title: Different chromatin and energy/redox responses of mouse morulae and blastocysts to slow freezing and vitrification.
abstract: BACKGROUND
  The ability to cryopreserve mammalian embryos has become an integral part of assisted reproduction, both in human and veterinary medicine. Despite differences in the size and physiological characteristics of embryos from various species, the embryos have been frozen by either of two procedures: slow freezing or vitrification. The aim of our study was to compare the effect of slow freezing and vitrification to the chromatin structure, energy status and reactive oxygen species production of mouse morulae and blastocysts.
  
  
  METHODS
  Mouse morulae and blastocysts were randomly allocated into vitrification, slow freezing and control groups. For slow freezing, Dulbecco phosphate buffered saline based 10% glicerol solution was used. For vitrification, G-MOPS™ based solution supplemented with 16% ethylene glycol, 16% propylene glycol, Ficoll (10 mg/ml) and sucrose (0.65 mol/l) was used. After warming, the chromatin integrity, mitochondrial distribution pattern and energy/oxidative status were compared among groups.
  
  
  RESULTS
  Cryopreservation affected chromatin integrity at a greater extent at the morula than the blastocyst stage. Chromatin damage induced by slow freezing was more relevant compared to vitrification. Slow freezing and vitrification similarly affected mitochondrial distribution pattern. Greater damage was observed at the morula stage and it was associated with embryo grade. Cryopreservation altered the quantitative bioenergy/redox parameters at a greater extent in the morulae than in the blastocysts. Effects induced by slow freezing were not related to embryo grade or mitochondrial pattern, as affected embryos were of all grades and with both mitochondrial patterns. However, effects induced by vitrification were related to mitochondrial pattern, as only embryos with homogeneous mitochondrial pattern in small aggregates had reduced energy status.
  
  
  CONCLUSIONS
  This study shows for the first time the joint assessment of chromatin damage and mitochondrial energy/redox potential in fresh and frozen mouse embryos at the morula and blastocyst stage, allowing the comparison of the effects of the two most commonly used cryopreservation procedures.
authors.affiliation: Department and Clinic of Obstetrics and Reproduction, Szent Istvan University, Budapest, Hungary. somoskoibence@gmail.com.
authors.forename: Bence
authors.initials: B
authors.lastname: Somoskoi
authors.affiliation: Veterinary Clinics and Animal Productions Unit, Department of Emergency and Organ Trasplantation (DETO), University of Bari Aldo Moro Valenzano, Bari, Italy. nicmartino@libero.it.
authors.forename: Nicola A
authors.initials: NA
authors.lastname: Martino
authors.affiliation: Department of Bioscience, Biotechnology and Pharmacological Science, University of Bari, 70126, Bari, Italy. r.cardone@biologia.uniba.it.
authors.forename: Rosa A
authors.initials: RA
authors.lastname: Cardone
authors.affiliation: Veterinary Clinics and Animal Productions Unit, Department of Emergency and Organ Trasplantation (DETO), University of Bari Aldo Moro Valenzano, Bari, Italy. giovannimichele.lacalandra@uniba.it.
authors.forename: Giovanni M
authors.initials: GM
authors.lastname: Lacalandra
authors.affiliation: Department of Bioscience, Biotechnology and Pharmacological Science, University of Bari, 70126, Bari, Italy. mariaelena.dellaquila@uniba.it.
authors.forename: Maria E
authors.initials: ME
authors.lastname: Dell'Aquila
authors.affiliation: Department and Clinic of Obstetrics and Reproduction, Szent Istvan University, Budapest, Hungary. cseh.sandor@aotk.szie.hu.
authors.forename: Sandor
authors.initials: S
authors.lastname: Cseh
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000378:metabolism / Q000502:physiology*; D002843:Chromatin / Q000378:metabolism* / Q000502:physiology; D015925:Cryopreservation / Q000379:methods*; D004622:Embryo, Mammalian / Q000378:metabolism / Q000502:physiology; D005260:Female; D005615:Freezing; D051379:Mice; D008928:Mitochondria / Q000378:metabolism / Q000502:physiology; D009028:Morula / Q000378:metabolism / Q000502:physiology*; D010084:Oxidation-Reduction; D017382:Reactive Oxygen Species / Q000378:metabolism; D058989:Vitrification
chemical_list: D002843:Chromatin; D017382:Reactive Oxygen Species