title: Ovarian tissue vitrification is more efficient than slow freezing in protecting oocyte and granulosa cell DNA integrity.
abstract: Ovarian tissue cryopreservation is the primary treatment modality currently available to women at risk of losing their ovarian function due to cytotoxic therapy. However, the impact of these techniques on the oocyte DNA integrity is not elucidated. Here we have investigated the effect of vitrification and conventional slow freezing of eight week old Swiss albino mouse ovarian tissues on the oocyte and granulosa cell DNA integrity using the comet assay. The intracellular levels of reactive oxygen species in oocytes was measured by 2',7'-dichlorodihydrofluorescein diacetate fluorescence. The cryopreservation of ovarian tissue by the slow freezing technique resulted in a significantly higher level of DNA fragmentation in oocytes in comparison to vitrification (p < 0.05) whereas DNA fragmentation in granulosa cells was significantly higher than the control (p < 0.01). Further, reactive oxygen species were significantly elevated in oocytes derived from slow freezing when compared to vitrification (p < 0.05). Therefore, we conclude that the ovarian tissue slow freeze-thawing makes the oocyte and granulosa cells more vulnerable to DNA damage whereas vitrification appears to be a safer method than slow freezing for ovarian tissue cryopreservation.
authors.affiliation: Division of Clinical Embryology, Department of Obstetrics and Gynecology, Kasturba Medical College, Manipal University , Manipal , India.
authors.forename: Freya Jothsna
authors.initials: FJ
authors.lastname: Mathias
authors.forename: Fiona
authors.initials: F
authors.lastname: D'Souza
authors.forename: Shubhashree
authors.initials: S
authors.lastname: Uppangala
authors.forename: Sujith Raj
authors.initials: SR
authors.lastname: Salian
authors.forename: Guruprasad
authors.initials: G
authors.lastname: Kalthur
authors.forename: Satish Kumar
authors.initials: SK
authors.lastname: Adiga
mesh_terms: D000818:Animals; D020552:Comet Assay; D015925:Cryopreservation / Q000379:methods*; D053938:DNA Fragmentation; D005260:Female; D005615:Freezing / Q000009:adverse effects; D006107:Granulosa Cells / Q000378:metabolism / Q000473:pathology*; D051379:Mice; D009865:Oocytes / Q000378:metabolism / Q000473:pathology*; D010053:Ovary / Q000378:metabolism / Q000473:pathology*; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000378:metabolism; D013997:Time Factors; D058989:Vitrification
chemical_list: D017382:Reactive Oxygen Species
keywords: Cryopreservation; DNA damage; oocyte DNA; ovarian tissue; slow freezing; vitrification