title: Oocytes recovered after ovarian tissue slow freezing have impaired H2AX phosphorylation and functional competence.
abstract: It has been shown that oocytes isolated from ovarian tissue cryopreservation acquire DNA damage during the process of freeze-thawing. Using a mouse model, here we have investigated the functional competence and phosphorylation of H2AX (γ-H2AX) in germinal vesicle (GV) and parthenogenetically activated oocytes derived from conventional ovarian tissue slow freezing and vitrification techniques. The number of GV-stage oocytes with γ-H2AX foci was not significantly different between the slow-freezing and vitrification groups. Although the in vitro maturation (IVM) potential of GV oocytes in the slow-freezing group showed a significant delay (P<0.0001) in the process of germinal vesicle breakdown, no difference in the maturation rate was observed between the two protocols. Nevertheless, parthenogenetic activation of IVM oocytes using strontium chloride showed a significantly lower activation rate in the slow-freezing group compared with the vitrification (P<0.05) and control (P<0.01) groups. Importantly, H2AX phosphorylation was significantly perturbed in the slow-freezing group in comparison to the control (P<0.05). Therefore, we conclude that impaired sensing of DNA strand breaks and repair processes are associated with the reduced functional competence of the oocytes recovered from the slow-freezing group, which may have a significant impact on the reproductive outcome.
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynaecology, Kasturba Medical College, Manipal University, Manipal-576 104, India.
authors.forename: Sam
authors.initials: S
authors.lastname: Sudhakaran
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynaecology, Kasturba Medical College, Manipal University, Manipal-576 104, India.
authors.forename: Shubhashree
authors.initials: S
authors.lastname: Uppangala
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynaecology, Kasturba Medical College, Manipal University, Manipal-576 104, India.
authors.forename: Sujith Raj
authors.initials: SR
authors.lastname: Salian
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynaecology, Kasturba Medical College, Manipal University, Manipal-576 104, India.
authors.forename: Sachin D
authors.initials: SD
authors.lastname: Honguntikar
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynaecology, Kasturba Medical College, Manipal University, Manipal-576 104, India.
authors.forename: Ramya
authors.initials: R
authors.lastname: Nair
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynaecology, Kasturba Medical College, Manipal University, Manipal-576 104, India.
authors.forename: Guruprasad
authors.initials: G
authors.lastname: Kalthur
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynaecology, Kasturba Medical College, Manipal University, Manipal-576 104, India.
authors.forename: Satish Kumar
authors.initials: SK
authors.lastname: Adiga
mesh_terms: D000818:Animals; D015925:Cryopreservation / Q000379:methods*; D005260:Female; D006657:Histones / Q000378:metabolism*; D051379:Mice; D009865:Oocytes / Q000378:metabolism*; D010766:Phosphorylation; D058989:Vitrification
chemical_list: D006657:Histones; C475961:gamma-H2AX protein, mouse