the mechanism of rutin functions in vitrified GV-stage oocytes in sheep, we analyzed the transcriptome and found that rutin mediated oocytes development and mitochondrial function, mainly by affecting oxidative phosphorylation and the mitophagy pathways. In conclusion, supplementing with 0.6 mM rutin in vitrification solution significantly enhanced developmental potential through improving mitochondrial function and decreased apoptosis potentially caused by mitophagy after vitrification of ovine GV-stage oocytes.
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Criopreservación , Mitocondrias , Oocitos , Rutina , Vitrificación , Animales , Rutina/farmacología , Oocitos/efectos de los fármacos , Oocitos/fisiología , Ovinos/fisiología , Mitocondrias/efectos de los fármacos , Vitrificación/efectos de los fármacos , Criopreservación/veterinaria , Especies Reactivas de Oxígeno/metabolismo , Femenino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Técnicas de Maduración In Vitro de los Oocitos/métodos , Antioxidantes/farmacología , Desarrollo Embrionario/efectos de los fármacosRESUMEN
Assisted reproductive technologies (ARTs) are performed worldwide in the equine industry to produce genetically valuable foals. Among them, ovum pick up (OPU) combined with intra-cytoplasmic sperm injection (ICSI) can now be more efficient than embryo transfer (ET) under optimal conditions. However, OPU is not a benign procedure for the mare and the process is costly. Improved efficiency is therefore in the interest of everyone, maximizing mare welfare and optimizing economics for the client. One of the key factors of success is the antral follicle count (AFC) at the time of OPU and subsequently the number of oocytes obtained. Variations in AFC are reported between individuals and between geographical areas. This leads to a significant increase in numbers of embryos produced per session in some countries compared to others, independent of the laboratory efficiency. This article revisits the basics of folliculogenesis involved in establishment of the antral follicle population and explores work in other species given the paucity of equine research in this area. The aim of the review is to elucidate interesting areas of further research that could generate essential information for clinicians and clients about the management and selection of the donor mare for OPU and potentially identify pharmacological targets for manipulation.
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Folículo Ovárico , Técnicas Reproductivas Asistidas , Caballos/fisiología , Animales , Femenino , Técnicas Reproductivas Asistidas/veterinaria , Oocitos/fisiología , Inyecciones de Esperma Intracitoplasmáticas/veterinaria , Inyecciones de Esperma Intracitoplasmáticas/métodos , Transferencia de Embrión/veterinaria , Transferencia de Embrión/métodos , Recuperación del Oocito/veterinaria , Recuperación del Oocito/métodos , EmbarazoRESUMEN
Calcium (Ca2+) is a second messenger for many signal pathways, and changes in intracellular Ca2+ concentration ([Ca2+]i) are an important signaling mechanism in the oocyte maturation, activation, fertilization, function regulation of granulosa and cumulus cells and offspring development. Ca2+ oscillations occur during oocyte maturation and fertilization, which are maintained by Ca2+ stores and extracellular Ca2+ ([Ca2+]e). Abnormalities in Ca2+ signaling can affect the release of the first polar body, the first meiotic division, and chromosome and spindle morphology. Well-studied aspects of Ca2+ signaling in the oocyte are oocyte activation and fertilization. Oocyte activation, driven by sperm-specific phospholipase PLCζ, is initiated by concerted intracellular patterns of Ca2+ release, termed Ca2+ oscillations. Ca2+ oscillations persist for a long time during fertilization and are coordinately engaged by a variety of Ca2+ channels, pumps, regulatory proteins and their partners. Calcium signaling also regulates granulosa and cumulus cells' function, which further affects oocyte maturation and fertilization outcome. Clinically, there are several physical and chemical options for treating fertilization failure through oocyte activation. Additionally, various exogenous compounds or drugs can cause ovarian dysfunction and female infertility by inducing abnormal Ca2+ signaling or Ca2+ dyshomeostasis in oocytes and granulosa cells. Therefore, the reproductive health risks caused by adverse stresses should arouse our attention. This review will systematically summarize the latest research progress on the aforementioned aspects and propose further research directions on calcium signaling in female reproduction.
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Señalización del Calcio , Oocitos , Oocitos/metabolismo , Oocitos/fisiología , Humanos , Señalización del Calcio/fisiología , Femenino , Animales , Calcio/metabolismo , Fertilización/fisiología , Células del Cúmulo/metabolismoRESUMEN
Zebrafish serve as a valuable model organism for studying germ cell biology and reproductive processes. The AB strain of zebrafish is proposed to exhibit a polygenic sex determination system, where most males initially develop juvenile ovaries before committing to male fate. In species with chromosomal sex determination, gonadal somatic cells are recognized as key determinants of germ cell fate. Notably, the loss of germ cells in zebrafish leads to masculinization, implying that germ cells harbor an intrinsic feminization signal. However, the specific signal triggering oogenesis in zebrafish remains unclear. In the present study, we identified foxl2l as an oocyte progenitor-specific gene essential for initiating oogenesis in germ cells. Results showed that foxl2l-knockout zebrafish bypassed the juvenile ovary stage and exclusively developed into fertile males. Further analysis revealed that loss of foxl2l hindered the initiation of oocyte-specific meiosis and prevented entry into oogenesis, leading to premature spermatogenesis during early gonadal development. Furthermore, while mutation of the pro-male gene dmrt1 led to fertile female differentiation, simultaneous disruption of foxl2l in dmrt1 mutants completely blocked oogenesis, with a large proportion of germ cells arrested as germline stem cells, highlighting the crucial role of foxl2l in oogenesis. Overall, this study highlights the unique function of foxl2l as a germ cell-intrinsic gatekeeper of oogenesis in zebrafish.
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Oogénesis , Proteínas de Pez Cebra , Pez Cebra , Animales , Pez Cebra/genética , Pez Cebra/fisiología , Oogénesis/fisiología , Oogénesis/genética , Femenino , Masculino , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismo , Células Germinativas/fisiología , Proteína Forkhead Box L2/genética , Proteína Forkhead Box L2/metabolismo , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Espermatogénesis/fisiología , Espermatogénesis/genética , Oocitos/fisiologíaRESUMEN
Ovum Pick Up (OPU) is a minimally invasive technique widely used in cattle and mares for oocyte retrieval, involving ultrasound-guided puncture of ovarian follicles. It has been demonstrated that this technique is safe for its repeated use in the same female without affecting her reproductive health, allowing for the retrieval of oocytes in individuals regardless of their reproductive status. The oocytes obtained through OPU can subsequently be used for in vitro embryo production (IVP) using assisted reproductive techniques (ARTs) or be cryopreserved in biobanks for their future use. Traditionally, the minimally invasive technique of choice performed in vivo in domestic and wild felines was LOPU (laparoscopic-guided ovum pick up). The present study was designed to explore if ultrasound-guided OPU in the domestic cat is safe and effective. In an initial series of ex vivo experiments (n = 92 ovaries, n = 434 oocytes), the effect of different aspiration pressures for oocyte collection was explored. These experiments identified 43 mmHg as the optimal aspiration pressure, resulting in the highest recovery rate and a favorable maturation and blastocyst rate. Subsequently, 16 grade I and II oocytes were retrieved by OPU and 101 oocytes were retrieved following ovariectomy and slicing. Sixteen oocytes obtained with each technique were subjected to in vitro maturation (IVM) and in vitro fertilization (IVF). A total of 14 presumptive zygotes were selected for in vitro culture (IVC) from each group (OPU and slicing), obtaining a cleavage rate of 57.1 % and 64.2 %, a morula rate of 28.5 % in both groups, and a blastocyst rate of 7.14 % and 14.2 % respectively. The hormonal stimulation protocol was well-tolerated, with no adverse effects observed. Moreover, no complications arose during the ovariectomy performed post-OPU. The use of this technique in domestic cats represents a significant step forward in terms of safety, replicability, and invasiveness, serving as a valuable model for its application in wild felids species. Additional research involving a greater number of animals is required to validate these encouraging findings.
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Fertilización In Vitro , Recuperación del Oocito , Animales , Gatos/fisiología , Femenino , Recuperación del Oocito/veterinaria , Recuperación del Oocito/métodos , Fertilización In Vitro/veterinaria , Fertilización In Vitro/métodos , Técnicas de Cultivo de Embriones/veterinaria , Ultrasonografía/veterinaria , Ultrasonografía/métodos , Oocitos/fisiología , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Técnicas de Maduración In Vitro de los Oocitos/métodosRESUMEN
The wide application of ovine oocyte vitrification is limited by its relatively low efficiency. Nanoparticle is potentially to be used in cryopreservation technology for its unique characteristics with high biocompatibility, potent antioxidant property as well as superiority in membrane permeation and heat transduction. However, the effect of nanoparticle on ovine oocyte cryopreservation as well as the underlying mechanism has not been systematically evaluated. The objective of this study was to investigate the impact of nanoparticles on ovine oocytes cryopreservation and further identify the underlying mechanism. Firstly, the effects of Hydroxyapatite (HA) and Fe3O4 nanoparticles on the developmental potential of vitrified ovine oocytes were determined, and the results showed that neither HA (VC = 85.95 ± 6.23 % vs. VH = 92.47 ± 8.11 %, P > 0.05) nor Fe3O4 (VC = 85.95 ± 6.23 % vs. VF = 89.39 ± 6.32 %, P > 0.05) had adverse effect on the survival rate of vitrified-thawed oocytes. Notably, both HA (VC = 77.78 ± 0.09 % vs. VH = 44.00 ± 0.09 %, Pï¼0.01) and Fe3O4 (VC = 77.78 ± 0.09 % vs. VF = 51.67 ± 0.15 %, Pï¼0.01) nanoparticles effectively reduced the level of oocyte apoptosis after freezing and thawing. What's more, HA could significantly improve the cleavage rate of frozen oocytes (VC = 33.79 ± 2.83 % vs. VH = 59.54 ± 4.13 %, Pï¼0.05). Moreover, reduced reactive oxygen species (ROS) level (VC = 13.66 ± 0.47 vs. VH = 12.61 ± 0.53, P < 0.05), increased glutathione (GSH) content (VC = 60.69 ± 7.89 vs. VH = 87.92 ± 1.05, P < 0.05) and elevated mitochondrial membrane potential (MMP) level (VC = 1.43 ± 0.04 vs. VH = 1.63 ± 0.01ï¼Pï¼0.01) were observed in oocytes treated with HA nanoparticles when compared with that of the control group. Furthermore, Smart-RNA sequence technology was utilized to identify differentially expressed mRNAs (DEMs) induced by nanoparticles during cryopreservation. When compared with the control counterparts, a total of 721 DEMs (309 up-regulated and 412 down-regulated mRNAs) were identified in oocytes treated with HA, while 702 DEMs (480 up-regulated and 222 down-regulated mRNAs) were identified in oocytes treated with Fe3O4. A comparison of DEMs showed that total 692 mRNAs were expressed in oocytes treated with HA and Fe3O4. Notably, we discovered that 15 mRNAs were specially highly expressed in oocytes treated with HA, and Focal adhesion signaling pathway mainly contributed to the improved ovine oocyte quality after vitrification by alleviating oxidative stress.
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Criopreservación , Durapatita , Nanopartículas , Oocitos , Estrés Oxidativo , Vitrificación , Animales , Oocitos/efectos de los fármacos , Oocitos/fisiología , Ovinos/fisiología , Estrés Oxidativo/efectos de los fármacos , Durapatita/farmacología , Criopreservación/veterinaria , Criopreservación/métodos , Femenino , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Calcium ions (Ca2+) regulate cell proliferation and differentiation and participate in various physiological activities of cells. The calcium transfer protein inositol 1,4,5-triphosphate receptor (IP3R), located between the endoplasmic reticulum (ER) and mitochondria, plays an important role in regulating Ca2+ levels. However, the mechanism by which IP3R1 affects porcine meiotic progression and embryonic development remains unclear. We established a model in porcine oocytes using siRNA-mediated knockdown of IP3R1 to investigate the effects of IP3R1 on porcine oocyte meiotic progression and embryonic development. The results indicated that a decrease in IP3R1 expression significantly enhanced the interaction between the ER and mitochondria. Additionally, the interaction between the ER and the mitochondrial Ca2+ ([Ca2+]m) transport network protein IP3R1-GRP75-VDAC1 was disrupted. The results of the Duolink II in situ proximity ligation assay (PLA) revealed a weakened pairwise interaction between IP3R1-GRP75 and VDAC1 and a significantly increased interaction between GRP75 and VDAC1 after IP3R1 interference, resulting in the accumulation of large amounts of [Ca2+]m. These changes led to mitochondrial oxidative stress, increased the levels of reactive oxygen species (ROS) and reduced ATP production, which hindered the maturation and late development of porcine oocytes and induced apoptosis. Nevertheless, after treat with [Ca2+]m chelating agent ruthenium red (RR) or ROS scavenger N-acetylcysteine (NAC), the oocytes developmental abnormalities, oxidative stress and apoptosis caused by Ca2+ overload were improved. In conclusion, our results indicated IP3R1 is required for meiotic progression and embryonic development by regulating mitochondrial calcium and oxidative damage.
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Calcio , Desarrollo Embrionario , Receptores de Inositol 1,4,5-Trifosfato , Meiosis , Mitocondrias , Estrés Oxidativo , Animales , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/genética , Porcinos , Mitocondrias/metabolismo , Mitocondrias/fisiología , Meiosis/fisiología , Calcio/metabolismo , Desarrollo Embrionario/fisiología , Especies Reactivas de Oxígeno/metabolismo , Oocitos/fisiología , FemeninoRESUMEN
Fertilization capacity and embryo survival rate are decreased in postovulatory aging oocytes, which results in a reduced reproductive rate in female animals. However, the key regulatory genes and related regulatory mechanisms involved in the process of postovulatory aging in oocytes remain unclear. In this study, RNA-Seq revealed that 3237 genes were differentially expressed in porcine oocytes between the MII and aging stages (MII + 24 h). The expression level of FOXM1 was increased at the aging stage, and FOXM1 was also observed to be enriched in many key biological processes, such as cell senescence, response to oxidative stress, and transcription, during porcine oocyte aging. Previous studies have shown that FOXM1 is involved in the regulation of various biological processes, such as oxidative stress, DNA damage repair, mitochondrial function, and cellular senescence, which suggests that FOXM1 may play a crucial role in the process of postovulatory aging. Therefore, in this study, we investigated the effects and mechanisms of FOXM1 on oxidative stress, mitochondrial function, DNA damage, and apoptosis during oocyte aging. Our study revealed that aging oocytes exhibited significantly increased ROS levels and significantly decreased GSH, SOD, T-AOC, and CAT levels than did oocytes at the MII stage and that FOXM1 inhibition exacerbated the changes in these levels in aging oocytes. In addition, FOXM1 inhibition increased the levels of DNA damage, apoptosis, and cell senescence in aging oocytes. A p21 inhibitor alleviated the effects of FOXM1 inhibition on oxidative stress, mitochondrial function, and DNA damage and thus alleviated the degree of senescence in aging oocytes. These results indicate that FOXM1 plays a crucial role in porcine oocyte aging. This study contributes to the understanding of the function and mechanism of FOXM1 during porcine oocyte aging and provides a theoretical basis for preventing oocyte aging and optimizing conditions for the in vitro culture of oocytes.
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Senescencia Celular , Daño del ADN , Proteína Forkhead Box M1 , Mitocondrias , Oocitos , Estrés Oxidativo , Animales , Oocitos/fisiología , Oocitos/metabolismo , Porcinos , Proteína Forkhead Box M1/metabolismo , Proteína Forkhead Box M1/genética , Mitocondrias/metabolismo , Femenino , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Regulación de la Expresión GénicaRESUMEN
The aim of the study was to investigate the relationship between ooplasm morphology, lipid content, glucose-6-phosphate dehydrogenase activity (G6PDH) and maturation potential of domestic cat oocytes. Cumulus-oocyte complexes were classified according to ooplasm morphology: evenly dark (dCOC), heterogeneous/mosaic (hCOC), or light/transparent (lCOC), however only dCOCs are thought to be the best-quality, the remaining ones are usually rejected, therefore little is known about their intracellular properties. Lipid droplets (LDs) were visualized and quantified using Oil Red O. G6PDH activity was assessed before in vitro maturation (IVM), using the brilliant cresyl blue (BCB) test. IVM-control oocytes underwent IVM without BCB staining. The dCOCs and hCOCs had different patterns of LD spatial distribution, but similar amounts of lipid, although this tended towards being lower in hCOCs. Low G6PDH activity (BCB+) was observed in 74 %, 60 % and 24 % (P < 0.01) of dCOCs, hCOCs, and lCOCs, respectively. Significantly more BCB+ /oocytes than BCB-/oocytes reached the metaphase II stage in all groups. The maturation rate of BCB+ /hCOCs was higher than that of IVM/hCOC-controls (40 % v.s. 20 %, P < 0.001), and was comparable to that of BCB+ /dCOCs (54 %; P > 0.05). lCOCs were the smallest (P < 0.01), contained fewer (P < 0.01) lipids than dCOCs or hCOCs, and displayed reduced maturational potential. Overall, LD content and distribution, as well as G6PDH activity, in cat oocytes were strongly associated with ooplasm morphology and oocyte maturational competence. Deeper understanding of the intrinsic properties of oocytes with different ooplasm morphology using the domestic cat model, may be particularly important in the context of the conservation of endangered felids.
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Glucosafosfato Deshidrogenasa , Oocitos , Animales , Gatos , Oocitos/fisiología , Glucosafosfato Deshidrogenasa/metabolismo , Femenino , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Células del Cúmulo/fisiología , Células del Cúmulo/metabolismo , Metabolismo de los Lípidos/fisiología , Gotas Lipídicas/metabolismo , LípidosRESUMEN
The present paper provides the first integrative assessment of the capacity of dietary, endogenous and other agents to induce hormetic dose responses in oocytes, their supportive cells such as granulosa cells, blastocyst formation and early stage embryo development with the goal of improving fertility and reproductive success. The analysis showed that numerous agents enhance oocyte maturation and blastocyst/embryonic development in an hormetic fashion. These findings indicate that numerous agents improve oocyte-related biological functioning under normal conditions as well as enhancing its capacity to prevent damage from numerous chemical toxins and related stressor agents, including heat and age-related processes in pre-post conditioning and concurrent exposures. The present assessment suggests that hormetic-based lifestyles and dietary interventions may offer the potential to enhance healthy reproductive performance with applications to animal husbandry and human biology. The present findings also significantly extend the generality of the hormesis dose response concept to multiple fundamental biological processes (i.e., oocyte maturation, fertilization and blastocyst/embryo development).
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Blastocisto , Desarrollo Embrionario , Hormesis , Oocitos , Oocitos/efectos de los fármacos , Oocitos/fisiología , Desarrollo Embrionario/efectos de los fármacos , Blastocisto/efectos de los fármacos , Blastocisto/fisiología , Humanos , Animales , FemeninoRESUMEN
Endoplasmic reticulum (ER) stress interferes with developmental processes in oocyte maturation and embryo development. Invitro growth (IVG) is associated with low developmental competence, and ER stress during IVG culture may play a role. Therefore, this study aimed to examine the effect of tauroursodeoxycholic acid (TUDCA), an ER stress inhibitor, on the IVG of bovine oocytes to understand the role of ER stress. Oocyte-granulosa cell complexes (OGCs) were collected from early antral follicles (1.5-1.8 mm) and allowed to grow in vitro for 5 days at 38.5 °C in a humidified atmosphere containing 5 % CO2. Basic growth culture medium was supplemented with TUDCA at various concentrations (0, 50, 100, 250, and 500 µM). After IVG, oocyte diameters were similar among groups, but the antrum formation rate tended to be higher in the TUDCA 100 µM group. The mRNA expression levels of ER stress-associated genes (PERK, ATF6, ATF4, CHOP, BAX, IRE1, and XBP1) in OGCs were downregulated in the TUDCA 100 µM group than those in the control group. Moreover, the TUDCA 100 µM group exhibited reduced ROS production with higher GSH levels and improved in vitro-grown oocyte maturation compared with those in the control group. In contrast, no difference in the developmental competence of embryos following invitro fertilization was observed between the control and TUDCA 100 µM groups. These results indicate that ER stress could impair IVG and subsequent maturation rate of bovine oocytes, and TUDCA could alleviate these detrimental effects. These outcomes might improve the quality of oocytes in IVG culture in assisted reproductive technology.
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Estrés del Retículo Endoplásmico , Técnicas de Maduración In Vitro de los Oocitos , Oocitos , Ácido Tauroquenodesoxicólico , Animales , Bovinos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Oocitos/efectos de los fármacos , Oocitos/fisiología , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Técnicas de Maduración In Vitro de los Oocitos/métodos , Ácido Tauroquenodesoxicólico/farmacología , Femenino , Especies Reactivas de Oxígeno/metabolismo , Desarrollo Embrionario/efectos de los fármacosRESUMEN
In vivo, the temperature inside preovulatory follicles of cows is approximately 1 °C lower than rectal temperature. However, standard bovine oocyte in vitro maturation (IVM) protocols use 38.5 °C based on rectal temperature. This study evaluated the effect of reducing IVM temperature to 37.5 °C on the proteomic profile of oocytes compared to the routine 38.5 °C. Nuclear maturation rate and cumulus cell (CC) expansion (30 COCs per group, 21 replicates) were assessed by observing the first polar body and using a subjective scoring method (0-4). Total nitrite concentrations in the culture medium were measured using the Griess method. Differential proteomics was performed using LC-MS/MS on pooled oocyte samples (500 matured oocytes per group, three replicates), followed by gene ontology enrichment, protein-protein interaction, and putative miRNA target analyses. No significant differences were observed between the groups in nuclear maturation, CC expansion, or nitrite concentration (P > 0.05). A total of 806 proteins were identified, with 7 up-regulated and 12 down-regulated in the treatment group compared to the control. Additionally, 12 proteins were unique to the control group, and 8 were unique to the treatment group. IVM at 37.5 °C resulted in the upregulation of proteins involved in protein folding and GTP binding, and the downregulation of enzymes with oxidoreductase activity and proteins involved in cytoskeletal fiber formation. Furthermore, 43 bovine miRNAs potentially regulating these genes (DES, HMOX2, KRT75, FARSA, IDH2, CARHSP1) were identified. We conclude that IVM of bovine oocytes at 37.5 °C induces significant proteomic changes without impacting nuclear maturation, cumulus cell expansion, or nitrite concentration in the IVM medium.
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Técnicas de Maduración In Vitro de los Oocitos , Oocitos , Proteómica , Animales , Bovinos , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Oocitos/fisiología , Femenino , Temperatura , ProteomaRESUMEN
The female reproductive system is essential to women's health, human reproduction and societal well-being. However, the clinical translation of traditional research models is restricted due to the uncertain effects and low efficiency. Emerging evidence shows that microfluidic chips provide valuable platforms for studying the female reproductive system, while no paper has ever comprehensively discussed the topic. Here, a total of 161 studies out of 14,669 records are identified in PubMed, Scopus, Web of Science, ScienceDirect and IEEE Xplore databases. Among these, 61 studies focus on oocytes, which further involves culture, cell surgeries (oocyte separation, rotation, enucleation, and denudation), evaluation and cryopreservation. Forty studies investigate embryo manipulation via microfluidic chips, covering in vitro fertilization, cryopreservation and functional evaluation. Forty-six studies reconstitute both the physiological and pathological statuses of in vivo organs, mostly involved in placenta and fetal membrane research. Fourteen studies perform drug screening and toxicity testing. In this review, we summarize the current application of microfluidic chips in studying the female reproductive system, the advancements in materials and methods, and discuss the future challenges. The present evidence suggests that microfluidic chips-assisted reproductive system reconstruction is promising and more studies are urgently needed.
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Dispositivos Laboratorio en un Chip , Femenino , Humanos , Animales , Microfluídica/métodos , Oocitos/fisiología , Criopreservación/métodos , Reproducción/fisiología , Embarazo , Técnicas Reproductivas Asistidas , Genitales Femeninos/fisiologíaRESUMEN
Objectives of the current study were to examine the effects of exogenous expression of PGC-1α, which is a transcription factor responsive for controlling mitochondrial DNA (mtDNA) replication, mitochondria quantity control, mitochondrial biogenesis, and reactive oxygen species (ROS) maintenance, in porcine oocytes during in-vitro maturation (IVM) on the developmental competence, as well as mitochondrial quantity and function. Exogenous over-expression of PGC-1α by injection of the mRNA construct into oocytes 20 h after the start of IVM culture significantly increased the copy number of mtDNA in the oocytes, but reduced the incidences of oocytes matured to the metaphase-II stage after the IVM culture for totally 44 h and completely suppressed the early development in vitro to the blastocyst stage following parthenogenetic activation. The exogenous expression of PGC-1α also significantly induced spindle defects and chromosome misalignments. Furthermore, markedly higher ROS levels were observed in the PGC-1α-overexpressed mature oocytes, whereas mRNA level of SOD1, encoded for a ROS scavenging enzyme, was decreased. These results conclude that forced expression of PGC-1α successfully increase mtDNA copy number but led to increased ROS production, evidently by downregulation of SOD1 gene expression, inducement of spindle aberration/chromosomal misalignment, and consequently reduction in the meiotic and developmental competences of porcine oocytes.
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Técnicas de Maduración In Vitro de los Oocitos , Oocitos , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Animales , Femenino , ADN Mitocondrial/genética , ADN Mitocondrial/metabolismo , Desarrollo Embrionario , Regulación del Desarrollo de la Expresión Génica , Técnicas de Maduración In Vitro de los Oocitos/veterinaria , Oocitos/metabolismo , Oocitos/fisiología , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/genética , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/metabolismo , Especies Reactivas de Oxígeno/metabolismo , PorcinosRESUMEN
Global warming poses significant challenges to the fertility of tropical dairy cattle. One promising approach to mitigate heat stress effects on reproductive function and reduce the carbon footprint is the use of integrated livestock-forest (ILF) systems. The aim of this study was to investigate the effects of two different systems, namely Full Sun (FS) and ILF, on maternal hyperthermia and oocyte quality of Holstein and Girolando heifers during the tropical summer season. The temperature-humidity index (THI) data revealed intense heat stress during the experiment. Both the system (P<0.01) and the breed (P<0.01) factors had a significant impact on vaginal temperature, being hyperthermia more pronounced in the FS system and in the Holstein breed. Over the five time points collected at a 33-day interval, we observed distinct patterns for ILF (P=0.65) and FS (P<0.001) systems, suggesting an adaptive response in animals kept in FS systems. Furthermore, oocyte quality assessment revealed an effect of the system for oocyte diameter (P<0.001) and levels of IGFBP2 (P<0.001), and caspase 3 levels showed a decrease in ILF compared to FS for both Holstein (P<0.001) and Girolando (P<0.001) breeds. Collectively, these parameters indicate that oocyte quality during the summer months was superior in animals maintained in the ILF system. In conclusion, the ILF system demonstrated promising results in attenuating maternal hyperthermia and mitigating its effects on oocyte quality. Additionally, our observations suggest that animals in the FS system may exhibit an adaptive response to heat stress.
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Respuesta al Choque Térmico , Oocitos , Animales , Bovinos/fisiología , Oocitos/fisiología , Femenino , Respuesta al Choque Térmico/fisiología , Trastornos de Estrés por Calor/veterinaria , Crianza de Animales Domésticos/métodos , Calor , Enfermedades de los Bovinos/prevención & controlRESUMEN
Introduction: the availability of oocytes is fundamental to in vitro fertilization (IVF). The factors associated with optimal or suboptimal oocyte recovery rates (ORR) in low-resource settings are not well known. This study aimed to determine the factors associated with ORR by comparing demographic and IVF cycle data of women undergoing IVF in our Centre. Methods: this was a prospective study of 110 infertile women undergoing IVF at Nisa Premier Hospital, Abuja Nigeria, from October 2020 to September 2021. All women had reached the stage of oocyte retrieval or further, after receiving ovarian stimulation with our routine protocols. Treatment was monitored by serial transvaginal ultrasonography. The oocyte retrieval procedures were performed under conscious sedation, 36 hours after the ovulatory trigger. Optimal ORR was when eggs were obtained from at least 80% of follicles punctured. Sub-optimal ORR was when it was less than 80%. Data analyses utilized SPSS statistical software and a p-value of < 0.05 was considered significant. Results: the mean age of all women was 34.1±4.9 years. Sixty-nine women (62.7%) had sub-optimal ORR while 41 (37.3%) had optimal ORR. Six women (5.5%) had no oocytes retrieved. Significantly more women with sub-optimal ORR were obese (70.6 vs 29.4%) and had higher follicle-stimulating hormone (FSH) levels (8.11 vs 6.34 miu/ml), p-value- 0.039. Women with sub-optimal ORR had higher mean prolactin levels (17.10 ± 13.93 miu/ml) than women with optimal ORR 11.43 ± 6.65 miu/ml), p-value- 0.019). Significantly more oocytes (5.99 vs 10.37, p-value 0.001), and MII oocytes (5.78 vs 7.56, p-value 0.035) were retrieved in women with optimal than sub-optimal ORR. The duration of stimulation, total amounts of gonadotropins administered, and fertilized oocytes were not significantly different among both groups (p-value >0.05). Conclusion: this study has shown the factors associated with ORR in our setting to be basal FSH, prolactin, and obesity.
Asunto(s)
Fertilización In Vitro , Infertilidad Femenina , Recuperación del Oocito , Oocitos , Inducción de la Ovulación , Humanos , Femenino , Adulto , Recuperación del Oocito/métodos , Fertilización In Vitro/métodos , Estudios Prospectivos , Nigeria , Inducción de la Ovulación/métodos , Infertilidad Femenina/terapia , Oocitos/fisiología , Embarazo , ObesidadRESUMEN
Muscarinic receptors are G protein-coupled receptors (GPCRs) that play a role in various physiological functions. Previous studies have shown that these receptors, along with other GPCRs, are voltage-sensitive; both their affinity toward agonists and their activation are regulated by membrane potential. To our knowledge, whether the effect of antagonists on these receptors is voltage-dependent has not yet been studied. In this study, we used Xenopus oocytes expressing the M2 muscarinic receptor (M2R) to investigate this question. Our results indicate that the potencies of two M2R antagonists, atropine and scopolamine, are voltage-dependent; they are more effective at resting potential than under depolarization. In contrast, the M2R antagonist AF-DX 386 did not exhibit voltage-dependent potency.Furthermore, we discovered that the voltage dependence of M2R activation by acetylcholine remains unchanged in the presence of two allosteric modulators, the negative modulator gallamine and the positive modulator LY2119620. These findings enhance our understanding of GPCRs' voltage dependence and may have pharmacological implications.
Asunto(s)
Antagonistas Muscarínicos , Oocitos , Receptor Muscarínico M2 , Xenopus laevis , Animales , Receptor Muscarínico M2/antagonistas & inhibidores , Receptor Muscarínico M2/metabolismo , Receptor Muscarínico M2/agonistas , Regulación Alostérica/efectos de los fármacos , Regulación Alostérica/fisiología , Antagonistas Muscarínicos/farmacología , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Oocitos/fisiología , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Atropina/farmacología , Escopolamina/farmacología , Acetilcolina/metabolismo , Acetilcolina/farmacología , Femenino , Sulfonamidas , TiadiazolesRESUMEN
Ovum pick-up (OPU) by transvaginal ultrasound guided follicle aspiration in mares is a common assisted reproductive technique used for oocyte recovery and in vitro production of horse embryos. There has been relatively little research into the factors influencing oocyte recovery in OPU from live mares. The objective of this study was to compare oocyte recovery and morphology of ultrasound-guided follicle puncture and aspiration in live mares and in postmortem excised ovaries, in order to validate an experimental model for research purposes of the efficiency of OPU in mares. Data from OPU performed in 12 mares from a commercial program (follicle numbers, oocyte recovery and oocyte morphology) were compared to that obtained from ultrasound-guided follicle puncture of 13 postmortem excised ovaries from slaughtered mares processed within 2 h of slaughter. In both groups, the OPU was performed by the same operator using the same equipment and OPU technique. The recovered oocytes per aspirated follicle was higher (P < 0.05) in the postmortem group (105/166, 63.2 %) than in live mares (138/261, 52.9 %). There was more (P < 0.05) expanded cumulus oocyte complexes in the postmortem than in the live mares (18 % vs. 2.9 %). Several oocytes (5 oocytes from 81 aspirated follicles) were found in the leaked fluid which overflowed during follicle flushing of postmortem ovaries. In conclusion, the higher recovery rate obtained in the excised ovaries and the finding of oocytes in the leaked fluid during OPU, suggests that there is still room for improvement in the in vivo OPU technique. Utilizing postmortem excised ovaries could offer an alternative for further research into factors affecting oocyte recovery and oocyte leakage during OPU procedures.
Asunto(s)
Recuperación del Oocito , Oocitos , Folículo Ovárico , Animales , Femenino , Caballos/fisiología , Recuperación del Oocito/veterinaria , Recuperación del Oocito/métodos , Oocitos/fisiología , Folículo Ovárico/diagnóstico por imagen , Ovario/diagnóstico por imagen , Ultrasonografía/veterinaria , Ultrasonografía/métodos , Recolección de Tejidos y Órganos/veterinaria , Recolección de Tejidos y Órganos/métodosRESUMEN
Oocyte selection is a crucial step of assisted reproductive treatment. The most common approach relies on the embryologist experience which is inevitably prone to human error. One potential approach could be the use of an electrical-based approach as an ameliorative alternative. Here, we developed a simple electrical microsensor to characterize mouse oocytes. The sensor is designed similarly to embryo culture dishes and is familiar to embryologists. Different microelectrode models were simulated for oocyte cells and a more sensitive model was determined. The final microsensor was fabricated. A differential measuring technique was proposed based on the cell presence/absence. We predicted oocyte quality by using three electrical characteristics, oocyte radii, and zona thicknesses, and also these predictions were compared with an embryologist evaluation. The evaluation of the oocyte membrane capacitance, as an electrophysiological characteristic, was found to be a more reliable method for predicting oocytes with fertilization and blastocyst formation success competence. It achieved 94% and 58% prediction accuracies, respectively, surpassing other methods and yielding lower errors. This groundbreaking research represents the first of its kind in this field and we hope that this will be a step towards improving the accuracy of the treatment.