R Gualtieri1, V Mollo, V Barbato, I Fiorentino, M Iaccarino, R Talevi. 1. Dipartimento di Biologia Strutturale e Funzionale, Università di Napoli 'Federico II', Complesso Universitario di Monte S Angelo, Via Cinthia, 80126 Napoli, Italy. roberto.gualtieri@unina.it
Abstract
BACKGROUND: The sensitivity of human oocytes to cryodamage may compromise their developmental competence following cryopreservation. Herein, we compared the ultrastructure and the response to the calcium (Ca²⁺) ionophore A23187 of fresh, slow-frozen and vitrified metaphase II (MII) human oocytes. METHODS: Supernumerary fresh MII oocytes, donated under written informed consent, were cryopreserved through either a slow cooling procedure based on propane-1,2-diol and 0.3 M sucrose or a closed vitrification system based on dimethylsulphoxide (DMSO) and ethylene glycol (EG). Ultrastructure of fresh and cryopreserved oocytes was assessed by transmission electron microscopy and compared through morphometrical analysis; intracellular calcium ([Ca²⁺](i)) dynamics was studied by evaluating the response to the Ca²⁺ ionophore A23187. RESULTS: Morphometric analysis demonstrated a markedly higher proportion of oocytes with large vacuoles, inward displacement of organelles from the pericortical toward the deep cytoplasm, and mitochondrial damage in slow-frozen compared with both fresh and vitrified oocytes. A23187 increased the [Ca²⁺](i) in all oocyte groups and the peak average increase in slow-frozen oocytes was significantly higher than in both fresh and vitrified oocytes. Moreover, the ability of slow-frozen oocytes to recover [Ca²⁺](i) to basal levels was significantly reduced compared with both fresh and vitrified oocytes. CONCLUSIONS: Closed vitrification based on DMSO and EG preserves the ultrastructural features and the ability to respond to the Ca²⁺ ionophore A23187 significantly better than does slow freezing with 0.3 M sucrose. Damage to organelles involved in the [Ca²⁺](i) modulation might reduce the developmental competence of cryopreserved oocytes.
BACKGROUND: The sensitivity of human oocytes to cryodamage may compromise their developmental competence following cryopreservation. Herein, we compared the ultrastructure and the response to the calcium (Ca²⁺) ionophore A23187 of fresh, slow-frozen and vitrified metaphase II (MII) human oocytes. METHODS: Supernumerary fresh MII oocytes, donated under written informed consent, were cryopreserved through either a slow cooling procedure based on propane-1,2-diol and 0.3 M sucrose or a closed vitrification system based on dimethylsulphoxide (DMSO) and ethylene glycol (EG). Ultrastructure of fresh and cryopreserved oocytes was assessed by transmission electron microscopy and compared through morphometrical analysis; intracellular calcium ([Ca²⁺](i)) dynamics was studied by evaluating the response to the Ca²⁺ ionophore A23187. RESULTS: Morphometric analysis demonstrated a markedly higher proportion of oocytes with large vacuoles, inward displacement of organelles from the pericortical toward the deep cytoplasm, and mitochondrial damage in slow-frozen compared with both fresh and vitrified oocytes. A23187 increased the [Ca²⁺](i) in all oocyte groups and the peak average increase in slow-frozen oocytes was significantly higher than in both fresh and vitrified oocytes. Moreover, the ability of slow-frozen oocytes to recover [Ca²⁺](i) to basal levels was significantly reduced compared with both fresh and vitrified oocytes. CONCLUSIONS: Closed vitrification based on DMSO and EG preserves the ultrastructural features and the ability to respond to the Ca²⁺ ionophore A23187 significantly better than does slow freezing with 0.3 M sucrose. Damage to organelles involved in the [Ca²⁺](i) modulation might reduce the developmental competence of cryopreserved oocytes.
Authors: E Fonseca; P Mesquita; C C Marques; M C Baptista; J Pimenta; J E Matos; G Soveral; R M L N Pereira Journal: Purinergic Signal Date: 2020-02-10 Impact factor: 3.765
Authors: Ana Arnanz; Neelke De Munck; Aşina Bayram; Ahmed El-Damen; Andrea Abdalla; Ibrahim ElKhatib; Laura Melado; Barbara Lawrenz; Human M Fatemi Journal: J Assist Reprod Genet Date: 2020-05-06 Impact factor: 3.412