title: Confocal laser scanning microscopy analysis of bioenergetic potential and oxidative stress in fresh and frozen-thawed human ovarian tissue from oncologic patients.
abstract: OBJECTIVE
  To evaluate the effectiveness of a bioenergy/oxidative stress assessment based on confocal laser scanning microscopy (CLSM) in association with morphology and ultrastructure analyses based on light microscopy (LM) and transmission electron microscopy (TEM), to monitor the preservation status of cryopreserved human ovarian tissue from cancer patients.
  
  
  METHODS
  Prospective study.
  
  
  METHODS
  University hospital.
  
  
  METHODS
  Fourteen young cancer patients.
  
  
  METHODS
  Human ovarian tissue biopsy, slow freezing/rapid thawing, LM, TEM, CLSM assessment of mitochondrial distribution and activity, and intracellular reactive oxygen species (ROS) localization and levels.
  
  
  METHODS
  In tissue examined before and after slow freezing/rapid thawing, follicular and stromal LM-based score of morphologic damage, ultrastructure, mitochondrial distribution pattern, reactive oxygen species (ROS) localization; mean ± standard deviation of stromal mitochondrial activity and ROS levels.
  
  
  RESULTS
  Severe (n = 6 patients), slight (n = 6 patients), or no (n = 2 patients) LM/TEM-based damage was found in fresh tissue. After freezing/thawing, no further morphologic/ultrastructural alterations were found; however, statistically significant reductions, increases, or no changes in mitochondrial activity and ROS levels were found in severely, slightly, and undamaged tissue, respectively.
  
  
  CONCLUSIONS
  Bioenergy/oxidative functional damage was found in tissue with severe LM/TEM-assessed damage. In tissue with slight LM/TEM-assessed damage, the CLSM-based bioenergy/oxidative stress assessment was the only test that allowed discrimination between tissue that had been better (low/no difference) or worse preserved (significant differences).
authors.affiliation: Gynaecology and Pathophysiology of Human Reproduction Unit, Sant'Orsola-Malpighi Hospital, DIMEC, University of Bologna, Bologna, Italy. Electronic address: raffaella.fabbri@unibo.it.
authors.forename: Raffaella
authors.initials: R
authors.lastname: Fabbri
authors.affiliation: Gynaecology and Pathophysiology of Human Reproduction Unit, Sant'Orsola-Malpighi Hospital, DIMEC, University of Bologna, Bologna, Italy.
authors.forename: Rossella
authors.initials: R
authors.lastname: Vicenti
authors.affiliation: Veterinary Clinics and Animal Productions Unit, Department of Emergency and Organ Transplantation (DETO), University of Bari Aldo Moro, Valenzano, Bari, Italy.
authors.forename: Nicola Antonio
authors.initials: NA
authors.lastname: Martino
authors.affiliation: Veterinary Clinics and Animal Productions Unit, Department of Emergency and Organ Transplantation (DETO), University of Bari Aldo Moro, Valenzano, Bari, Italy.
authors.forename: Maria Elena
authors.initials: ME
authors.lastname: Dell'Aquila
authors.affiliation: Surgical Pathology, Sant'Orsola-Malpighi Hospital, DIMES, University of Bologna, Bologna, Italy.
authors.forename: Gianandrea
authors.initials: G
authors.lastname: Pasquinelli
authors.affiliation: Gynaecology and Pathophysiology of Human Reproduction Unit, Sant'Orsola-Malpighi Hospital, DIMEC, University of Bologna, Bologna, Italy.
authors.forename: Maria
authors.initials: M
authors.lastname: Macciocca
authors.affiliation: Gynaecology and Pathophysiology of Human Reproduction Unit, Sant'Orsola-Malpighi Hospital, DIMEC, University of Bologna, Bologna, Italy.
authors.forename: Valentina
authors.initials: V
authors.lastname: Magnani
authors.affiliation: Gynaecology and Pathophysiology of Human Reproduction Unit, Sant'Orsola-Malpighi Hospital, DIMEC, University of Bologna, Bologna, Italy.
authors.forename: Roberto
authors.initials: R
authors.lastname: Paradisi
authors.affiliation: Gynaecology and Pathophysiology of Human Reproduction Unit, Sant'Orsola-Malpighi Hospital, DIMEC, University of Bologna, Bologna, Italy.
authors.forename: Stefano
authors.initials: S
authors.lastname: Venturoli
mesh_terms: D000293:Adolescent; D000328:Adult; D015925:Cryopreservation / Q000379:methods; D004734:Energy Metabolism / Q000502:physiology*; D005260:Female; D006801:Humans; D018613:Microscopy, Confocal / Q000379:methods; D010051:Ovarian Neoplasms / Q000378:metabolism* / Q000473:pathology; D010053:Ovary / Q000378:metabolism* / Q000473:pathology*; D018384:Oxidative Stress / Q000502:physiology*; D011446:Prospective Studies; D017382:Reactive Oxygen Species / Q000378:metabolism; D055815:Young Adult
chemical_list: D017382:Reactive Oxygen Species
keywords: Bioenergy/oxidative stress assessment; confocal laser scanning microscopy; human ovarian tissue cryopreservation by slow freezing/rapid thawing; light microscopy histology; transmission electron microscopy