Histone 3 lysine 9 acetylation is a biomarker of the effects of culture on zygotes

Acetylation of histone proteins is a major determinant of chromatin structure and function. Fertilisation triggers a round of chromatin remodelling that prepares the genome for the first round of transcription from the new embryonic genome. In this study we confirm that fertilisation leads to a marked progressive increase in the level of histone 3 lysine 9 acetylation in both the paternally and maternally derived genomes. The culture of zygotes in simple defined media caused a marked increase in the global level of acetylation and this affected the male pronucleus more than the female. The culture created a marked asymmetry in staining between the two pronuclei that was not readily detected in zygotes collected directly from the reproductive tract and was ameliorated to some extent by optimized culture media. The increased acetylation caused by culture resulted in increased transcription ofHspa1b, a marker of embryonic genome activation. Pharmacological analyses showed the hyperacetylation of H3K9 and the increased expression ofHspa1bcaused by culture were due to the altered net activity of a range of histone acetylases and deacetylases. The marked hyperacetylation of histone 3 lysine 9 caused by culture of zygotes may serve as an early biomarker for the effects of culture on the normal function of the embryo. The results also provide further evidence for an effect of the stresses associated with assisted reproductive technologies on the normal patterns of epigenetic reprogramming in the early embryo.

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History of oocyte and embryo metabolism

Reproduction Fertility and Development ◽

10.1071/rd14278 ◽

2015 ◽

Vol 27 (4) ◽

pp. 567 ◽

Cited By ~ 16

Author(s):

Henry J. Leese

Keyword(s):

Reproductive Tract ◽

Assisted Reproductive Technologies ◽

Culture Media ◽

Cell Signalling ◽

Reproductive Technologies ◽

Early Embryo ◽

Female Reproductive Tract ◽

1960S And 1970S ◽

The 1960S ◽

Embryo Metabolism

The basic pattern of metabolism in mammalian oocytes and early embryos was established in the 1960s and 1970s, largely in terms of the consumption of oxygen and the utilisation of nutrients present in culture media at the time, mainly glucose, pyruvate and lactate. The potential importance of endogenous fuels was also recognised but was largely ignored, only to be rediscovered quite recently. The 1980s and 1990s saw the arrival of a ‘new generation’ of culture media, characterised metabolically by the addition of amino acids, an initiative driven strongly by the need to improve embryo culture and selection methods in assisted reproductive technologies. This trend has continued alongside some basic metabolic studies and the general recognition of the importance of metabolism in all aspects of biology. A framework for future studies on oocyte and early embryo metabolism has been provided by: (1) the developmental origins of health and disease concept and recognition of the relationship between development, epigenetics and metabolism; (2) the need to understand cell signalling within, and between the cells of, the early embryo; and (3) the importance of identifying the mechanisms underlying dialogue between the oocyte and early embryo and the female reproductive tract.

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Extracellular Vesicles in the Oviduct: Progress, Challenges and Implications for the Reproductive Success

Bioengineering ◽

10.3390/bioengineering6020032 ◽

2019 ◽

Vol 6 (2) ◽

pp. 32 ◽

Cited By ~ 20

Author(s):

Almiñana ◽

Bauersachs

Keyword(s):

Extracellular Vesicles ◽

Reproductive Tract ◽

Assisted Reproductive Technologies ◽

Current Knowledge ◽

Cell Communication ◽

Reproductive Technologies ◽

Early Embryo ◽

Female Reproductive Tract ◽

Functional Aspects ◽

Reproductive Events

The oviduct is the anatomical part of the female reproductive tract where the early reproductive events take place, from gamete transport, fertilization and early embryo development to the delivery of a competent embryo to the uterus, which can implant and develop to term. The success of all these events rely upon a two-way dialogue between the oviduct (lining epithelium and secretions) and the gametes/embryo(s). Recently, extracellular vesicles (EVs) have been identified as major components of oviductal secretions and pointed to as mediators of the gamete/embryo-maternal interactions. EVs, comprising exosomes and microvesicles, have emerged as important agents of cell-to-cell communication by the transfer of biomolecules (i.e., mRNAs, miRNAs, proteins) that can modulate the activities of recipient cells. Here, we provide the current knowledge of EVs in the oviductal environment, from isolation to characterization, and a description of the EVs molecular content and associated functional aspects in different species. The potential role of oviductal EVs (oEVs) as modulators of gamete/embryo-oviduct interactions and their implications in the success of early reproductive events is addressed. Lastly, we discuss current challenges and future directions towards the potential application of oEVs as therapeutic vectors to improve pregnancy disorders, infertility problems and increase the success of assisted reproductive technologies.

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207 Female Reproductive Fluids and Epigenetics

Journal of Animal Science ◽

10.1093/jas/skab054.188 ◽

2021 ◽

Vol 99 (Supplement_1) ◽

pp. 114-114

Author(s):

Sebastian Canovas ◽

Raquel Romar ◽

Pilar Coy

Keyword(s):

Assisted Reproductive Technologies ◽

Culture Media ◽

Morphological Changes ◽

Adult Life ◽

Specific Effect ◽

Reproductive Technologies ◽

Early Embryo ◽

Vitro Fertilization

Abstract Physiological fertilization, and early embryo development, involves dramatic transcriptomic, epigenetic and morphological changes in a short temporal window. During this period gametes and early embryos are surrounded by reproductive fluids (oviductal and uterine), which contain nutrients, growth factors, hormones and extracellular vesicles acting as carriers of DNA, RNA, proteins and other factors with putative roles in intercellular communication. Under in vitro conditions, and in the absence of these fluids, embryos derived from Assisted Reproductive Technologies (ART) reveal transcriptional and epigenetic differences compared with in vivo embryos, which could result in long-term phenotypic consequences in adult life. Therefore, reproductive fluids supplementation in the culture medium offers an alternative to imitate physiological conditions and decrease these consequences. In vitro, oviductal fluid (OF) can modulate capacitation-associated events and sperm-zona pellucida interactions and contribute to the control of polyspermy in pigs. The use of in vitro fertilization media supplemented with reproductive fluids (Natur-IVF) improves embryo quality and blastocysts hatching ability. Moreover, Natur-IVF embryos show expression and methylation patterns closer to in vivo blastocysts. In cows, supplementation of culture media with reproductive fluids, or some isolated factors, improves blastocyst rate and survival after embryo transfer, and reverses the expression of some altered genes. However, considering the complexity of the oviductal and uterine fluids, it seems difficult that the use of just a few factors in isolation can reverse all undesired consequences of the IVP. On the other hand, sex-specific embryonic plasticity, as a consequence of the oviductal regulatory signals, have been proposed. Thus, we have analysed the sex-specific effect of supplementation with reproductive fluids in bovine embryos and data reveal sex-dependent impact in DNA methylation. All these results confirm that developmental programme can be modulated by reproductive fluids and it shows sex-specific effects. This strategy allows the possibility of minimizing undesired in vitro derived consequences.

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Reproductive fluids, used for the in vitro production of pig embryos, result in healthy offspring and avoid aberrant placental expression of PEG3 and LUM.

10.21203/rs.3.rs-42227/v2 ◽

2020 ◽

Author(s):

Evelynne Paris-Oller ◽

Sergio Navarro-Serna ◽

Cristina Soriano-Úbeda ◽

Jordana Sena Lopes ◽

Carmen Matas ◽

...

Keyword(s):

Reproductive Performance ◽

Assisted Reproductive Technologies ◽

Culture Media ◽

Reproductive Technologies ◽

In Vitro Production ◽

Aberrant Expression ◽

Low Efficiency ◽

The Impact

Abstract Background: In vitro embryo production (IVP) and embryo transfer (ET) are two very common assisted reproductive technologies (ART) in human and cattle. However, in pig, the combination of either procedures, or even their use separately, is still considered suboptimal due to the low efficiency of IVP plus the difficulty of performing ET in the long and contorted uterus of the sow. In addition, the potential impact of these two ART on the health of the offspring is unknown. We investigated here if the use of a modified IVP system, with natural reproductive fluids (RF) as supplements to the culture media, combined with a minimally invasive surgery to perform ET, affects the output of the own IVP system as well as the reproductive performance of the mother and placental molecular traits.Results: The blastocyst rates obtained by both in vitro systems, conventional (C-IVP) and modified (RF-IVP), were similar. Pregnancy and farrowing rates were also similar. However, when compared to in vivo control (artificial insemination, AI), litter sizes of both IVP groups were lower, while placental efficiency was higher in AI than in RF-IVP. Gene expression studies revealed aberrant expression levels for PEG3 and LUM in placental tissue for C-IVP group when compared to AI, but not for RF-IVP group.Conclusions: The use of reproductive fluids as additives for the culture media in pig IVP does not improve reproductive performance of recipient mothers but could mitigate the impact of artificial procedures in the offspring.

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Microbiota and Human Reproduction: The Case of Female Infertility

High-Throughput ◽

10.3390/ht9020012 ◽

2020 ◽

Vol 9 (2) ◽

pp. 12 ◽

Cited By ~ 2

Author(s):

Rossella Tomaiuolo ◽

Iolanda Veneruso ◽

Federica Cariati ◽

Valeria D’Argenio

Keyword(s):

Reproductive Tract ◽

Assisted Reproductive Technologies ◽

Embryo Implantation ◽

Reproductive Technologies ◽

Human Reproduction ◽

Microbial Dysbiosis ◽

Technical Issues ◽

Almost All ◽

Generation Sequencing

During the last decade, the availability of next-generation sequencing-based approaches has revealed the presence of microbial communities in almost all the human body, including the reproductive tract. As for other body sites, this resident microbiota has been involved in the maintenance of a healthy status. As a consequence, alterations due to internal or external factors may lead to microbial dysbiosis and to the development of pathologies. Female reproductive microbiota has also been suggested to affect infertility, and it may play a key role in the success of assisted reproductive technologies, such as embryo implantation and pregnancy care. While the vaginal microbiota is well described, the uterine microbiota is underexplored. This could be due to technical issues, as the uterus is a low biomass environment. Here, we review the state of the art regarding the role of the female reproductive system microbiota in women’s health and human reproduction, highlighting its contribution to infertility.

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Which Low-Abundance Proteins are Present in the Human Milieu of Gamete/Embryo Maternal Interaction?

International Journal of Molecular Sciences ◽

10.3390/ijms20215305 ◽

2019 ◽

Vol 20 (21) ◽

pp. 5305 ◽

Cited By ~ 3

Author(s):

Canha-Gouveia ◽

Paradela ◽

Ramos-Fernández ◽

Prieto-Sánchez ◽

Sánchez-Ferrer ◽

...

Keyword(s):

Embryo Development ◽

Reproductive Tract ◽

Culture Media ◽

Adult Life ◽

Early Embryo ◽

Female Reproductive Tract ◽

High Throughput Analysis ◽

Maternal Interaction ◽

Oviductal Fluid ◽

Abundance Proteins

The improvement of the embryo culture media is of high relevance due to its influence on successful implantation rates, pregnancy, neonatal outcomes, and potential effects in adult life. The ideal conditions for embryo development are those naturally occurring in the female reproductive tract, i.e., the oviductal and uterine fluids. To shed light on the differences between chemical and natural media, we performed the first comparative study of the low abundance proteins in plasma, uterine, and oviductal fluid collected, simultaneously, from healthy and fertile women that underwent a salpingectomy. The rationale for this design derives from the fact that high-abundant proteins in these fluids are usually those coming from blood serum and frequently mask the detection of low abundant proteins with a potentially significant role in specific processes related to the embryo–maternal interaction. The proteomic analysis by 1D-nano LC ESI-MSMS detected several proteins in higher amounts in oviductal fluid when compared to uterine and plasma samples (RL3, GSTA1, EZRI, DPYSL3, GARS, HSP90A). Such oviductal fluid proteins could be a target to improve fertilization rates and early embryo development if used in the culture media. In conclusion, this study presents a high-throughput analysis of female reproductive tract fluids and contributes to the knowledge of oviductal and uterine secretome.

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163 EVALUATION OF VIABILITY OF BULL SEMEN COLLECTED BY ELECTRO-EJACULATION USING COMMERCIAL SEMEN EXTENDER AND 2 CULTURE MEDIA AT CONTROLLED ROOM TEMPERATURE

Reproduction Fertility and Development ◽

10.1071/rdv29n1ab163 ◽

2017 ◽

Vol 29 (1) ◽

pp. 190

Author(s):

A. M. Raseona ◽

O. A. Ajao ◽

L. D. Nethengwe ◽

L. R. Madzhie ◽

T. L. Nedambale ◽

...

Keyword(s):

Sperm Motility ◽

Assisted Reproductive Technologies ◽

Culture Media ◽

Reproductive Technologies ◽

Sperm Viability ◽

Progressive Motility ◽

Bull Semen ◽

Significant Difference ◽

Semen Extender ◽

Motility Rate

Preservation of semen is an important process to ensure that semen quality is sufficient for assisted reproductive technologies. The aim of this study was to evaluate the viability of bull semen collected by electro-ejaculation using commercial semen extender and 2 modified culture media stored at controlled RT (24°C) for 72 h. Two Nguni bulls were used for semen collection; after collection, the semen was evaluated macroscopically for volume, pH, and colour, and microscopically for sperm motility, viability, and morphology. Uncontaminated semen samples with progressive motility >70% and morphological defects <20% were pooled after collection before being aliquoted into 3 extenders, namely Triladyl, modified Ham’s F10, and TCM-199 culture media, at a dilution ratio of 1:4 and then stored at controlled RT (24°C). Sperm motility rate was analysed using the computer-aided sperm analyser after 0, 24, 48, and 72 h of storage. Sperm morphology and viability was performed after staining the sperm cells with spermac and nigrosine-eosin stain, respectively. The study was replicated 4 times and data were analysed using ANOVA. Triladyl had a higher sperm viability rate (41.3%) and total motility rate (96.3%) for 72 h (P < 0.01) compared with the 2 modified culture media, Ham’s F10 (26.5 and 86.8%) and TCM-199 (25.0 and 86.7%), respectively. However, Ham’s F10 had higher progressive motility rate (37.8%) as compared with the other extenders, TCM-199 (31.7%) and Triladyl (23.4). There was no significant difference (P > 0.05), in viability rate between Ham’s F10 (26.5%) and TCM-199 (25.0%). No significant difference (P > 0.05) in straight line velocity was observed for the three extenders. Furthermore, no significant difference was observed in total sperm abnormalities, except for reacted acrosomes and absent tails (P > 0.05), between the 2 Nguni bulls. Nguni semen can be preserved in Triladyl or modified Ham’s F10 and TCM-199 culture media stored at 24°C and stay viable for 72 h. Triladyl proved to be the best suitable extender of the 3 extenders, showing higher sperm viability and total motility rate as compared with Ham’s F10 and TCM-199 modified culture media.

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Embryo Culture Conditions and the Epigenome

Seminars in Reproductive Medicine ◽

10.1055/s-0038-1675777 ◽

2018 ◽

Vol 36 (03/04) ◽

pp. 211-220 ◽

Cited By ~ 4

Author(s):

Sneha Mani ◽

Monica Mainigi

Keyword(s):

Embryo Culture ◽

Assisted Reproductive Technologies ◽

Culture Media ◽

Reproductive Technologies ◽

Common Mechanism ◽

Success Rates ◽

Blastocyst Development ◽

Increased Risk ◽

Modifiable Factors

AbstractAssisted reproductive technologies (ARTs) lead to an increased risk for pregnancy complications, congenital abnormalities, and specific imprinting disorders. Epigenetic dysfunction is thought to be one common mechanism which may be affecting these outcomes. The timing of multiple ART interventions overlaps with developmental time periods that are particularly vulnerable to epigenetic change. In vitro embryo culture is known to impact blastocyst development, in vitro fertilization (IVF) success rates, as well as neonatal outcomes. Embryo culture, in contrast to other procedures involved in ART, is obligatory, and has the highest potential for causing alterations in epigenetic reprograming. In this review, we summarize progress that has been made in exploring the effects of embryo culture, culture media, and oxygen tension on epigenetic regulation in the developing embryo. In humans, it is difficult to isolate the role of embryo culture on epigenetic perturbations. Therefore, additional well-controlled animal studies isolating individual exposures are necessary to minimize the epigenetic effects of modifiable factors utilized during ART. Findings from these studies will likely not only improve IVF success rates but also reduce the risk of adverse perinatal outcomes.

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Microphysiologic systems in female reproductive biology

Experimental Biology and Medicine ◽

10.1177/1535370217697386 ◽

2017 ◽

Vol 242 (17) ◽

pp. 1690-1700 ◽

Cited By ~ 10

Author(s):

Alexandria N Young ◽

Georgette Moyle-Heyrman ◽

J Julie Kim ◽

Joanna E Burdette

Keyword(s):

Reproductive Biology ◽

Embryo Development ◽

Reproductive Tract ◽

New Technologies ◽

Assisted Reproductive Technologies ◽

Reproductive Technologies ◽

Female Reproductive Tract ◽

The Body ◽

Technological Advancement ◽

Organ Systems

Microphysiologic systems (MPS), including new organ-on-a-chip technologies, recapitulate tissue microenvironments by employing specially designed tissue or cell culturing techniques and microfluidic flow. Such systems are designed to incorporate physiologic factors that conventional 2D or even 3D systems cannot, such as the multicellular dynamics of a tissue–tissue interface or physical forces like fluid sheer stress. The female reproductive system is a series of interconnected organs that are necessary to produce eggs, support embryo development and female health, and impact the functioning of non-reproductive tissues throughout the body. Despite its importance, the human reproductive tract has received less attention than other organ systems, such as the liver and kidney, in terms of modeling with MPS. In this review, we discuss current gaps in the field and areas for technological advancement through the application of MPS. We explore current MPS research in female reproductive biology, including fertilization, pregnancy, and female reproductive tract diseases, with a focus on their clinical applications. Impact statement This review discusses existing microphysiologic systems technology that may be applied to study of the female reproductive tract, and those currently in development to specifically investigate gametes, fertilization, embryo development, pregnancy, and diseases of the female reproductive tract. We focus on the clinical applicability of these new technologies in fields such as assisted reproductive technologies, drug testing, disease diagnostics, and personalized medicine.

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191 TIMING OF OVULATION IN THE GONADOTROPHIN-STIMULATED SOUTHERN HAIRY-NOSED WOMBAT, LASIORHINUS LATIFRONS

Reproduction Fertility and Development ◽

10.1071/rdv17n2ab191 ◽

2005 ◽

Vol 17 (2) ◽

pp. 246 ◽

Cited By ~ 1

Author(s):

G.V. Druery ◽

M.D. Rival ◽

D.A. Taggart ◽

G.A. Shimmin ◽

A.B. Horsup ◽

...

Keyword(s):

Reproductive Tract ◽

Assisted Reproductive Technologies ◽

Reproductive Technologies ◽

Ovarian Activity ◽

Corpora Lutea ◽

Transabdominal Ultrasonography ◽

Group 2 ◽

Follicular Response ◽

Circulating Levels ◽

Breeding Techniques

The southern hairy-nosed wombat (SHW), Lasiorhinus latifrons, is a model species in which to develop assisted breeding techniques for the endangered northern hairy-nosed wombat, Lasiorhinus krefftii. We recently showed that anoestrus SHW respond to eutherian gonadotrophins by production of multiple ovarian follicles, but ovulation had not occurred at the time of examination 24 h post-LH (Druery GV et al. 2003 Theriogenology 59, 391 abst). This study investigated the timing of ovulation in six anoestrus captive adult female SHW (n = 3 per group) after ovarian superstimulation using porcine FSH (200 mg total, Folltropin-V, Bioniche, Belleville, Ontario, Canada) administered s.c. at 12-h intervals over 7 days. Ovulation was triggered by a single s.c. dose of porcine LH (25 mg Lutropin-V, Bioniche) 12 h after the final FSH injection. Superstimulatory response was determined by laparoscopy immediately after the final FSH injection on Day 7 prior to LH. Group 1 was re-examined at 33, 36, and 39 h post-LH, and Group 2 at 42, 45, and 48 h post-LH, for evidence of ovulations using laparoscopy and transabdominal ultrasonography. Laparoscopy on Day 7 revealed an ovarian follicular response in all six females, which coincided with the highest levels of estradiol. The reproductive tract also responded to the treatment (swollen fimbriae and enlarged, highly vascular uteri). Multiple follicles (range 16–31) up to 11 mm in diameter were observed in five females. One female had ovulated, as determined by the presence of corpora lutea. Transabdominal ultrasonographic imaging was unable to confirm the number of follicles in stimulated ovaries. Ovulation had commenced by 36 h post-LH, with the majority occurring 39–45 h post-LH. Ovulation was recorded as having occurred if a dark red, highly vascular crater on the surface of the newly formed corpus hemorrhagicum was observed. Increased circulating levels of progesterone were confirmed 9 days after the last laparoscopies. These results have important implications for the development of assisted reproductive technologies in the SHW: (1) transabdominal ultrasound imaging is ineffective for determining ovarian activity; (2) laparoscopy is a well-tolerated, repeatable minor surgical procedure that can be used for intrauterine AI in this species in which nonsurgical AI is unlikely to succeed (Paris DBBP et al. 2003 Theriogenology 59, 401 abst); and (3) knowledge of the timing of ovulation will enable insemination of spermatozoa into the uterus prior to ovulation. Financial support was provided by Dr. M. Jacobson, and hormones were supplied by Bioniche.

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