BACKGROUND Cryopreservation has been extensively used in assisted reproductive technology, agriculture and conservation programmes for endangered species. The literature reports largely positive results regarding the survival of frozen-thawed cells and clinical outcomes. Nonetheless, it is unclear whether or not cryopreservation of sperm, oocytes and embryos causes any disruption in their genetic integrity. Drawing on the available published evidence, this review paper describes in detail the physical and biochemical factors of cryopreservation that could potentially affect genomic integrity. METHODS A critical review of the published literature using PubMed with particular emphasis on studies which include assessment of genetic stability after cryopreservation of oocyte, sperm and embryos. The search was performed in 2014 and covered the period from the beginning of electronic records until July 2014. No language restrictions were applied. RESULTS Cryopreservation is associated with extensive damage to cell membranes, and results in alteration of the functional and metabolic status of the cells and mitochondria. Some evidence suggests an increase in DNA single-strand breaks, and degree of DNA condensation or fragmentation in sperm after cryopreservation. The extent of these changes may vary between different individuals and different techniques. The addition of antioxidants to the cryopreservation media and the use of well-controlled cooling regimes could potentially improve such outcomes. Limited numbers of studies on oocytes provide controversial results regarding the effect on DNA fragmentation, sister chromatid exchange (SCE) and aneuploidy. The only study on human embryos suggested that vitrification affects DNA integrity to a much lesser extent than slow freezing. Animal studies show increases in mitochondrial DNA mutations in embryos after cryopreservation. The limited numbers of long-term follow-up studies in humans provide reassurance that derives mostly from retrospective studies with some methodological weaknesses. CONCLUSIONS This review provides an overview of studies performed to date on the effect of cryopreservation on the oocyte, sperm and embryos. Controversy of the reported data has highlighted the gaps in our knowledge not only for clinical studies, but also for basic research in human embryos. New perspectives for future research are proposed.

/pdf/the-effect-of-cryopreservation-on-the-genome-of-gametes-and-pisg9trds7.pdf

The effect of cryopreservation on the genome of gametes and embryos: principles of cryobiology and critical appraisal of the evidence

Study question Do the extreme conditions of vitrification affect mitochondrial health and reactive oxygen species (ROS) levels of human oocytes? Summary answer Vitrification of discarded human oocytes shifts the intracellular redox potential towards oxidation but does not alter the mitochondrial potential or intracellular ROS levels. What is known already Recent studies have reflected increased ROS levels in warmed young oocytes and have highlighted the temporal dynamic loss of mitochondrial potential that could, therefore, lead to a decrease in ATP production, impairing embryo development. Mitochondrial function can also be evaluated in vivo by the FAD/NAD(P)H autofluorescence ratio, which reflects the respiratory chain activity and is considered as a marker of the intracellular redox state. Study design, size, duration A total of 629 discarded Metaphase II (MII) oocytes collected from June 2013 to April 2014 were included in this control (fresh oocytes, n= 270) versus treatment (vitrified oocytes, n= 359) study. Participants/materials, setting, methods Discarded MII oocytes were donated to research by young ( 36 years old) women who underwent ovarian stimulation for IVF at a university-affiliated private fertility clinic. Redox state was assessed by measuring the FAD/NAD(P)H autofluorescence ratio, while ROS and mitochondrial activity were reported by in vivo labelling with carboxy-H2DCFDA and JC-1, respectively. Main results and the role of chance Young and aged oocytes showed high and similar survival rates (81.8 versus 83.1%, not significant). Confocal microscopy revealed that the FAD/NAD(P)H ratio was significantly higher in vitrified oocytes than in fresh oocytes, suggesting a significant shift towards the oxidized state in oocytes after vitrification, regardless of the maternal age. Mitochondrial distribution was not affected by vitrification. Furthermore, it was not possible to resolve any difference in mitochondrial potential using JC-1 potentiometric dye or in reactive oxygen species (ROS) production (assessed with H2-DCFDA staining) between fresh and vitrified oocytes. Therefore, measurement of intracellular redox potential by autofluorescence imaging may be a more sensitive method to assess oxidative stress or mitochondrial demise in human oocytes because it showed a higher resolving power than JC-1 staining and displayed less variability than H2-DCFDA staining. Limitations, reasons for caution Owing to sample availability, MII discarded oocytes (in vitro matured oocytes and unfertilized oocytes 20 h after ICSI) were included in the study. These discarded oocytes do not necessarily reflect the physiological condition of the MII human oocyte. Wider implications of the findings Although vitrified oocytes yield comparable clinical outcomes compared with fresh oocytes, lower cleavage and blastocyst rates can be observed during in vitro culture. Data here obtained suggest that the redox state of human oocytes could be affected by vitrification. Therefore, the importance of adding protective antioxidant molecules to the vitrification solution and to the post-warming culture medium to improve embryo cleavage deserves some research. Study funding/competing interests This research project was supported by the Valencian Government (Val+i+D program, M.N.-C.), INCLIVA Foundation for health research (G.S.-A.) and by the University of L'Aquila and Regione Abruzzo ('Reti per l'Alta Formazione' - P.O.F.S.E. Abruzzo 2007-2013 G.D.E.). No conflicts of interest were declared.

/pdf/impact-of-vitrification-on-the-mitochondrial-activity-and-1w84frbke5.pdf

Impact of vitrification on the mitochondrial activity and redox homeostasis of human oocyte

Background
Recent advances in vitrification technology have markedly improved the efficacy of oocyte cryopreservation in terms of oocyte survival and pregnancy, as well as live birth rates. However, there still remains room for improvement in terms of vitrification techniques.

Oocyte vitrification: advances, progress and future goals.

Cryopreservation of oocytes is becoming a valuable method for fertility preservation in women. However, various unphysiological alterations occur in the oocyte during the course of cryopreservation, one of which is the disappearance of the meiotic spindle. Fortunately, the meiotic spindle does regenerate after thawing the frozen oocytes, which enables completion of meiosis and further development after fertilization. Nonetheless, the mechanistic understanding of the meiotic spindle regeneration after cryopreservation is still scarce. Here, to gain insight into the mechanisms of the spindle disappearance and regeneration, we examined the status of spindle microtubules as well as the key components of the microtubule-organizing center (MTOC), specifically gamma-Tubulin, NEDD1, and Pericentrin, in mature (metaphase II) mouse oocytes at different steps of vitrification, a major cryopreservation technique. We found that the configuration of the spindle microtubules dynamically changed during the process of vitrification and that spindle regeneration was preceded by excessive microtubule polymerization, followed by reduction into the normal size and shape. Also, all three MTOC components exhibited disappearance and reappearance during the vitrification process, although Pericentrin appeared to regenerate in earlier steps compared to the other components. Furthermore, we found that the localization of the MTOC components to the spindle poles persisted even after depolymerization of spindle microtubules, suggesting that the MTOC components are impacted by vitrification independently from the integrity of the microtubules. The present study would set the stage for future investigations on the molecular mechanisms of the meiotic spindle regeneration, which may contribute to further improving protocols for oocyte cryopreservation.

https://bioone.org/journals/biology-of-reproduction/volume-89/issue-5/biolreprod.113.108167/Impact-of-Vitrification-on-the-Meiotic-Spindle-and-Components-of/10.1095/biolreprod.113.108167.pdf

Impact of Vitrification on the Meiotic Spindle and Components of the Microtubule-Organizing Center in Mouse Mature Oocytes

https://www.fertstert.org/article/S0015-0282(14)00583-4/pdf

Effect of oocyte vitrification on deoxyribonucleic acid methylation of H19, Peg3, and Snrpn differentially methylated regions in mouse blastocysts

Background Cryopreservation is now considered as an efficient way to store human oocytes to preserve fertility. However, little is known about the effects of this technology on oocyte gene expression. The aim of this study was to examine the effect of the two cryopreservation procedures, slow freezing and vitrification, on the gene expression profile of human metaphase II (MII) oocytes. Methods Unfertilized MII oocytes following ICSI failure were cryopreserved either by slow freezing or by the Cryotip method for vitrification. After thawing, total RNA was extracted and analyzed using Affymetrix Human Genome U133 Plus 2.0 GeneChip arrays. The gene expression profiles and associated biological pathways in slowly frozen/thawed and vitrified MII oocytes were determined and compared with those of non-cryopreserved MII oocytes used as controls. Results Both cryopreservation procedures negatively affected the gene expression profile of human MII oocytes in comparison with controls. However, slowly frozen and vitrified MI oocytes displayed specific gene expression signatures. Slow freezing was associated with down-regulation of genes involved in chromosomal structure maintenance (KIF2C and KIF3A) and cell cycle regulation (CHEK2 and CDKN1B) that may lead to a reduction in the oocyte developmental competence. In vitrified oocytes, many genes of the ubiquitination pathway were down-regulated, including members of the ubiquitin-specific peptidase family and subunits of the 26S proteasome. Such inhibition of the degradation machinery might stabilize the maternal protein content that is necessary for oocyte developmental competence. Conclusions The low pregnancy rates commonly observed when using human MII oocytes after slow freezing-thawing may be explained by the alterations of the oocyte gene expression profile.

https://www.hal.inserm.fr/inserm-00726705/document

Slow freezing and vitrification differentially modify the gene expression profile of human metaphase II oocytes

Handle everyday research tasks with reliable, citation-backed results

Your personal Research Agent to handle research tasks with citation-backed results

Popular Tasks used by Researchers

How can I help with your research?

Meet SciSpace

Get more enhanced response by uploading the PDFs you want me to reference.

No relevant PDFs in your library

SciSpace is the AI research assistant for academics. Run systematic literature reviews on 280M+ papers, and write papers with cited sources. Trusted by 1M+ students, PhDs & researchers.

SciSpace | AI for Research

Analyze PDFs

Code & Manuscripts

Funding & Grants

Literature & Patents

Medical & Clinical Data

Systematic Review

Visualize & Present

Web & Data

Build a Google Scholar-like website for your research.

Build a website

Create charts and images for your research

Create a Chart

Write a paper for submission to a journal

Draft a manuscript

Patent Search

Design eye-catching scientific posters in minutes.

Scientific Poster Generation

Systematic Literature Review

One task is running at the moment. Your messages will be shown right after.

Drag and drop or click here to browse

Loved by <highlight>1 million+</highlight> researchers

Extract a list of specific topics and their sources from unstructured text

Topics

Compare and analyze relevant papers that matches with your search

Papers

Get insights from PDFs and bookmarked papers from your library

My library

Recent searches

Try searching for:

Catch AI-generated content in scholarly and non-scholarly content

{ai} Detector

Ai Writer

Get PDF Summaries, highlighted text explanations 

Chat with PDF

Effortlessly create in-text citations and bibliographies in APA and 2,500 other formats

Citation generator

Get explanations, summaries, and answers on academic papers

Ease up your research workflow with {scispace}'s cohort of exciting AI tools

Elevate your academic writing skills and convey your ideas the way you want

Paraphraser

Explore our range of reading and writing tools

Your file is being prepared and should be ready in a few minutes. If it's a large file, it might take a bit longer. You can close this window, and we'll email you the file when it's done.

You have reached a maximum limit of <strong>{limit}</strong> columns in the table. Remove at least <strong>1</strong> column to add or create another one.

K. Roman

Author Tools

Chat about Author

Papers

Slow freezing and vitrification differentially modify the gene expression profile of human metaphase II oocytes