Mitochondria, fundamental organelles in cell metabolism, and ATP synthesis are responsible for generating reactive oxygen species (ROS), calcium homeostasis, and cell death. Mitochondria produce most ROS, and when levels exceed the antioxidant defenses, oxidative stress (OS) is generated. These changes may eventually impair the electron transport chain, resulting in decreased ATP synthesis, increased ROS production, altered mitochondrial membrane permeability, and disruption of calcium homeostasis. Mitochondria play a key role in the gamete competence to facilitate normal embryo development. However, iatrogenic factors in assisted reproductive technologies (ART) may affect their functional competence, leading to an abnormal reproductive outcome. Cryopreservation, a fundamental technology in ART, may compromise mitochondrial function leading to elevated intracellular OS that decreases sperm and oocytes’ competence and the dynamics of fertilization and embryo development. This article aims to review the role played by mitochondria and ROS in sperm and oocyte function and the close, biunivocal relationships between mitochondrial damage and ROS generation during cryopreservation of gametes and gonadal tissues in different species. Based on current literature, we propose tentative hypothesis of mechanisms involved in cryopreservation-associated mitochondrial dysfunction in gametes, and discuss the role played by antioxidants and other agents to retain the competence of cryopreserved reproductive cells and tissues.

Reactive oxygen species (ROS) generated at low levels during mitochondrial respiration have key roles in several signaling pathways. Oxidative stress (OS) arises when the generation of ROS exceeds the cell’s antioxidant scavenging ability and leads to cell damage. Physiological ROS production in spermatozoa regulates essential functional characteristics such as motility, capacitation, acrosome reaction, hyperactivation, and sperm-oocyte fusion. OS can have detrimental effects on sperm function through lipid peroxidation, protein damage, and DNA strand breakage, which can eventually affect the fertility of an individual. Substantial evidence in the literature indicates that spermatozoa experiencing OS during in vitro manipulation procedures in human- and animal-assisted reproduction are increasingly associated with iatrogenic ROS production and eventual impairment of sperm function. Although a direct association between sperm OS and human assisted reproductive techniques (ART) outcomes after in vitro fertilization (IVF) and/or intracytoplasmic sperm injection (ICSI) is still a matter of debate, studies in animal models provide enough evidence on the adverse effects of sperm OS in vitro and defective fertilization and embryo development. This review summarized the literature on sperm OS in vitro, its effects on functional ability and embryo development, and the approaches that have been proposed to reduce iatrogenic sperm damage and altered embryonic development.

Spermatozoa are extremely vulnerable to oxidative stress caused by the unbalance between concentrations of reactive oxygen species and antioxidant scavenging systems present inside the male reproductive tract. In spite of a large number of clinical studies that claimed the beneficial effects of antioxidant oral administration on sperm physiology and fertility, only a few studies were addressed to evaluate their effects on spermatozoa in vitro. Main aims of the present study were to assess the influence of zinc, D-aspartate and coenzyme Q10, included in the dietary supplement Genadis (Merck Serono), on human sperm motility, DNA fragmentation and lipid peroxidation.

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. 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. 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. 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.