Vitrification versus controlled-rate freezing in cryopreservation of human ovarian tissue

Keros, Victoria; Xella, Susanna; Hultenby, Kjell; Pettersson, Karin; Sheikhi, Maryam; Volpe, Annibale; Hreinsson, Julius; Hovatta, Outi

doi:10.1093/humrep/dep079

Cited by 294 publications

(267 citation statements)

References 49 publications

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“…First, we examined two freezing methods and several cryoprotectants by evaluating the spermatogenic potential of tissues following different methods of cryopreservation. The two freezing methods were slow freezing and vitrification, the latter of which is now widely used in clinical settings for oocytes, embryos and ovarian tissues [12][13][14] . As cryoprotectants, dimethylsulphoxide (DMSO), propanediol (PROH) and Cell Banker 1 (CB) were used for slow freezing and Stem Cell Keep (SCK) was used for vitrification.…”

Section: Resultsmentioning

confidence: 99%

Offspring production with sperm grown in vitro from cryopreserved testis tissues

et al. 2014

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With the increasing cure rate of paediatric cancers, infertility, as one of the adverse effects of treatments, has become an important concern for patients and their families. Since semen cryopreservation is applicable only for post-pubertal patients, alternative pre-pubertal measures are necessary. Here we demonstrate that testis tissue cryopreservation is a realistic measure for preserving the fertility of an individual. Testis tissues of neonatal mice were cryopreserved either by slow freezing or by vitrification. After thawing, they were cultured on agarose gel and showed spermatogenesis up to sperm formation. Microinsemination was performed with round spermatids and sperm, leading to eight offspring in total. They grew healthily and produced progeny upon natural mating between them. This strategy, the cryopreservation of testis tissues followed by in vitro spermatogenesis, is promising to preserve the fertility of male paediatric cancer patients in the future.

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Section: Resultsmentioning

confidence: 99%

Offspring production with sperm grown in vitro from cryopreserved testis tissues

et al. 2014

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“…In vitro culture experiments were conducted using thawed ovarian samples from 5 patients, presenting the higher number of follicles according to the previous histological examination. As shown in previous studies, culture of ovarian pieces allows cellular interactions between follicles and surrounding stroma indispensable to sustain follicular growth initiation [8,25,34]. In order to retain the 3D structure of the follicles, a low-attachment culture system limiting stroma cell adhesion on the culture support was used.…”

Section: Discussionmentioning

confidence: 99%

“…Morphology of follicles was evaluated on the basis of parameters previously described by Keros et al [25]. Follicles were classified as intact if no overt signs of oocyte and GCs degeneration were noted.…”

Section: Follicle Viability Assessmentmentioning

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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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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“…Vitrification is a simple, convenient, and efficient method for embryo and oocyte cryopreservation in both animal and human models (Fujihira et al 2004, Campos-Chillò n et al 2009, Cao et al 2009, Keros et al 2009, Selman et al 2009, Zhou et al 2010. However, data on ovarian tissue vitrification are still limited, and the results are often conflicting and controversial (Choi et al 2008, Isachenko et al 2009, Kagawa et al 2009.…”

Section: Discussionmentioning

confidence: 99%

“…Lower concentrations of cryoprotectant are required in freezing media for slow cooling, which reduces the risk of toxic and osmotic damage to cells, but this is insufficient to prevent ice crystal formation jeopardizing cell survival during the freezing procedures (Vajta 2006). Vitrification, though still regarded as a relatively novel freezing method, has recently been reported as an effective alternative method for the cryopreservation of ovarian tissue in various species, including mouse (Wang et al 2009), rat (Deng et al 2009), pig (Gandolfi et al 2006), goat (Santos et al 2007), sheep (Al-aghbari & Menino 2002, Courbiere et al 2006, monkey (Yeoman et al 2005), and human (Keros et al 2009, Zhou et al 2010. It combines a high cooling rate with a high concentration of cryoprotectant in the vitrification media, which rapidly dehydrates the cells and prevents water from precipitating as ice (Vajta 2006).…”

Section: Introductionmentioning

confidence: 99%

Successful in vitro culture of pre-antral follicles derived from vitrified murine ovarian tissue: oocyte maturation, fertilization, and live births

Wang¹,

Catt²,

Pangestu³

et al. 2011

Reproduction

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Cryopreservation of ovarian tissue is an important option for preserving the fertility of cancer patients undergoing chemotherapy and radiotherapy. In this study, we examined the viability and function of oocytes derived in vitro from pre-antral follicles as an alternative method for restoring fertility. Pre-antral follicles (specified as secondary follicle with a diameter around 100-130 mm) were mechanically isolated from vitrified-warmed and fresh adult mouse ovarian tissues and cultured for 12 days followed by an ovulation induction protocol at the end of this period to initiate oocyte maturation. Oocytes were then released from these follicles, fertilized in vitro, and cultured to the blastocyst stage and vitrified. After storage in liquid nitrogen for 2 weeks, groups of vitrified blastocysts were warmed and transferred into pseudo-pregnant recipient females. Although most of the isolated mouse pre-antral follicles from fresh (79.4%) and vitrified (75.0%) ovarian tissues survived the 12-day in vitro culture period, significantly fewer mature oocytes developed from vitrifiedwarmed pre-antral follicles than from the fresh controls (62.2 vs 86.4%, P!0.05). No difference was observed in embryo cleavage rates between these two groups, but the proportion of embryos that developed into blastocysts in the vitrification group was only half that of the controls (24.2 vs 47.2%, P!0.05). Nevertheless, live births of healthy normal pups were achieved after transfer of vitrified blastocysts derived from both experimental groups. This study shows that successful production of healthy offspring using an in vitro follicle culture system is feasible, and suggests that this procedure could be used in cancer patients who wish to preserve their fertility using ovarian tissue cryopreservation.

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Vitrification versus controlled-rate freezing in cryopreservation of human ovarian tissue

Abstract: Vitrification using a combination of PrOH, EG, DMSO and PVP was comparable to slow freezing in terms of preserving follicles in human ovarian tissue. Ovarian stroma had significantly better morphological integrity after vitrification than after controlled-rate freezing.

Cited by 294 publications

References 49 publications

Offspring production with sperm grown in vitro from cryopreserved testis tissues

Offspring production with sperm grown in vitro from cryopreserved testis tissues

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

Successful in vitro culture of pre-antral follicles derived from vitrified murine ovarian tissue: oocyte maturation, fertilization, and live births

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