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Keywords = ovine oocytes

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14 pages, 939 KiB  
Review
Revisiting Male Fertility in Livestock: The Case of Bull Sperm RNA
by Rene A. Ramírez-Sosa, Francisco J. Jahuey-Martínez, Monserrath Felix-Portillo and José A. Martínez-Quintana
Biology 2025, 14(8), 969; https://doi.org/10.3390/biology14080969 (registering DOI) - 1 Aug 2025
Viewed by 282
Abstract
To achieve the goals of productivity and sustainability across diverse livestock systems, reproductive factors play a pivotal role. Historically, reproductive research has primarily focused on females, as they are responsible for maintaining pregnancy and delivering offspring following oocyte fertilization. However, since the early [...] Read more.
To achieve the goals of productivity and sustainability across diverse livestock systems, reproductive factors play a pivotal role. Historically, reproductive research has primarily focused on females, as they are responsible for maintaining pregnancy and delivering offspring following oocyte fertilization. However, since the early 2000s, the biological significance of sperm RNAs has been increasingly recognized in various livestock species. These RNAs contribute both genetically and epigenetically at the time of fertilization and during early embryonic development. Multiple types of sperm RNA have been identified in bovine, porcine, ovine, buffalo, and caprine spermatozoa. Notably, transcriptomic profiling has shown potential to differentiate between high- and low-fertility males, even when conventional semen quality values appear normal in both groups. This opens the possibility for more accurate identification of highly fertile sires. Nevertheless, a definitive marker or set of markers has yet to be established, likely due to the transcriptome’s sensitivity to environmental conditions and to the variability in evaluation methodologies. Therefore, global scientific efforts should aim to establish standardized, robust protocols, as sperm RNA represents a promising avenue for enhancing the sustainability of animal production systems. Full article
(This article belongs to the Special Issue The Biology of Animal Reproduction)
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26 pages, 17371 KiB  
Article
Transcriptomic Profile of Early Antral Follicles: Predictive Somatic Gene Markers of Oocyte Maturation Outcome
by Alessia Peserico, Barbara Barboni, Chiara Camerano Spelta Rapini, Chiara Di Berardino, Giulia Capacchietti, Angelo Canciello, Fani Konstantinidou, Marisa Donato, Liborio Stuppia and Valentina Gatta
Cells 2025, 14(10), 704; https://doi.org/10.3390/cells14100704 - 12 May 2025
Viewed by 733
Abstract
Early antral follicles (EAfs) offer oocyte potential in Assisted Reproductive Technology (ART), but most fail to mature under current in vitro maturation (IVM) protocols. This study examines transcriptomic profiles of the follicular wall (FW) compartment during IVM in ovine EAfs using a 3D [...] Read more.
Early antral follicles (EAfs) offer oocyte potential in Assisted Reproductive Technology (ART), but most fail to mature under current in vitro maturation (IVM) protocols. This study examines transcriptomic profiles of the follicular wall (FW) compartment during IVM in ovine EAfs using a 3D follicle-enclosed oocyte (FEO) culture to identify somatic gene markers predicting oocyte maturation success. Differentially expressed genes (DEGs) were identified across three comparisons: pre- vs. post-hCG in FW enclosing mature/fertilizable (1) or immature (2) oocytes, and post-hCG between FW supporting successful vs. failed maturation (3). Network analysis highlighted key modulated and HUB genes. Two DEG categories emerged: genes regulating meiosis resumption and genes defining follicular signatures linked to oocyte competence. Meiosis resumption involved ECM remodeling, hypoxia, and relaxin signaling activation, while proliferative and metabolic pathways were downregulated. MMP13 and EGFR regulated the ECM pathway, working for meiosis resumption, while TGFB1 predicted failure. Oocyte competence involves ECM activation and the suppression of stress and cell cycle pathways, with ITIH4 being conducive to central HUB tuning inflammation and angiogenesis-dependent maturation. This study reveals molecular mechanisms behind follicle maturation, identifying transcriptomic signatures for FW releasing mature/fertilizable and incompetent oocytes. It confirms known biomarkers and uncovers new regulators, offering tools to assess follicle quality, improve IVF–oocyte selection, and enhance fertility preservation. Full article
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18 pages, 7425 KiB  
Article
Exploration of Conditions for the Scaled Application of Laparoscopic Ovum Pick-Up in Sheep and Comparison of Follicular Development Differences Among Breeds
by Dongxu Li, Xiangli Wu, Ying Chen, Yangsheng Wu, Gulimire Abudureyimu, Hongyang Liang, Xiuling Ma, Wei Zhang, Liqin Wang and Jiapeng Lin
Int. J. Mol. Sci. 2025, 26(5), 1989; https://doi.org/10.3390/ijms26051989 - 25 Feb 2025
Viewed by 725
Abstract
In small ruminants, laparotomy for ovarian exploration followed by oocyte collection has been progressively replaced by laparoscopic puncture of follicles, which has become an important method for obtaining oocytes in vivo. However, the superovulation protocols and collection frequency used for laparoscopic ovum pick-up [...] Read more.
In small ruminants, laparotomy for ovarian exploration followed by oocyte collection has been progressively replaced by laparoscopic puncture of follicles, which has become an important method for obtaining oocytes in vivo. However, the superovulation protocols and collection frequency used for laparoscopic ovum pick-up (LOPU) in sheep still require further investigation. This study explored the factors influencing LOPU efficiency in sheep, including Controlled Internal Drug Release (CIDR) for estrus synchronization, FSH source and dose, and recovery intervals. The optimal superovulation protocol (using the CIDR device, a total of 16 mg of long-acting recombinant ovine FSH (LR-FSH) administered in two doses, and a one-month interval between LOPU sessions) was subsequently identified. Ovarian follicles were collected via LOPU from Hu sheep and Altay sheep for transcriptomic and metabolomic sequencing to explore interbreed differences in follicular development. The results indicated that LOPU efficiency was significantly higher in the CIDR group (p < 0.05) and with a 30-day recovery interval (p < 0.05). No significant differences in LOPU efficiency were observed between FSH sources or hormone doses. Furthermore, Hu sheep exhibited significantly higher LOPU efficiency and more antral follicles than Altay sheep. Transcriptomic analysis of follicular contents and metabolomic profiling of follicular fluid revealed that differentially expressed genes and metabolites were primarily enriched in pathways related to steroidogenesis, amino acid metabolism, and fatty acid metabolism. This study provides an optimized treatment protocol to enhance LOPU efficiency and integrates multi-omics analyses to elucidate the molecular mechanisms underlying follicular development differences among various breeds. Full article
(This article belongs to the Special Issue Molecular Research on Embryo Developmental Potential)
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12 pages, 1774 KiB  
Article
Study on the Technology of Laparoscopic Ovum Pick-Up and In Vitro Embryo Production in Chongming Goats
by Xiangli Wu, Dongxu Li, Ying Chen, Yangsheng Wu, Gulimire Abudureyimu, Wei Zhang, Kelu Deng, Zhen Huang, Jiapeng Lin and Liqin Wang
Biology 2024, 13(9), 699; https://doi.org/10.3390/biology13090699 - 6 Sep 2024
Cited by 2 | Viewed by 1902
Abstract
The application of laparoscopic ovum pick-up (LOPU) and in vitro production of embryos (IVEP) technologies has opened up a new path for purebred breeding and breed improvement in goats. However, due to the complexity of the procedures and multiple influencing factors, these technologies [...] Read more.
The application of laparoscopic ovum pick-up (LOPU) and in vitro production of embryos (IVEP) technologies has opened up a new path for purebred breeding and breed improvement in goats. However, due to the complexity of the procedures and multiple influencing factors, these technologies have not been widely adopted in goat production. This study explores factors affecting the efficiency of LOPU in goats by comparing the use of controlled internal drug release (CIDR) for estrus synchronization, conventional FSH versus long-acting recombinant ovine FSH (R-FSH) for superovulation, and the timing of LOPU at 48 h, 60 h, and 72 h of follicular development. The metrics evaluated included the recovery rate of cumulus–oocyte complexes (COCs), the average number of ovarian follicles, the average number of COCs, and the average number of available COCs. The results demonstrated that the efficiency of LOPU was significantly higher with two doses of R-FSH compared to the conventional FSH superovulation protocol and the control group (p < 0.05), with two doses of R-FSH providing a higher LOPU efficiency than one dose (p < 0.05). Using CIDR for estrus synchronization showed no significant difference in LOPU efficiency compared to the non-CIDR group. Similarly, the efficiency of LOPU showed no significant difference between the hormone treatments at 48 h, 60 h, and 72 h. By exploring and optimizing the factors influencing LOPU, we ultimately established a LOPU technology system for goats that meets the production needs of small ruminants. Full article
(This article belongs to the Special Issue Reproductive Management of Goats and Sheep)
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12 pages, 16877 KiB  
Article
Effects of Exogenous Regulation of PPARγ on Ovine Oocyte Maturation and Embryonic Development In Vitro
by Hengbin Yu, Yue Zhang, Yidan Zhang, Shuaitong Chen, Zhenghang Li, Wenhui Pi, Weibin Zeng and Guangdong Hu
Vet. Sci. 2024, 11(9), 397; https://doi.org/10.3390/vetsci11090397 - 28 Aug 2024
Viewed by 2888
Abstract
Lactating oocytes consume a lot of energy during maturation, a large part of which comes from lipid metabolism. PPARγ is a key regulator of lipid metabolism. In this study, rosiglitazone (RSG), an activator of PPARγ, was added to a mature medium to investigate [...] Read more.
Lactating oocytes consume a lot of energy during maturation, a large part of which comes from lipid metabolism. PPARγ is a key regulator of lipid metabolism. In this study, rosiglitazone (RSG), an activator of PPARγ, was added to a mature medium to investigate its effects on the levels of spindle and the chromosome arrangement, lipid deposition, reactive oxygen species (ROS), and glutathione (GSH) levels, oocyte secretion factors, apoptosis and lipid metabolism-related gene expression, and subsequent embryonic development during the maturation of sheep oocytes. The oocyte secretion factor affects gene expression related to apoptosis and lipid metabolism and subsequent embryonic development. The results showed that the proportion of spindle and normal chromosome arrangements increased in the 5 μM RSG treatment group, the lipid content increased after cell maturation, the ROS level decreased, and the GSH level increased. The expressions of oocyte secretion factor (GDF9 and BMP15), anti-apoptosis gene (BCL2), and lipid metabolism-related genes (ACAA1, CPT1A, PLIN2) were increased in the 5 μM treatment group. Finally, the development of blastocysts was examined. After the oocytes were treated with 5 μM RSG, the blastocyst rate and the gene expression of the totipotency gene (OCT4) were increased. It was concluded that increasing PPARγ activity during ovine oocyte maturation could promote lipid metabolism, reduce oxidative stress, and improve the ovine oocyte maturation rate and subsequent embryo development. Full article
(This article belongs to the Special Issue Mechanisms of Reproductive Control in the Animals)
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21 pages, 6951 KiB  
Article
Screening and Mechanism Study of Three Antagonistic Drugs, Oxysophoridine, Rutin, and Phellodendrine, against Zearalenone-Induced Reproductive Toxicity in Ovine Oocytes
by Zongshuai Li, Tian Ma, Yali Liu, Wanruo Liu, Xingxu Zhao, Gaiping Zhang, Jianlin Wang and Yong Zhang
Antioxidants 2024, 13(6), 752; https://doi.org/10.3390/antiox13060752 - 20 Jun 2024
Cited by 2 | Viewed by 1731
Abstract
Zearalenone (ZEN) is a common fungal toxin with reproductive toxicity in various grains. It poses a serious threat to ovine and other animal husbandry industries, as well as human reproductive health. Therefore, investigating the mechanism of toxicity and screening antagonistic drugs are of [...] Read more.
Zearalenone (ZEN) is a common fungal toxin with reproductive toxicity in various grains. It poses a serious threat to ovine and other animal husbandry industries, as well as human reproductive health. Therefore, investigating the mechanism of toxicity and screening antagonistic drugs are of great importance. In this study, based on the natural compound library and previous Smart-seq2 results, antioxidant and anti-apoptotic drugs were selected for screening as potential antagonistic drugs. Three natural plant compounds (oxysophoridine, rutin, and phellodendrine) were screened for their ability to counteract the reproductive toxicity of ZEN on ovine oocytes in vitro using quantitative polymerase chain reaction (qPCR) and reactive oxygen species detection. The compounds exhibited varying pharmacological effects, notably impacting the expression of antioxidant (GPX, SOD1, and SOD2), autophagic (ATG3, ULK2, and LC3), and apoptotic (CAS3, CAS8, and CAS9) genes. Oxysophoridine promoted GPX, SOD1, ULK2, and LC3 expression, while inhibiting CAS3 and CAS8 expression. Rutin promoted SOD2 and ATG3 expression, and inhibited CAS3 and CAS9 expression. Phellodendrine promoted SOD2 and ATG3 expression, and inhibited CAS9 expression. However, all compounds promoted the expression of genes related to cell cycle, spindle checkpoint, oocyte maturation, and cumulus expansion factors. Although the three drugs had different regulatory mechanisms in enhancing antioxidant capacity, enhancing autophagy, and inhibiting cell apoptosis, they all maintained a stable intracellular environment and a normal cell cycle, promoted oocyte maturation and release of cumulus expansion factors, and, ultimately, counteracted ZEN reproductive toxicity to promote the in vitro maturation of ovine oocytes. This study identified three drugs that antagonize the reproductive toxicity of ZEN on ovine oocytes, and compared their mechanisms of action, providing data support and a theoretical basis for their subsequent application in the ovine breeding industry, reducing losses in the breeding industry, screening of ZEN reproductive toxicity antagonists and various toxin antagonists, improving the study of ZEN reproductive toxicity mechanisms, and even protection of human reproductive health. Full article
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15 pages, 29915 KiB  
Article
Smart-seq2 Technology Reveals a Novel Mechanism That Zearalenone Inhibits the In Vitro Maturation of Ovine Oocytes by Influencing TNFAIP6 Expression
by Zongshuai Li, Yali Liu, Tian Ma, Chen Lv, Yina Li, Hongwei Duan, Xingxu Zhao, Jianlin Wang and Yong Zhang
Toxins 2023, 15(10), 617; https://doi.org/10.3390/toxins15100617 - 17 Oct 2023
Cited by 4 | Viewed by 2400
Abstract
Zearalenone (ZEN), a non-steroidal estrogenic fungal toxin widely present in forage, food, and their ingredients, poses a serious threat to animal and human reproductive health. ZEN also threatens ovine, a major source of human food and breeding stock. However, the mechanisms underlying the [...] Read more.
Zearalenone (ZEN), a non-steroidal estrogenic fungal toxin widely present in forage, food, and their ingredients, poses a serious threat to animal and human reproductive health. ZEN also threatens ovine, a major source of human food and breeding stock. However, the mechanisms underlying the impact of ZEN on the in vitro maturation (IVM) of ovine oocytes remain unclear. This study aimed to elucidate these mechanisms using the Smart-seq2 technology. A total of 146 differentially expressed genes were obtained, using Smart-seq2, from sheep oocytes cultured in vitro after ZEN treatment. ZEN treatment inhibited RUNX2 and SPP1 expression in the PI3K signaling pathway, leading to the downregulation of THBS1 and ultimately the downregulation of TNFAIP6; ZEN can also decrease TNFAIP6 by reducing PTPRC and ITGAM. Both inhibit in vitro maturation of ovine oocytes and proliferation of cumulus cells by downregulating TNFAIP6. These findings provide data and a theoretical basis for elucidating ZEN’s toxicity mechanisms, screening therapeutic drugs, and reducing ZEN-related losses in the ovine industry. Full article
(This article belongs to the Section Mycotoxins)
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18 pages, 3584 KiB  
Article
New Strategies for Conservation of Gentile di Puglia Sheep Breed, an Autochthonous Capital of Millennial Tradition in Southern Italy
by Letizia Temerario, Davide Monaco, Antonella Mastrorocco, Nicola Antonio Martino, Sándor Cseh, Giovanni Michele Lacalandra, Elena Ciani and Maria Elena Dell’Aquila
Animals 2023, 13(14), 2371; https://doi.org/10.3390/ani13142371 - 20 Jul 2023
Cited by 6 | Viewed by 2306
Abstract
Gentile di Puglia (GdP) is an autochthonous sheep breed of Southern Italy included among ovine breeds threatened by genetic erosion and extinction risk, which have been given attention by local and international institutions, thus emphasizing the need for germplasm conservation actions. In the [...] Read more.
Gentile di Puglia (GdP) is an autochthonous sheep breed of Southern Italy included among ovine breeds threatened by genetic erosion and extinction risk, which have been given attention by local and international institutions, thus emphasizing the need for germplasm conservation actions. In the present study, two assisted reproduction approaches, finalized for GdP conservation, were performed: (1) on-farm reproductive efficiency evaluation, expressed as pregnancy rate (PR), twin pregnancy rate (tPR), and body condition score (BCS), for three consecutive breeding cycles and (2) pre-pubertal lambs’ immature cumulus–oocyte complex (COC) retrieval, vitrification, in vitro maturation (IVM), and assessment of meiotic stage and bioenergetic-oxidative status compared with those of other Italian and European commercial breeds. PR and tPR were progressively reduced over time. In all clinical examination times, BCS was significantly lower in nonpregnant ewes compared with pregnant ones. Fresh GdP pre-pubertal lamb COCs achieved meiotic maturation and showed healthy bioenergetic–oxidative status after IVM. Vitrification reduced the oocyte maturation rate in all groups. However, mature oocytes retained their cytoplasmic maturity, expressed as a mitochondria distribution pattern and activity, indicating promising developmental competence. In conclusion, clinical- and biotechnological-assisted reproduction approaches can support conservation strategies of GdP and other local sheep breeds in Southern Italy. Full article
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24 pages, 4024 KiB  
Article
IVM Advances for Early Antral Follicle-Enclosed Oocytes Coupling Reproductive Tissue Engineering to Inductive Influences of Human Chorionic Gonadotropin and Ovarian Surface Epithelium Coculture
by Alessia Peserico, Chiara Di Berardino, Giulia Capacchietti, Chiara Camerano Spelta Rapini, Liliana Liverani, Aldo Roberto Boccaccini, Valentina Russo, Annunziata Mauro and Barbara Barboni
Int. J. Mol. Sci. 2023, 24(7), 6626; https://doi.org/10.3390/ijms24076626 - 1 Apr 2023
Cited by 7 | Viewed by 2642
Abstract
In vitro maturation (IVM) is not a routine assisted reproductive technology (ART) for oocytes collected from early antral (EA) follicles, a large source of potentially available gametes. Despite substantial improvements in IVM in the past decade, the outcomes remain low for EA-derived oocytes [...] Read more.
In vitro maturation (IVM) is not a routine assisted reproductive technology (ART) for oocytes collected from early antral (EA) follicles, a large source of potentially available gametes. Despite substantial improvements in IVM in the past decade, the outcomes remain low for EA-derived oocytes due to their reduced developmental competences. To optimize IVM for ovine EA-derived oocytes, a three-dimensional (3D) scaffold-mediated follicle-enclosed oocytes (FEO) system was compared with a validated cumulus-oocyte complex (COC) protocol. Gonadotropin stimulation (eCG and/or hCG) and/or somatic cell coculture (ovarian vs. extraovarian-cell source) were supplied to both systems. The maturation rate and parthenogenetic activation were significantly improved by combining hCG stimulation with ovarian surface epithelium (OSE) cells coculture exclusively on the FEO system. Based on the data, the paracrine factors released specifically from OSE enhanced the hCG-triggering of oocyte maturation mechanisms by acting through the mural compartment (positive effect on FEO and not on COC) by stimulating the EGFR signaling. Overall, the FEO system performed on a developed reproductive scaffold proved feasible and reliable in promoting a synergic cytoplasmatic and nuclear maturation, offering a novel cultural strategy to widen the availability of mature gametes for ART. Full article
(This article belongs to the Special Issue Gonadotropin Cell Transduction Mechanisms 2.0)
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26 pages, 4253 KiB  
Review
Motility Assessment of Ram Spermatozoa
by Madeleine Van de Hoek, Jessica P. Rickard and Simon P. de Graaf
Biology 2022, 11(12), 1715; https://doi.org/10.3390/biology11121715 - 26 Nov 2022
Cited by 32 | Viewed by 6770
Abstract
For successful fertilisation to occur, spermatozoa need to successfully migrate through the female reproductive tract and penetrate the oocyte. Predictably, poor sperm motility has been associated with low rates of fertilisation in many mammalian species, including the ram. As such, motility is one [...] Read more.
For successful fertilisation to occur, spermatozoa need to successfully migrate through the female reproductive tract and penetrate the oocyte. Predictably, poor sperm motility has been associated with low rates of fertilisation in many mammalian species, including the ram. As such, motility is one of the most important parameters used for in vitro evaluation of ram sperm quality and function. This review aims to outline the mechanical and energetic processes which underpin sperm motility, describe changes in motility which occur as a result of differences in sperm structure and the surrounding microenvironment, and assess the effectiveness of the various methods used to assess sperm motility in rams. Methods of subjective motility estimation are convenient, inexpensive methods widely used in the livestock industries, however, the subjective nature of these methods can make them unreliable. Computer-assisted sperm analysis (CASA) technology accurately and objectively measures sperm motility via two-dimensional tracing of sperm head motion, making it a popular method for sperm quality assurance in domesticated animal production laboratories. Newly developed methods of motility assessment including flagellar tracing, three-dimensional sperm tracing, in vivo motility assessment, and molecular assays which quantify motility-associated biomarkers, enable analysis of a new range of sperm motion parameters with the potential to reveal new mechanistic insights and improve ram semen assessment. Experimental application of these technologies is required to fully understand their potential to improve semen quality assessment and prediction of reproductive success in ovine artificial breeding programs. Full article
(This article belongs to the Section Developmental and Reproductive Biology)
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25 pages, 5013 KiB  
Article
When Electrospun Fiber Support Matters: In Vitro Ovine Long-Term Folliculogenesis on Poly (Epsilon Caprolactone) (PCL)-Patterned Fibers
by Chiara Di Berardino, Liliana Liverani, Alessia Peserico, Giulia Capacchietti, Valentina Russo, Nicola Bernabò, Umberto Tosi, Aldo Roberto Boccaccini and Barbara Barboni
Cells 2022, 11(12), 1968; https://doi.org/10.3390/cells11121968 - 19 Jun 2022
Cited by 12 | Viewed by 2800
Abstract
Current assisted reproduction technologies (ART) are insufficient to cover the slice of the population needing to restore fertility, as well as to amplify the reproductive performance of domestic animals or endangered species. The design of dedicated reproductive scaffolds has opened the possibility to [...] Read more.
Current assisted reproduction technologies (ART) are insufficient to cover the slice of the population needing to restore fertility, as well as to amplify the reproductive performance of domestic animals or endangered species. The design of dedicated reproductive scaffolds has opened the possibility to better recapitulate the reproductive 3D ovarian environment, thus potentially innovating in vitro folliculogenesis (ivF) techniques. To this aim, the present research has been designed to compare ovine preantral follicles in vitro culture on poly(epsilon-caprolactone) (PCL)-based electrospun scaffolds designed with different topology (Random vs. Patterned fibers) with a previously validated system. The ivF performances were assessed after 14 days under 3D-oil, Two-Step (7 days in 3D-oil and on scaffold), or One-Step PCL protocols (14 days on PCL-scaffold) by assessing morphological and functional outcomes. The results show that Two- and One-Step PCL ivF protocols, when performed on patterned scaffolds, were both able to support follicle growth, antrum formation, and the upregulation of follicle marker genes leading to a greater oocyte meiotic competence than in the 3D-oil system. In conclusion, the One-Step approach could be proposed as a practical and valid strategy to support a synergic follicle-oocyte in vitro development, providing an innovative tool to enhance the availability of matured gametes on an individual basis for ART purposes. Full article
(This article belongs to the Section Reproductive Cells and Development)
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22 pages, 3636 KiB  
Article
Equine Chorionic Gonadotropin as an Effective FSH Replacement for In Vitro Ovine Follicle and Oocyte Development
by Chiara Di Berardino, Alessia Peserico, Giulia Capacchietti, Martina Crociati, Maurizio Monaci, Umberto Tosi, Annunziata Mauro, Valentina Russo, Nicola Bernabò and Barbara Barboni
Int. J. Mol. Sci. 2021, 22(22), 12422; https://doi.org/10.3390/ijms222212422 - 17 Nov 2021
Cited by 11 | Viewed by 3748
Abstract
The use of assisted reproductive technologies (ART) still requires strategies through which to maximize individual fertility chances. In vitro folliculogenesis (ivF) may represent a valid option to convey the large source of immature oocytes in ART. Several efforts have been made [...] Read more.
The use of assisted reproductive technologies (ART) still requires strategies through which to maximize individual fertility chances. In vitro folliculogenesis (ivF) may represent a valid option to convey the large source of immature oocytes in ART. Several efforts have been made to set up ivF cultural protocols in medium-sized mammals, starting with the identification of the most suitable gonadotropic stimulus. In this study, Equine Chorionic Gonadotropin (eCG) is proposed as an alternative to Follicle Stimulating Hormone (FSH) based on its long superovulation use, trans-species validation, long half-life, and low costs. The use of 3D ivF on single-ovine preantral (PA) follicles allowed us to compare the hormonal effects and to validate their influence under two different cultural conditions. The use of eCG helped to stimulate the in vitro growth of ovine PA follicles by maximizing its influence under FBS-free medium. Higher performance of follicular growth, antrum formation, steroidogenic activity and gap junction marker expression were recorded. In addition, eCG, promoted a positive effect on the germinal compartment, leading to a higher incidence of meiotic competent oocytes. These findings should help to widen the use of eCG to ivF as a valid and largely available hormonal support enabling a synchronized in vitro follicle and oocyte development. Full article
(This article belongs to the Special Issue Gonadotropin Cell Transduction Mechanisms)
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14 pages, 2886 KiB  
Article
3D Liquid Marble Microbioreactors Support In Vitro Maturation of Prepubertal Ovine Oocytes and Affect Expression of Oocyte-Specific Factors
by Daniela Bebbere, Stefano Mario Nieddu, Federica Ariu, Davide Piras and Sergio Ledda
Biology 2021, 10(11), 1101; https://doi.org/10.3390/biology10111101 - 25 Oct 2021
Cited by 14 | Viewed by 2610
Abstract
In vitro oocyte maturation (IVM) is a well-established technique. Despite the high IVM rates obtained in most mammalian species, the developmental competence of IVM oocytes is suboptimal. The aim of this work was to evaluate the potential beneficial effects of a liquid marble [...] Read more.
In vitro oocyte maturation (IVM) is a well-established technique. Despite the high IVM rates obtained in most mammalian species, the developmental competence of IVM oocytes is suboptimal. The aim of this work was to evaluate the potential beneficial effects of a liquid marble microbioreactor (LM) as a 3D culture system to mature in vitro prepubertal ovine oocytes, as models of oocytes with intrinsic low competence. Cumulus–oocyte complexes of prepubertal sheep ovaries were in vitro matured in a LM system with hydrophobic fumed-silica-nanoparticles (LM group) or in standard conditions (4W control group). We evaluated: (a) maturation and (b) developmental rates following in vitro fertilization (IVF) and embryo culture; (c) expression of a panel of genes. LM and 4W groups showed similar IVM and IVF rates, while in vitro development to blastocyst stage approached significance (4W: 14.1% vs. LM: 28.3%; p = 0.066). The expression of GDF9, of enzymes involved in DNA methylation reprogramming and of the subcortical maternal complex was affected by the IVM system, while no difference was observed in terms of cell-stress-response. LM microbioreactors provide a suitable microenvironment to induce prepubertal sheep oocyte IVM and should be considered to enhance the developmental competence of oocytes with reduced potential also in other species, including humans. Full article
(This article belongs to the Section Developmental and Reproductive Biology)
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12 pages, 1399 KiB  
Article
Seasonal Effect on Developmental Competence, Oxidative Status and Tubulin Assessment of Prepubertal Ovine Oocyte
by Elisa Serra, Sergio Domenico Gadau, Giovanni Giuseppe Leoni, Salvatore Naitana and Sara Succu
Animals 2021, 11(7), 1886; https://doi.org/10.3390/ani11071886 - 24 Jun 2021
Cited by 2 | Viewed by 2315
Abstract
The reproductive seasonality of domestic animals is often manipulated in order to have more reproductive periods for commercial purposes related to the production of milk and meat. It is scientifically proven that such an alteration of the reproductive activity in sheep entails a [...] Read more.
The reproductive seasonality of domestic animals is often manipulated in order to have more reproductive periods for commercial purposes related to the production of milk and meat. It is scientifically proven that such an alteration of the reproductive activity in sheep entails a deterioration in oocyte quality, leading to an inability to generate embryos. Since oocytes obtained from prepubertal ewes can be incorporated into an in vitro embryo production system and considering that their quality is crucial to the success of in vitro procedures, the aim of this work was to investigate the effect of seasons on the quality of prepubertal ovine oocytes collected in autumn and spring. Ovaries were collected from a local slaughterhouse from 30–40-day-old suckling lambs during both seasons. Following 24 h of in vitro maturation, oocytes developmental competence, reactive oxygen species (ROS) intracellular levels, and mitochondrial activity were evaluated, and a tubulin assessment was performed. The results on embryo production, as a percentage of first divisions and number of blastocysts obtained, were significantly higher in oocytes collected in the spring. Mitochondrial activity in oocytes was higher, and ROS production significantly lower, in spring than in autumn. Tubulin PTMs (tyrosinated and acetylated α-tubulin) showed a higher immunoreactivity in oocytes collected in spring compared with autumn sampling. Our data showed that seasons may affect the developmental competence, energetic status, and tubulin assessment of oocytes recovered from prepubertal ewes. Therefore, special care should be taken when choosing the period of the year for prepuberal ovine oocytes collection aimed at in vitro embryo reproduction programs. Full article
(This article belongs to the Special Issue Reproductive Management)
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16 pages, 1762 KiB  
Article
The Roles of the miRNAome and Transcriptome in the Ovine Ovary Reveal Poor Efficiency in Juvenile Superovulation
by Xiaosheng Zhang, Chunxiao Dong, Jing Yang, Yihai Li, Jing Feng, Biao Wang, Jinlong Zhang and Xiaofei Guo
Animals 2021, 11(1), 239; https://doi.org/10.3390/ani11010239 - 19 Jan 2021
Cited by 12 | Viewed by 3899
Abstract
Juvenile superovulation can provide a wealth of oocyte material for embryo production, animal cloning, and genetic modification research, but embryos derived from juvenile oocytes show poor efficiency in subsequent developmental capacity. In order to reveal the formation mechanism of large numbers of follicles [...] Read more.
Juvenile superovulation can provide a wealth of oocyte material for embryo production, animal cloning, and genetic modification research, but embryos derived from juvenile oocytes show poor efficiency in subsequent developmental capacity. In order to reveal the formation mechanism of large numbers of follicles and poor oocyte quality in juvenile ovaries under superovulation treatment, differentially expressed microRNAs (miRNAs) and messenger RNAs (mRNAs) were characterized and investigated in the ovaries of lambs and adult sheep using high-throughput sequencing technology. The majority of differentially expressed miRNAs (337/358) were upregulated in lamb libraries. The expression levels of mRNAs related to hormone receptors (follicle-stimulating hormone receptor, FSHR; luteinizing hormone/choriogonadotropin receptor, LHCGR; estrogen receptor 1, ESR1), steroid hormone secretion (cytochrome P450 family 11 subfamily A member 1, CYP11A1; cytochrome P450 family 17 subfamily A member 1, CYP17A1; cytochrome P450 family 19 subfamily A member 1, CYP19A1), and oocyte quality (pentraxin 3, PTX3; BCL2 apoptosis regulator, BCL2; caspase 3, CASP3) were significantly different between the lamb and adult libraries. The miRNA aor-miR-143, which targets FSHR, was highly and differentially expressed, and PTX3 was predicted to be targeted by oar-miR-485-3p and oar-miR-377-3p in the ovine ovary. A considerable number of miRNAs were predicted to inhibit ESR1 expression in lamb ovaries. In conclusion, oar-miR-143 and FSHR molecules, among others, might regulate follicle formation, and oar-miR-485-3p, oar-miR-377-3p, and PTX3, among others, may be associated with oocyte quality. These identified miRNAs and mRNAs will be beneficial for the prediction of ovarian superovulation potential and screening of oocytes. Full article
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