Long-Term Advantages of Ovarian Reserve Maintenance and Follicle Development Using Adipose Tissue-Derived Stem Cells in Ovarian Tissue Transplantation

Cacciottola, Luciana; Nguyen, Thu Yen Thi; Chiti, Maria Costanza; Camboni, Alessandra; Amorim, Christiani Andrade; Donnez, Jacques; Dolmans, Marie-Madeleine

doi:10.3390/jcm9092980

Cited by 16 publications

(16 citation statements)

References 49 publications

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“…Statistical significance was expressed using different letters (p < 0.05). Scale bar: 20 μm preserved after 6 months of grafting with ASCs, showing similar endocrine function resumption to the standard transplantation procedure [29]. In the present study, we were able to prove that ASCs have a positive impact on grafted tissue by significantly reducing follicle loss both 3 and 10 days after grafting.…”

Section: Discussionsupporting

confidence: 73%

“…Increased primordial follicle activation observed soon after ovarian tissue transplantation has been associated with activation of the PI3K/Akt pathway [14,43]. Maintenance of the quiescent state appears to be related to FOXO1-3 activity, whose cytoplasmic shift is related to the start of cross-talk between the oocyte and granulosa cells before effective follicle growth, namely transformation from flattened to cuboidal granulosa cells [29]. Indeed, its constitutive presence or absence in specific knockout animal models was found to be responsible for primordial follicle recruitment, but not further follicle growth [44,45].…”

Section: Follicle Activationmentioning

confidence: 99%

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Adipose tissue–derived stem cells protect the primordial follicle pool from both direct follicle death and abnormal activation after ovarian tissue transplantation

Cacciottola

Courtoy

Nguyen

et al. 2020

J Assist Reprod Genet

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Purpose To investigate whether adipose tissue-derived stem cells (ASCs) protect the primordial follicle pool, not only by decreasing direct follicle loss but also by modulating follicle activation pathways. Methods Twenty nude mice were grafted with frozen-thawed human ovarian tissue from 5 patients. Ten mice underwent standard ovarian tissue transplantation (OT group), while the remaining ten were transplanted with ASCs and ovarian tissue (2-step/ASCs+OT group). Ovarian grafts were retrieved on days 3 (n = 5) and 10 (n = 5). Analyses included histology (follicle count and classification), immunohistochemistry (c-caspase-3 for apoptosis and LC3B for autophagy), and immunofluorescence (FOXO1 for PI3K/Akt activation and YAP for Hippo pathway disruption). Subcellular localization was determined in primordial follicles on high-resolution images using structured illumination microscopy. Results The ASCs+OT group showed significantly higher follicle density than the OT group (p = 0.01). Significantly increased follicle atresia (p < 0.001) and apoptosis (p = 0.001) were observed only in the OT group. In primordial follicles, there was a significant shift in FOXO1 to a cytoplasmic localization in the OT group on days 3 (p = 0.01) and 10 (p = 0.03), indicating follicle activation, while the ASCs+OT group and non-grafted controls maintained nuclear localization, indicating quiescence. Hippo pathway disruption was encountered in primordial follicles irrespective of transplantation, with nuclear YAP localized in their granulosa cells. Conclusion We demonstrate that ASCs exert positive effects on the ovarian reserve, not only by protecting primordial follicles from direct death but also by maintaining their quiescence through modulation of the PI3K/Akt pathway.

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Section: Discussionsupporting

confidence: 73%

Section: Follicle Activationmentioning

confidence: 99%

Adipose tissue–derived stem cells protect the primordial follicle pool from both direct follicle death and abnormal activation after ovarian tissue transplantation

Cacciottola

Courtoy

Nguyen

et al. 2020

J Assist Reprod Genet

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“…The preparation of the transplantation site using ASCs has been proven to decrease hypoxia in grafted tissues (22) by boosting revascularization through ASC differentiation into endothelial-like lineages and growth factor secretion (24). This ensures both the protection of preexisting vessels in the host component and the earlier migration of endothelial progenitors from the graft component, as shown in our findings, resulting in enhanced primordial follicle survival after short-term (22,23) and long-term (38) grafting. In the light of these results, we believe it is crucial to investigate the mechanisms by which ASCs have a positive impact on ovarian tissue recovery from the potential injury caused by either hypoxia or oxidative stress after OTT.…”

Section: Discussionsupporting

confidence: 62%

“…Boosting neoangiogenesis has proven to be effective in enhancing follicle outcomes in other experimental models that administered VEGF and fibroblast growth factor after OTT (43,44). ASCs exert a protective effect on the follicle pool in grafted ovarian tissues, as widely demonstrated (22,24,38,45), because of their ability to shorten the hypoxic period responsible for follicle loss that occurs soon after transplantation.…”

Section: Figurementioning

confidence: 97%

Modulating hypoxia and oxidative stress in human xenografts using adipose tissue-derived stem cells

et al. 2021

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Objective: To investigate whether adipose tissue-derived stem cells (ASCs) modulate hypoxia and oxidative stress in human ovarian tissue transplants. Design: Prospective experimental study Setting: Gynecological research unit in a university hospital Patient(s): Cryopreserved ovarian cortex from 5 adult women. Intervention(s): Thirty mice were grafted with frozen-thawed human ovarian tissue, with or without ASCs (2-step/ASCsþovarian tissue [OT] group and OT group). The ovarian grafts were retrieved on days 3 (n ¼ 5), 10 (n ¼ 5), and 21 (n ¼ 5). The 10 animals grafted for 21 days underwent in vivo evaluations using microdialysis. One piece of ovarian tissue per patient was fixed for analysis after thawing (non-grafted controls). Main Outcome Measure(s): Direct reactive oxygen species were collected every second day after grafting by means of microdialysis. Analyses of ovarian fragments included immunolabeling for double CD34 (revascularization by host and graft components); immunofluorescence for hypoxia-inducible factor 1a (hypoxia-related response), nuclear factor erythroid 2-related factor 2 (oxidative stressrelated response), and 8-hydroxy-deoxyguanosine (oxidative stress-related DNA damage); and gene expression (quantitative reverse transcription polymerase chain reaction) for vascular endothelial growth factor-A (neoangiogenesis), superoxide dismutase 2 (antioxidant activity), and nuclear respiratory factor 1 (mitochondrial biogenesis). Result(s): Reactive oxygen species peaked earlier in the ASC group (day 2) compared with that in the OT group (day 10) after grafting. Total vascularization was stable in the ASC group at all time points, while it was lower in the OT group 3 days after grafting. Hypoxiainducible factor 1a expression, also detected in non-grafted controls, was significantly lower in the ASC group than in the OT group on days 3 and 10. The increase in VEGF gene expression lasted significantly longer in the ASC group than in the OT group. There was no significant upturn in the oxidative stress-related response (nuclear factor erythroid 2-related factor 2 pathway) or oocyte DNA damage (8-hydroxy-deoxyguanosine) in any of the grafted groups. Conclusion(s): Use of ASCs allows faster ovarian graft reperfusion and mitigates the hypoxia-related response through rapid revascularization, sustained by prolonged increase in vascular endothelial growth factor after grafting. No evidence of oxidative stress-related damage was detected irrespective of the transplantation strategy.

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“…Enhancement of graft revascularization by delivering both angiogenic and antiapoptotic factors [20][21][22][23][24] or antioxidants to decrease oxidative stress [25][26][27] has led to limited improvements in graft outcomes [9]. Recently, the use of adipose tissue-derived stem cells in an experimental mice model of one or two-step transplantation strategy evidenced a real improvement of oxygenation and vascularization of the ovarian tissue in the early post-transplantation period leading to a better follicular survival [28][29][30]. To improve the lifetime of auto-transplanted fragments and ultimately enhance the pregnancy rate among young patients who are cured of cancer, supplemental studies are clearly required to identify additional strategies to limit follicular burnout.…”

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confidence: 99%

Pharmacological inhibition of the PI3K/PTEN/Akt and mTOR signalling pathways limits follicle activation induced by ovarian cryopreservation and in vitro culture

2021

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Background Cryopreservation and transplantation of ovarian tissue (OTCTP) represent a promising fertility preservation technique for prepubertal patients or for patients requiring urgent oncological management. However, a major obstacle of this technique is follicle loss due to, among others, accelerated recruitment of primordial follicles during the transplantation process, leading to follicular reserve loss in the graft and thereby potentially reducing its lifespan. This study aimed to assess how cryopreservation itself impacts follicle activation. Results Western blot analysis of the PI3K/PTEN/Akt and mTOR signalling pathways showed that they were activated in mature or juvenile slow-frozen murine ovaries compared to control fresh ovaries. The use of pharmacological inhibitors of follicle signalling pathways during the cryopreservation process decreased cryopreservation-induced follicle recruitment. The second aim of this study was to use in vitro organotypic culture of cryopreserved ovaries and to test pharmacological inhibitors of the PI3K/PTEN/Akt and mTOR pathways. In vitro organotypic culture-induced activation of the PI3K/PTEN/Akt pathway is counteracted by cryopreservation with rapamycin and in vitro culture in the presence of LY294002. These results were confirmed by follicle density quantifications. Indeed, follicle development is affected by in vitro organotypic culture, and PI3K/PTEN/Akt and mTOR pharmacological inhibitors preserve primordial follicle reserve. Conclusions Our findings support the hypothesis that inhibitors of mTOR and PI3K might be an attractive tool to delay primordial follicle activation induced by cryopreservation and culture, thus preserving the ovarian reserve while retaining follicles in a functionally integrated state.

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Long-Term Advantages of Ovarian Reserve Maintenance and Follicle Development Using Adipose Tissue-Derived Stem Cells in Ovarian Tissue Transplantation

Cited by 16 publications

References 49 publications

Adipose tissue–derived stem cells protect the primordial follicle pool from both direct follicle death and abnormal activation after ovarian tissue transplantation

Adipose tissue–derived stem cells protect the primordial follicle pool from both direct follicle death and abnormal activation after ovarian tissue transplantation

Modulating hypoxia and oxidative stress in human xenografts using adipose tissue-derived stem cells

Pharmacological inhibition of the PI3K/PTEN/Akt and mTOR signalling pathways limits follicle activation induced by ovarian cryopreservation and in vitro culture

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