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Lucia Olexiková
Linda Dujíčková
Alexander V. Makarevich
Elena Kubovičová
Peter Chrenek


Oocytes during cryopreservation are exposed to adverse conditions and factors that result in various damages reducing their developmental capacity. Most of such lesions may not be visible to light microscopy when assessing morphology. The aim of our study was to examine the ultrastructure of bovine in vitro matured (IVM) oocytes following cryopreservation. IVM oocytes were vitrified using ultra-rapid cooling technique in minimum volume on the electron microscopy grids and stored in liquid nitrogen for several weeks. After warming the oocytes were fixed, dehydrated and embedded into resin. Ultrastructure of oocytes was analysed on ultrathin sections obtained from embedded oocytes. Several alterations and damages to cell organelles of vitrified oocytes (smooth endoplasmic reticulum (SER), mitochondria and lipid droplets) were revealed in contrast to the fresh oocytes. Some membranes of large vesicles of SER were damaged and vesicles were obviously fused. Similarly, fusion of small lipid droplets to form large lipid droplets was visible in vitrified/warmed oocytes. Mitochondria showed signs of slight vacuolation, loss of mitochondrial matrix or less recognizable cristae. Differences were found also in the cortical area of oocytes (cortical granules, oolemma, zona pellucida and microvilli). However, these damages were less extensive than were presented in vitrified bovine oocytes previously, what indicates the suitability of our vitrification technique. In conclusion, ultrastructure assay can reveal individual membrane damages to organelles inside the oocyte, what may help in explaining developmental failures of oocytes following vitrification and warming.

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