https://escholarship.umassmed.edu/cgi/viewcontent.cgi?article=1006&context=covid19

SARS-CoV-2

Oocyte quality affects early embryonic survival, the establishment and maintenance of pregnancy, fetal development, and even adult disease. Quality, or developmental competence, is acquired during folliculogenesis as the oocyte grows, and during the period of oocyte maturation. Assisted reproductive technologies involving ovarian hyperstimulation, or collection of immature oocytes followed by maturation in vitro, perturb this process and result in oocytes with reduced quality. In domestic livestock species, offspring have been produced using in vitro oocyte maturation, although only a small percentage of the original pool of immature oocytes is capable of developing to the blastocyst stage and subsequently resulting in pregnancy. In vitro maturation, as it is currently undertaken, does not support the correct development of oocyte competence. Follicle size affects oocyte quality, potentially implicating messenger RNA or protein stores as factors involved in oocyte competence. Oocytes from preantral follicles grown in vitro are competent to resume meiosis, although development to the blastocyst stage is decreased. An offspring from oocytes produced using this technique was normal at birth but experienced delayed onset health issues, highlighting the importance of oocyte quality long after embryogenesis. Metabolism may play a critical role in oocyte quality because glycolytic activity in mature oocytes is correlated with increased embryonic development. Communication between the oocyte and its surrounding cumulus cells is also important for the development of a competent oocyte. Ovarian stimulation causes delayed embryonic development, increased abnormal blastocyst formation, fetal growth retardation, and increased fetal loss. Thus, although meiosis and even early development may be completed successfully, there are a variety of other processes occurring within the cytoplasm of the oocyte that are required for complete developmental competence. However, the cellular mechanisms that impart oocyte quality are unclear. Until the mechanisms involved in oocyte quality are elucidated, any effort to use assisted reproductive technologies in animals for production or biomedical purposes will be inefficient at best.

The effect of oocyte quality on development.

Effect of the addition of insulin-transferrin-selenium and/or L-ascorbic acid to the in vitro maturation of prepubertal bovine oocytes on cytoplasmic maturation and embryo development.

Published reports have indicated that rates of preimplantation embryo aneuploidy in a control donor population may vary between IVF centres. This suggests that location-specific conditions, in the clinic, IVF or genetics laboratory, may be influencing the chromosome dynamics or diagnosis. More recent reports suggest that rates of embryo mosaicism, representing mitotic errors, may vary between IVF centres. This would suggest perhaps a laboratory-controlled variable is influencing mitotic cell division during preimplantation embryo development. Various IVF laboratory-controlled factors may be impacting chromosome separation and segregation. Variables including type of culture media, pH, temperature, osmolality and oxygen concentration could all be possible factors influencing embryo aneuploidy. Furthermore, laboratory techniques, method of insemination, laser use or handling of biopsied cells may also influence genetic results. These IVF laboratory variables will be reviewed and literature suggesting a possible link to mitotic aneuploidy/mosaicism discussed.

Controversies in ART: can the IVF laboratory influence preimplantation embryo aneuploidy?

As the coronavirus pandemic is far from ending, more questions regarding the female reproductive system, particularly fertility issues, arise. The purpose of this paper is to bring light upon the possible link between COVID-19 and women’s reproductive health. This review emphasizes the effect of SARS-CoV-2 on the hormones, endometrium and menstrual cycle, ovarian reserve, follicular fluid, oocytes, and embryos. The results showed that endometrial samples did not express SARS-CoV-2 RNA. Regarding the menstrual cycle, there is a large range of alterations, but they were all reversible within the following months. The ovarian reserve was not significantly affected in patients recovering from both mild and severe infection in most cases, except one, where the levels of AMH were significantly lower and basal follicle-stimulating hormone (FSH) levels were increased. All COVID-19 recovered patients had positive levels of SARS-CoV-2 IgG in the follicular fluid. The amount of retrieved and mature oocytes and the fertilization rate were unharmed in three studies, except for one study, where the quantity of retrieved and mature oocytes was reduced in patients with higher levels of SARS-CoV-2 antibodies. The numbers of blastocysts, top-quality embryos, and euploid embryos were affected in most of the studies reviewed.

/pdf/the-effects-of-sars-cov-2-infection-on-female-fertility-a-2ny6jjlp.pdf

The Effects of SARS-CoV-2 Infection on Female Fertility: A Review of the Literature

Objective: To study mRNA and protein expression of SARS-CoV-2 entry receptors (ACE2, CD147) and proteases (TMPRSS2, CTSL) in human oocytes, embryos, cumulus and granulosa cells. Design: Research Study. Setting: Clinical IVF treatment center. Patients: IVF patients treated at the Colorado Center for Reproductive Medicine. Interventions: Oocytes (GV, MII) and embryos (1 cell (1C); blastocyst (BL)) were donated to research at disposition by IVF patients. Follicular cells (cumulus (CC); granulosa (GC)) were collected from women undergoing egg retrieval after ovarian stimulation without an ovulatory trigger for IVM/IVF treatment cycles. Main Outcome Measures: The presence or absence of ACE2, CD147, TMPRSS2, and CTSL mRNA using RT-qPCR and protein using capillary Western blotting in human oocytes, embryos and ovarian follicular cells. Result s: RT- q-PCR analysis revealed high abundance of ACE2 gene transcripts in GV and MII oocytes compared to CC, GC, and BL. ACE2 protein was only present in MII oocytes, 1C, and BL embryos, but other ACE2 protein variants were observed in all the samples. TMPRSS2 protein was present in all samples while mRNA was observed only at the blastocyst stage. All the samples were positive for CD147 and CTSL mRNA expression. However, cumulus and granulosa cells were the only samples that showed co-expression of both CD147 and CTSL proteins in low abundance. Conclusions: Cumulus and granulosa cells are least susceptible to SARS-CoV-2 infection due to the lack of required receptors and proteases combination (ACE2/TMPRSS2 or CD147/CTSL) in high abundance. Co-expression of the ACE2 and TMPRSS2 proteins in MII oocytes, zygotes, and blastocysts demonstrate that these gametes and embryos have the cellular machinery required and thus are potentially susceptible to SARS-CoV-2 infection if exposed to the virus. However, we do not yet know if infection occurs in vivo, or in vitro in an ART setting.

Human eggs, zygotes, and embryos express the receptor angiotensin 1-converting enzyme 2 and transmembrane serine protease 2 protein necessary for severe acute respiratory syndrome coronavirus 2 infection.

https://www.fertstert.org/article/S0015-0282(18)31607-8/pdf

Media evaporation in a dry culture incubator; effect of dish, drop size and oil on media osmolality

Objective To characterize fatty acid (FA) profile of commercially available albumin products and determine their effect on embryonic development. Design Research study. Setting Private research facility. Animal(s) Outbred mice aged 4–8 weeks. Intervention(s) Gas chromatography-mass spectrometry was used to quantify the FA content of 15 commercial albumins. Embryos were produced in media containing different albumin products, with or without carnitine or exogenous FA supplementation, to determine their effect on embryo development in vitro. Main Outcome Measure(s) Total micrograms of FA per milligram of albumin for the 15 albumin products, blastocyst development, cell number, allocation to the trophectoderm (TE) or inner cell mass (ICM), and evaluation of morphology during implantation. Result(s) The albumin products contained 0.07–16.77 μg total FA/mg albumin. Compared to media with with >1.4 μg FA/mg albumin, media with Conclusion(s) Commercial albumin products contain FAs, which vary in abundance. These FAs have different effects on embryo development and quality before and during the implantation stage. Several of these albumin preparations are routinely used for human-assisted reproductive technologies; therefore, serious consideration is warranted when selecting a product for clinical use.

Fatty acids present in commercial albumin preparations differentially affect development of murine embryos before and during implantation

Oocyte metabolism, particularly the pentose phosphate pathway (PPP), is a critical component of porcine oocyte maturation. The objective of this study was to determine the effects of manipulating the PPP on nuclear (completion of meiosis) and cytoplasmic (glycolytic and PPP activity, intracellular glutathione (GSH) content, and developmental competence) maturation of porcine oocytes. Cumulus-oocyte complexes were recovered from abattoir ovaries and matured for 42-44 h in the presence of 0.01 IU/ml FSH and LH, 10 ng/ml EGF, and 0.6 mM cysteine with or without 0.1 μM of diphenyleneiodonium (DPI; PPP inhibitor) or pyrroline-5-carboxylic acid (PC; PPP stimulator). Following maturation, a portion of the oocytes were incubated with 5-[H]glucose and 1-[C]-glucose to determine the activity of glycolysis and the PPP, respectively. At the conclusion of metabolic measurements, oocytes were stained to evaluate meiotic progression. Remaining oocytes were either frozen for subsequent GSH analysis (DTNB-GSSG reductase recycling assay) or inseminated (5 x 10 sperm/ml; 5 h) and cultured to determine developmental competence.

/pdf/manipulation-of-the-pentose-phosphate-pathway-in-porcine-znyhiek0a9.pdf

Manipulation of the Pentose Phosphate Pathway in Porcine Oocytes: Effects on Nuclear and Cytoplasmic Maturation

Chromatin reorganization governs gene expression regulation during pre-implantation development. However, the global chromatin landscape and its dynamics in this period remain unexplored in bovine. In this study, we constructed a genome-wide map of accessible chromatin in bovine oocytes and early embryos using an improved assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq). We analysed pools of 20 germinal vesicles or MII oocytes or 2-, 4-, 8-, 16-cell, morula, and blastocyst stage in vitro-produced embryos. We conducted ATAC-seq on six pools for each stage and an additional four pools of in vivo-derived morula and blastocysts and six replicates using individual Day 14 elongating embryos. We obtained ~110 million paired end reads uniquely mapped to the bovine reference genome for each stage. Hierarchical clustering, t-distributed stochastic neighbour embedding, and principal component analysis showed four distinct patterns for open chromatin status: (1) low accessibility in germinal vesicles and MII oocytes and in 2- and 4-cell embryos; (2) significantly elevated accessibility in 8-cell, 16-cell, and morula embryos; (3) less accessibility in blastocysts; and (4) extremely high accessibility in elongating embryos. This dynamic and sequential chromatin remodelling is consistent with transcription activation during the bovine minor embryonic genome activation from fertilization to 4-cell, major embryonic genome activation at 8-cell, first differentiation at blastocyst and drastic transcription initiation for embryo elongation. Genome-wide characteristics of accessible chromatin showed (1) accessible chromatin near the transcription start sites of active genes and CpG-rich promoters; (2) widespread accessible chromatin regions extensively overlapped with transposable elements; (3) distal peaks preferentially enriched for repeats including LINE, SINE, and LTR from 8-cell to morula embryos, especially for LTR, whereas enrichment in simple repeats were found from oocytes to 4-cell and in elongating embryos; and (4) highly stage-specific transcription factor motifs in distal peaks were unveiled. By integrating the maps of chromatin accessibility with bovine embryo transcriptomes and DNA methylomes, we found promoter accessibility and DNA methylation in bovine embryos correlated with both gene activities and CpG densities. Most importantly, we constructed the regulatory networks of stage-specific expressed genes and stage-specific activated genes with three omics datasets in bovine early embryos and revealed conserved and distinctive transcriptional regulatory networks between in vivo- and in vitro-derived embryos. This comprehensive analysis revealed critical features of the chromatin landscape and epigenetic reprogramming during bovine early embryo development.

R.L. Krisher

Papers

Human eggs, zygotes, and embryos express the receptor angiotensin 1-converting enzyme 2 and transmembrane serine protease 2 protein necessary for severe acute respiratory syndrome coronavirus 2 infection.

Media evaporation in a dry culture incubator; effect of dish, drop size and oil on media osmolality

Fatty acids present in commercial albumin preparations differentially affect development of murine embryos before and during implantation

Manipulation of the Pentose Phosphate Pathway in Porcine Oocytes: Effects on Nuclear and Cytoplasmic Maturation

2 The landscape of accessible chromatin in bovine oocytes and early embryos