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Cryopreservation and xenografting of human ovarian fragments: medulla decreases the phosphatidylserine translocation rate

Isachenko, V; Todorov, P; Isachenko, E; Rahimi, G; Hanstein, B; Salama, M; Mallmann, P; ... Merzenich, M; + view all (2016) Cryopreservation and xenografting of human ovarian fragments: medulla decreases the phosphatidylserine translocation rate. Reproductive Biology and Endocrinology , 14 , Article 79. 10.1186/s12958-016-0213-6. Green open access

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Abstract

Background: Phosphatidylserine is the phospholipid component which plays a key role in cell cycle signaling, specifically in regards to necrosis and apoptosis. When a cell affected by some negative factors, phosphatidylserine is no longer restricted to the intracellular side of membrane and can be translocated to the extracellular surface of the cell. Cryopreservation can induce translocation of phosphatidylserine in response to hypoxia, increasing intracellular Ca2+, osmotic disruption of cellular membranes, generation of reactive oxygen species and lipid peroxidation. As such the aim of this study was to test the level of phosphatidylserine translocation in frozen human medulla-contained and medulla-free ovarian tissue fragments. Methods: Ovarian fragments from twelve patients were divided into small pieces of two types, medulla-free cortex (Group 1, n = 42, 1.5–3.0 × 1.5–3.0 × 0.5–0.8 mm) and cortex with medulla (Group 2, n = 42, 1.5–3.0 × 1.5–3.0 × 1.5–2.0 mm), pre-cooled after operative removal to 5 °C for 24 h and then conventionally frozen with 6 % dimethyl sulfoxide, 6 % ethylene glycol and 0.15 M sucrose in standard 5-ml cryo-vials. After thawing at +100 °C and step-wise removal of cryoprotectants in 0.5 M sucrose, ovarian pieces were xenografted to SCID mice for 45 days. The efficacy of tissues cryopreservation, taking into account the presence or absence of medulla, was evaluated by the development of follicles (histology with hematoxylin-eosin) and through the intensity of translocation of phosphatidylserine (FACS with FITC-Annexin V and Propidium Iodide). Results: For Groups 1 and 2, the mean densities of follicles per 1 mm3 were 9.8, and 9.0, respectively. In these groups, 90 and 90 % preantral follicles appeared morphologically normal. However, FACS analysis showed a significantly decreased intensity of translocation of phosphatidylserine (FITC-Annexin V positive) after cryopreservation of tissue with medulla (Group 2, 59.6 %), in contrast with tissue frozen without medulla (Group 1, 78.0 %, P < 0.05). In Groups 1 and 2 it was detected that 21.6 and 40.0 % cells were viable (FITC-Annexin V negative, Propidium Iodide negative). Conclusion: The presence of medulla in ovarian pieces is beneficial for post-thaw development of cryopreserved human ovarian tissue.

Type: Article
Title: Cryopreservation and xenografting of human ovarian fragments: medulla decreases the phosphatidylserine translocation rate
Open access status: An open access version is available from UCL Discovery
DOI: 10.1186/s12958-016-0213-6
Publisher version: http://dx.doi.org/10.1186/s12958-016-0213-6
Language: English
Additional information: Copyright © The Author(s) 2016. This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Keywords: Cancer, Cryopreservation, Human ovarian tissue, Phosphatidylserine translocation, Medulla, Xenotransplantation
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL EGA Institute for Womens Health
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL EGA Institute for Womens Health > Reproductive Health
URI: https://discovery.ucl.ac.uk/id/eprint/1537082
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