Skip to main content
Key Specialties
Cardiology Diabetology Endocrinology Gastroenterology Geriatrics and Gerontology Gynecology and Obstetrics Hematology Infectious Disease Internal Medicine Nephrology Neurology Oncology Pediatrics Respiratory Medicine Rheumatology Surgery
Congress Coverage
2024 ACC Congress 2023 ESMO Congress 2023 EASD Congress 2023 ERS Congress 2023 ESC Congress
Clinical Collections
Advances in Alzheimer’s

Keynote webinar | Spotlight on medication adherence

LIVE: Thursday 27th June 2024, 18:00-19:30 (CEST)
Join our expert panel to discover why you need to understand the drivers of non-adherence in your patients, and how you can optimize medication adherence in your clinics to drastically improve patient outcomes.
ADVANCED SEARCH
Log in
Top
Reproductive Biology and Endocrinology
Published in: Reproductive Biology and Endocrinology 1/2013

Open Access 01-12-2013 | Research

Vitrification preserves chromatin integrity, bioenergy potential and oxidative parameters in mouse embryos

Authors: Nicola A Martino, Maria E Dell’Aquila, Rosa A Cardone, Bence Somoskoi, Giovanni M Lacalandra, Sandor Cseh

Published in: Reproductive Biology and Endocrinology | Issue 1/2013

Login to get access

Abstract

Background

The aim of this study was to evaluate the effects of vitrification on morpho-functional parameters (blastomere/chromatin integrity and bioenergy/oxidative potential) of mouse preimplantation embryos.

Methods

In vivo produced mouse (4/16-cell, morulae and blastocyst-stage) embryos were randomly divided into vitrification and control groups. For vitrification, embryos were exposed to a 2-step loading of ethylene glycol and propylene glycol, before being placed in a small nylon loop and submerged into liquid nitrogen. After warming, the cryoprotectants were diluted by a 3-step procedure. Embryo morphology, chromatin integrity and energy/oxidative status were compared between groups.

Results

Vitrification induced low grade blastomere cytofragmentation (P < 0.05) and low chromatin damage only in embryos at the morula stage (P < 0.001). Mitochondrial (mt) distribution pattern was affected by vitrification only in early embryos (P < 0.001). Mitochondrial activity did not change upon vitrification in morula-stage embryos but it was reduced in blastocyst-stage embryos (P < 0.05). Intracellular ROS levels significantly increased in embryos at the morula and blastocyst stages (P < 0.001). Colocalization of active mitochondria and ROS increased only in vitrified blastocysts.

Conclusions

In conclusion, this study elucidates the developmentally-related and mild effects of vitrification on morphology, nuclear and bioenergy/oxidative parameters of mouse embryos and demonstrates that vitrification is a suitable method for preserving predictive parameters of embryo ability to induce a full-term pregnancy.
Appendix
Available only for authorised users
Literature
1.
Fuller B, Paynter S, Watson P: Cryopreservation of human gametes and embryos. Life in the frozen state. Edited by: Fuller B, Lane N, Benson E. 2004, Boca Raton: CRC Press, 505-541.CrossRef
2.
Rall WF, Fahy GM: Ice-free cryopreservation of mouse embryos at -196 degree C by vitrification. Nature. 1985, 313: 573-575. 10.1038/313573a0.CrossRefPubMed
3.
Vajta G: Vitrification of oocytes and embryos of domestic animals. Anim Reprod Sci. 2000, 60–61: 357-364.CrossRefPubMed
4.
Rall WF: Factors affecting the survival of mouse embryos cryopreserved by vitrification. Cryobiology. 1987, 24: 387-402. 10.1016/0011-2240(87)90042-3.CrossRefPubMed
5.
Liebermann J, Nawroth F, Isachenko V, Isachenko E, Rahimi G, Tucker MJ: Potential importance of vitrification in reproductive medicine. Biol Reprod. 2002, 67: 1671-1680. 10.1095/biolreprod.102.006833.CrossRefPubMed
6.
Van Blerkom J, Davis P, Alexander S: Differential mitochondrial distribution in human pronuclear embryos leads to disproportionate inheritance between blastomeres: Relationship to microtubular organization, ATP content and competence. Hum Reprod. 2000, 15: 2621-2633. 10.1093/humrep/15.12.2621.CrossRefPubMed
7.
Van Blerkom J: Mitochondrial function in the human oocyte and embryo and their role in developmental competence. Mitochondrion. 2011, 11: 797-813. 10.1016/j.mito.2010.09.012.CrossRefPubMed
8.
Zhao XM, Fu XW, Hou YP, Yan CL, Suo L, Wang YP, Zhu HB, Dinnyés A, Zhu SE: Effect of vitrification on mitochondrial distribution and membrane potential in mouse two pronuclear (2-PN) embryos. Mol Reprod Dev. 2009, 76: 1056-1063. 10.1002/mrd.21064.CrossRefPubMed
9.
Shi LY, Jin HF, Kim JG, Kumar M, Balasubramanian S, Choe SY, Rho GJ: Ultra-structural changes and developmental potential of porcine oocytes following vitrification. Anim Reprod Sci. 2007, 100: 128-140. 10.1016/j.anireprosci.2006.06.020.CrossRefPubMed
10.
Bilodeau JF, Chatterjee S, Sirard MA, Gagnon C: Levels of antioxidant defenses are decreased in bovine spermatozoa after a cycle of freezing and thawing. Mol Reprod Dev. 2000, 55: 282-288. 10.1002/(SICI)1098-2795(200003)55:3<282::AID-MRD6>3.0.CO;2-7.CrossRefPubMed
11.
Ahn HJ, Sohn IP, Kwon HC, Jo DH, Park YD, Min CK: Characteristics of the cell membrane fluidity, actin fibers, and mitochondrial dysfunctions of frozen-thawed two-cell mouse embryos. Mol Reprod Dev. 2002, 61: 466-476. 10.1002/mrd.10040.CrossRefPubMed
12.
Somfai T, Ozawa M, Noguchi J, Kaneko H, Kuriani Karja NW, Farhudin M, Dinnyés A, Nagai T, Kikuchi K: Developmental competence of in vitro-fertilized porcine oocytes after in vitro maturation and solid surface vitrification: effect of cryopreservation on oocyte antioxidative system and cell cycle stage. Cryobiology. 2007, 55: 115-126. 10.1016/j.cryobiol.2007.06.008.CrossRefPubMed
13.
Tatone C, Di Emidio G, Vento M, Ciriminna R, Artini PG: Cryopreservation and oxidative stress in reproductive cells. Gynecol Endocrinol. 2010, 26: 563-567. 10.3109/09513591003686395.CrossRefPubMed
14.
Finkel T, Holbrook NJ: Oxidants, oxidative stress and the biology of ageing. Nature. 2000, 408: 239-247. 10.1038/35041687.CrossRefPubMed
15.
Agarwal A, Allamaneni SS: Role of free radicals in female reproductive diseases and assisted reproduction. Reprod Biomed Online. 2004, 9: 338-347. 10.1016/S1472-6483(10)62151-7.CrossRefPubMed
16.
Cadenas E, Davies KJ: Mitochondrial free radical generation, oxidative stress, and aging. Free Radic Biol Med. 2000, 29: 222-230. 10.1016/S0891-5849(00)00317-8.CrossRefPubMed
17.
Winyard PG, Moody CJ, Jacob C: Oxidative activation of antioxidant defence. Trends Biochem Sci. 2005, 30: 453-461. 10.1016/j.tibs.2005.06.001.CrossRefPubMed
18.
Brookes PS, Yoon Y, Robotham JL, Anders MW, Sheu SS: Calcium, ATP, and ROS: a mitochondrial love-hate triangle. Am J Physiol Cell Physiol. 2004, 287: 817-833. 10.1152/ajpcell.00139.2004.CrossRef
19.
Klambauer P, Keresztes Z, Kanyo K, Varga E, Kriston R, Vass N, Jávor A, Konc J, Solti L, Cseh S: Vitrification of cleavage stage mouse embryos by the cryoloop procedure. Acta Vet Hung. 2009, 57 (3): 399-410. 10.1556/AVet.57.2009.3.6.CrossRefPubMed
20.
Ambruosi B, Filioli Uranio M, Sardanelli AM, Pocar P, Martino NA, Paternoster MS, Amati F, Dell’Aquila ME: In vitro acute exposure to DEHP affects oocyte meiotic maturation, energy and oxidative stress parameters in a large animal model. PLoS One. 2011, 6 (11): e27452-10.1371/journal.pone.0027452.PubMedCentralCrossRefPubMed
21.
Martino NA, Lacalandra GM, Filioli Uranio M, Ambruosi B, Caira M, Silvestre F, Pizzi F, Desantis S, Accogli G, Dell'Aquila ME: Oocyte mitochondrial bioenergy potential and oxidative stress: within-/between-subject, in vivo vs IVM and age-related variations in a sheep model. Fertil Steril. 2012, 97 (3): 720-728. 10.1016/j.fertnstert.2011.12.014.CrossRefPubMed
22.
Poot M, Zhang YZ, Kramer JA, Wells KS, Jones LJ, Hanzel DK, Lugade AG, Singer VL, Haugland RP: Analysis of mitochondrial morphology and function with novel fixable fluorescent stains. J Histochem Cytochem. 1996, 44: 1363-1372. 10.1177/44.12.8985128.CrossRefPubMed
23.
Torner H, Brussow KP, Alm H, Ratky J, Pohland R, Tuchscherer A, Kanitz W: Mitochondrial aggregation patterns and activity in porcine oocytes and apoptosis in surrounding cumulus cells depends on the stage of pre-ovulatory maturation. Theriogenology. 2004, 61: 1675-1689. 10.1016/j.theriogenology.2003.09.013.CrossRefPubMed
24.
Yang HW, Hwang KJ, Kwon HC, Kim HS, Choi K, Oh KS: Detection of reactive oxygen species (ROS) and apoptosis in human fragmented embryos. Hum Reprod. 1998, 13: 998-1002. 10.1093/humrep/13.4.998.CrossRefPubMed
25.
Valentini L, Iorga AI, De Santis T, Ambruosi B, Reynaud K, Chastant-Maillard S, Guaricci AC, Caira M, Dell'Aquila ME: Mitochondrial distribution patterns in canine oocytes as related to the reproductive cycle stage. Anim Reprod Sci. 2010, 117: 166-177. 10.1016/j.anireprosci.2009.03.008.CrossRefPubMed
26.
Van Soom A: Assessment of mammalian embryo quality: what can we learn from embryo morphology. Reprod BioMed Online. 2003, 7: 104-110.CrossRef
27.
Han Z, Chung YG, Gao S, Latham KE: Maternal factors controlling blastomere fragmentation /in early mouse embryos. Biol Reprod. 2005, 72: 612-618. 10.1095/biolreprod.104.035444.CrossRefPubMed
28.
Liu J, Tang S, Xu W, Wang Y, Yin B, Zhang Y: Detrimental effects of antibiotics on mouse embryos in chromatin integrity, apoptosis and expression of zygotically activated genes. Zygote. 2010, 19: 137-145.CrossRefPubMed
29.
Van Blerkom J, Davis P, Mathwig V, Alexander S: Domains of high-polarized and low-polarized mitochondria may occur in mouse and human oocytes and early embryos. Hum Reprod. 2002, 17 (2): 393-406. 10.1093/humrep/17.2.393.CrossRefPubMed
30.
Van Blerkom J: High-polarized mitochondria (ΔΨm HIGH) are spatially polarized in human oocytes and early embryos in stable subplasmalemmal domains: developmental significance and the concept of vanguard mitochondria. Reprod BioMed Online. 2006, 13 (2): 246-254. 10.1016/S1472-6483(10)60622-0.CrossRefPubMed
31.
Ambruosi B, Lacalandra GM, Iorga AI, De Santis T, Mugnier S, Matarrese R, Goudet G, Dell'aquila ME: Cytoplasmic lipid droplets and mitochondrial distribution in equine oocytes: implications on oocyte maturation, fertilization and developmental competence after ICSI. Theriogenology. 2009, 71: 1093-1104. 10.1016/j.theriogenology.2008.12.002.CrossRefPubMed
32.
Zhang C, Liu C, Li D, Yao N, Yuan X, Yu A, Lu C, Ma X: Intracellular redox imbalance and extracellular amino acid metabolic abnormality contribute to arsenic-induced developmental retardation in mouse preimplantation embryos. J Cell Physiol. 2010, 222 (2): 444-455. 10.1002/jcp.21966.CrossRefPubMed
33.
Kawamura Y, Uchijima Y, Horike N, Tonami K, Nishiyama K, Amano T, Asano T, Kurihara Y, Kurihara H: Sirt3 protects in vitro-fertilized mouse preimplantation embryos against oxidative stress-induced p53-mediated developmental arrest. J Clin Invest. 2010, 2;120 (8): 2817-2828.CrossRef
34.
Zinchuk V, Grossenbacher-Zinchuc O: Recent advances in quantitative colocalization analysis:focus on neuroscience. Prog Histochem Cytochem. 2009, 44: 125-172. 10.1016/j.proghi.2009.03.001.CrossRefPubMed
35.
Jurisicova A, Varmuza S, Casper R: Programmed cell death and human embryo fragmentation. Mol Hum Reprod. 1996, 2: 93-98. 10.1093/molehr/2.2.93.CrossRefPubMed
36.
Han Z, Mtango NR, Zhong Z, Vassena R, Latham KE: Early transcription from the maternal genome controlling blastomere integrity in mouse two-cell stage embryos. Am J Physiol. 2010, 298: 1235-1244. 10.1152/ajpcell.00393.2009.CrossRef
37.
Chi HJ, Koo JJ, Choi SY, Jeong HJ, Roh SI: Fragmentation of embryos is associated with both necrosis and apoptosis. Fertil & Steril. 2011, 96: 187-192. 10.1016/j.fertnstert.2011.04.020.CrossRef
38.
Vutyavanich T, Sreshthaputra O, Piromlertamorn W, Nunta S: Closed-system solid surface vitrification versus slow programmable freezing of mouse 2-cell embryos. J Assist Reprod Genet. 2009, 26: 285-290. 10.1007/s10815-009-9324-8.PubMedCentralCrossRefPubMed
39.
Tsang WH, Chow KL: Cryopreservation of Mammalian Embryos: Advancement of Putting Life on Hold. Birth Defects Res. 2010, 90: 163-175. 10.1002/bdrc.20186.CrossRef
40.
AbdelHafez F, Xu J, Goldberg J, Desai N: Vitrification in open and closed carriers at different cell stages: assessment of embryo survival, development, DNA integrity and stability during vapor phase storage for transport. BMC Biotechnol. 2011, 11: 1-10. 10.1186/1472-6750-11-1.CrossRef
41.
Piccoli C, Sardanelli A, Scrima R, Ripoli M, Quarato G, D'Aprile A, Bellomo F, Scacco S, De Michele G, Filla A, Iuso A, Boffoli D, Capitanio N, Papa S: Mitochondrial respiratory dysfunction in familiar parkinsonism associated with PINK1 mutation. Neurochem Res. 2008, 33 (12): 2565-2574. 10.1007/s11064-008-9729-2.CrossRefPubMed
42.
Dell’Aquila ME, Cho YS, Martino NA, Filioli Uranio M, Rutigliano L, Hinrichs K: OMICS for the identification of biomarkers for oocyte competence, with special reference to the mare as a prospective model for human reproductive medicine. Meiosis – Molecular Mechanism and Cytogenetic Diversity. Edited by: InTech. 2012, ISBN 978-953-51-0118-5
43.
Van Blerkom J: Mitochondria in early mammalian development. Semin Cell Dev Biol. 2009, 20: 354-364. 10.1016/j.semcdb.2008.12.005.CrossRefPubMed
44.
Perkins GA, Tjong J, Brown JM, Poquiz PH, Scott RT, Kolson DR, Ellisman MH, Spirou GA: The micro-architecture of mitochondria at active zones: electron tomography reveals novel anchoring scaffolds and cristae structured for high-rate metabolism. J Neurosci. 2010, 20;30 (3): 1015-1026.CrossRef
Metadata
Title
Vitrification preserves chromatin integrity, bioenergy potential and oxidative parameters in mouse embryos
Authors
Nicola A Martino
Maria E Dell’Aquila
Rosa A Cardone
Bence Somoskoi
Giovanni M Lacalandra
Sandor Cseh
Publication date
01-12-2013
Publisher
BioMed Central
Published in
Reproductive Biology and Endocrinology / Issue 1/2013
Electronic ISSN: 1477-7827
DOI
https://doi.org/10.1186/1477-7827-11-27

Other articles of this Issue 1/2013

Reproductive Biology and Endocrinology 1/2013 Go to the issue

Research

Assessment of cells in the ascitic fluid of women with ovarian hyperstimulation syndrome: the clinical implications for subsequent ovarian malignancy

Research

Associations among serum pro- and anti-inflammatory cytokines, metabolic mediators, body condition, and uterine disease in postpartum dairy cows

Research

Measuring thyroid peroxidase antibodies on the day nulliparous women present for management of miscarriage: a descriptive cohort study

Research

NF-kB overexpression and decreased immunoexpression of AR in the muscular layer is related to structural damages and apoptosis in cimetidine-treated rat vas deferens

Research

Levothyroxine and lung cancer in females: the importance of oxidative stress

Research

A retrospective study on IVF/ICSI outcome in patients with anti-nuclear antibodies: the effects of prednisone plus low-dose aspirin adjuvant treatment