Use of trypan blue staining to assess the quality of ovarian cryopreservation

Fauque, Patricia; Amor, Anis Ben; Joanne, Christiane; Agnani, G.; Bresson, Jean Luc; Roux, Christophe

doi:10.1016/j.fertnstert.2006.08.115

Cited by 58 publications

(57 citation statements)

References 50 publications

(37 reference statements)

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“…The follicles were thus cryopreserved without subsequent irreversible DNA damage which can be the consequence of apoptosis and/or oxidative stress activation [43,48]. These findings show clearly the beneficial effects of our procedure, since several studies in human reported a significant increase of DNA fragmentation in follicles after ovarian cortex cryopreservation with Gosden's procedure [12,33,48]. Supplementation of the collection medium as well as the freezing medium with taurine and L-glutamine, which have been found to play an antioxidant role by reducing cryopreservationinduced oxidative stress, might explain this result [4,18,40].…”

Section: Discussionmentioning

confidence: 59%

“…The morphology of follicles was well maintained (70.5±5.2 %) by our original protocol. For morphological examination of follicles, we used well-defined histological criteria unlike other investigators who considered atresia as the sole morphological consequence of follicular cryoinjuries [12,19]. This might explain the significant difference we observed between fresh and thawed morphologically intact follicles at resting stage.…”

Section: Discussionmentioning

confidence: 99%

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Quality and functionality of human ovarian tissue after cryopreservation using an original slow freezing procedure

Sanfilippo

Canis

Romero

et al. 2012

J Assist Reprod Genet

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Purpose To evaluate the efficiency of an original slow freezing protocol on the quality and function of human ovarian cortex. Methods Human ovarian tissues were cryopreserved using a freezing medium supplemented with propanediol and raffinose as cryoprotectants and antioxidants (L-glutamine, taurine). Samples were then frozen using a faster cooling rate than the usual one. Viability and morphology of follicles, DNA fragmentation in follicles and stroma as well as histology of the vascular endothelium were analyzed before and after freezing/thawing. Moreover, a functional analysis was performed based on the evaluation of follicular growth and development in thawed ovarian tissues that were cultured in vitro. Results Our freezing/thawing protocol allows preservation of a high proportion of viable follicles and the preservation of the different follicle developmental stages (p>0.05 versus fresh control). 70.5±5.2 % of follicles retained an intact morphology after cryopreservation (p=0.04). Stroma cells but not follicles exhibited a slight increase of DNA fragmentation after thawing (p<0.05). Microvessel endothelium within thawed tissues appeared to be preserved. Granulosa cells showed signs of proliferation in follicles cultured for 12 days. Secretion of 17β-oestradiol significantly increased during in vitro culture. Conclusions This protocol leads to good preservation of ovarian integrity and functionality post-thawing and thus appears as a suitable technique of ovarian tissue cryopreservation in clinical settings. Further research could be extended to optimize conditions of in vitro culture.

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Section: Discussionmentioning

confidence: 59%

Section: Discussionmentioning

confidence: 99%

Quality and functionality of human ovarian tissue after cryopreservation using an original slow freezing procedure

Sanfilippo

Canis

Romero

et al. 2012

J Assist Reprod Genet

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show abstract

“…In studies of cryopreserved ovarian cortical biopsies from PCOS patients a similar proportion of live follicles was seen [48]. Collectively, the results on human and ovine cryopreserved ovaries indicate a viability of around 80% of the immature follicles/cells.…”

Section: Discussionmentioning

confidence: 64%

Whole sheep ovary cryopreservation: evaluation of a slow freezing protocol with dimethylsulphoxide

Milenković

Wallin

Ghahremani

et al. 2010

J Assist Reprod Genet

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“…Various techniques to examine ovarian tissue viability have been reported in the literature, [8,28,34,[50][51][52][53] but a golden standard does not exist. With respect to the result from our current study, one should bear in mind that only small biopsies were available for histology and the Calcein assay, meaning that some sample bias with respect to the density of follicles in the tissue did occurr.…”

Section: Discussionmentioning

confidence: 99%