title: Three dimensional in vitro culture of preantral follicles following slow-freezing and vitrification of mouse ovarian tissue.
abstract: To evaluate the effects slow-freezing and vitrification on three dimensional in vitro culture of preantral follicles, ovaries of 12-14 days old female NMRI mice were isolated and randomly assigned to fresh control, slow-freezing and vitrification groups. Slow-freezing was performed using programmable freezer. Vitrification was carried out in a medium consisting of ethylene glycol (EG) and dimethyl sulphoxide (Me2SO) by needle immersion method. middle sized preantral follicles were mechanically isolated and cultured for 12 days in 0.7% sodium alginate gel. The follicles development and quantitative expression of oocyte specific genes (Bmp15, Gdf9, Fgf8) and the growth related genes (Igf1, Kit, Kit-l) were assessed after 1, 8 and 12 days of culture. Both cryopreserved groups showed reduction of follicular survival rates compared to the control group on days 8 and 12 of culture (P < 0.05). Antrum formation rates reduced in slow-freezing after 12 days of culture (P < 0.05). Evaluation of gene expression showed reduction of Bmp15, Gdf9, Fgf8, Kit and Kit-l during 12 days of culture (P < 0.05). Kit and Kit-l expression in slow-freezing group significantly reduced on day 8 of culture (p < 0.05). Igf1 expression was lower in slow-freezing group on 1st day of culture than vitrification and control groups (P < 0.05). Finally, intergroup comparison showed same expression pattern of genes after 12 days of culture. Thus, cryopreservation of mouse ovaries by both methods can preserve most developmental parameters and expression of maturation genes. However, vitrification is a better method for cryopreservation of mouse ovaries due to greater antrum formation and expression of growth related markers.
authors.affiliation: Department of Anatomy, Faculty of Medical Sciences, Tarbiat Modares University, P.O. Box14115-111, Tehran, Iran.
authors.forename: Fatemeh
authors.initials: F
authors.lastname: Asgari
authors.affiliation: Department of Anatomy, Faculty of Medical Sciences, Tarbiat Modares University, P.O. Box14115-111, Tehran, Iran; Department of Embryology, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran. Electronic address: mr_valojerdi@modares.ac.ir.
authors.forename: Mojtaba Rezazadeh
authors.initials: MR
authors.lastname: Valojerdi
authors.affiliation: Department of Embryology, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran.
authors.forename: Bita
authors.initials: B
authors.lastname: Ebrahimi
authors.affiliation: Department of Embryology, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran.
authors.forename: Roya
authors.initials: R
authors.lastname: Fatehi
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D005260:Female; D005615:Freezing; D051379:Mice; D009865:Oocytes / Q000187:drug effects / Q000254:growth & development; D006080:Ovarian Follicle / Q000187:drug effects; D058989:Vitrification
chemical_list: D003451:Cryoprotective Agents
keywords: Ovary cryopreservation; Preantral follicle; Three dimensional in vitro culture