Qualitative and morphometric analysis of the ultrastructure of human oocytes cryopreserved by two alternative slow cooling protocols

Abstract

Purpose To ascertain possible cell damage from cryopreservation, the ultrastructure of human oocytes cryopreserved by slow cooling was assessed. Materials and methods Cryopreservation was performed through two protocols with one-step or two-step propanediol. Fresh control oocytes were examined for comparison. Samples were processed for transmission electron microscopy analysis. Results By light microscopy, both fresh and frozen-thawed oocytes appeared regularly rounded, with intact zona pellucida, and homogeneous cytoplasm. By electron microscopy observation, organelles were abundant and uniformly dispersed. Mitochondria-smooth endoplasmic reticulum associations appeared regular. However, both the amount and density of cortical granules appeared abnormally reduced in frozen-thawed samples. Slight to moderate vacuolization was also found in the ooplasm of oocytes of both frozen groups. Conclusions Slow cooling ensures a good overall preservation of human oocytes. However, cytoplasmic vacuolization and cortical granule loss appears associated with cryopreservation, irrespective of the protocol used.

Effect of vitrification on mitochondrial membrane potential in human metaphase II oocytes

“…Many studies reported that, cryopreservation can affect subcellular structure of oocytes such as chromosomes, spindle, plasmalemma, cortial granules, mitochondria, and so on [20][21][22]. ΔΨm is one of the most sensitive indices of freezing injury.…”

Ultrastructural markers of quality are impaired in human metaphase II aged oocytes: a comparison between reproductive and in vitro aging

“…After 2-5 days at 4°C, the samples were processed for light microscopy (LM) and TEM examination as described in [18][19][20]22].…”

“…In humans, vacuoles may occur commonly in response to a cellular injury, as for example in procedures of oocyte cryopreservation at MII stage [17][18][19][20]22].…”

“…Undoubtedly, transmission electron microscopy (TEM) and confocal microscopy cannot be used proactively in ART. However, TEM is effective in evaluating ooplasmic quality [17][18][19][20][21][22][23][24] and confocal microscopy is a technique of choice to assess nuclear (spindle) impairment [25]. Up to date, only rare reports described the ultrastructure of human, mature, metaphase II (MII), IvA oocytes [26,27].…”

Vitamin E-analog Trolox prevents endoplasmic reticulum stress in frozen-thawed ovarian tissue of capuchin monkey (Sapajus apella)

“…Cytoplasmic vacuolization is one of the most common signs of damage in cryopreserved oocytes (Nottola et al 2009;Coticchio et al 2010) and ovarian tissue ) and has been adopted as a specific marker for cryodamage (Coticchio et al 2010). Silva et al (2000) have suggested that damage to smooth endoplasmic reticulum (ER) leads to vacuolization.…”