Cryopreservation of sheep ovarian tissue: Effect of different cryoprotectants and equilibration regimen on ovarian follicles structure

The ovaries resected from twenty-nine day old female rats primed with 10 IU of PMS for 48 hours were incubated for 120 minutes with 2-20 μg/ml of Hachimijogan (HZ) and Tokishakuyakusan (TS). The animals in another group were injected intravenously with 20 μg of HZ and TS 48 hours after PMS injection, and 20 minutes later decapitated. Deoxyribonucleic acid (DNA) α-nucleotidyltransferase activity and cyclic AMP accumulation in ovarian tissue were significantly increased by HZ and TS. These results suggest that HZ and TS enhance DNA α-nucleotidyltransferase activity via cyclic AMP by ovarian follicles.

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Application of Omni-ATAC to Profile Chromatin Accessibility Before and After Ovarian Tissue Cryopreservation

10.1101/2021.04.29.21256316 ◽

2021 ◽

Author(s):

Jennifer A. Shannon ◽

Aishwarya Sundaresan ◽

Orhan Bukulmez ◽

Zexu Jiao ◽

Sarah Capelouto ◽

...

Keyword(s):

Epithelial Mesenchymal Transition ◽

Ovarian Tissue ◽

Regulatory Elements ◽

Ovarian Follicles ◽

Chromatin Accessibility ◽

Ovarian Tissue Cryopreservation ◽

Post Transplantation ◽

Mesenchymal Transition ◽

Before And After ◽

Tissue Cryopreservation

AbstractOvarian tissue cryopreservation and subsequent autologous transplantation has allowed resumption of endocrine function as well as fertility in certain populations. However, graft function is short-lived due to ischemia and aberrant follicular activation post-transplantation. While many studies have focused on gene expression, we wanted to determine whether cryopreservation itself had a deleterious effect on regulatory elements that might influence transcriptional integrity and graft performance. In this study, we used Omni-ATAC to assess genome-wide chromatin accessibility in primary human follicles before and after cryopreservation. Omni-ATAC from fresh ovarian follicles identified active regulatory elements expected to be functional in oocytes and granulosa cells, and gene ontology was consistent with RNA translation/processing and DNA repair. While promoter accessibility was largely maintained in cryopreserved ovarian follicles, we observed a widespread increase in the number of accessible enhancers. Transcription factor motif analysis and gene ontology suggested that this dysregulation was focused around the epithelial-mesenchymal transition. Indeed, transcription factor binding was noted in major pathways involved in this transition: TGF-β and Wnt signaling. Overall, our work provides the first genomic analysis of active regulatory elements in matched fresh and cryopreserved ovarian follicles as they undergo the process of ovarian tissue cryopreservation. Our data suggest that the process of cryopreservation activates an epithelial-mesenchymal transition state, which may lead to graft burn-out post-transplantation. Optimizing this technique in relation to this transition may therefore be an important step towards improving graft longevity and patient outcomes in fertility preservation.Summary sentenceCryopreservation of ovarian cortical tissue results in activation of differentiation and EMT pathways in follicles, which may explain graft burnout after autotransplantation.

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Radiation-induced ovarian follicle loss occurs without overt stromal changes

Reproduction ◽

10.1530/rep-18-0089 ◽

2018 ◽

Vol 155 (6) ◽

pp. 553-562 ◽

Cited By ~ 13

Author(s):

Bruce F Kimler ◽

Shawn M Briley ◽

Brian W Johnson ◽

Austin G Armstrong ◽

Susmita Jasti ◽

...

Keyword(s):

Low Dose ◽

Ovarian Tissue ◽

Ovarian Follicle ◽

Ovarian Follicles ◽

Specific Protein ◽

Reproductive Aging ◽

Ovarian Stroma ◽

Cesium 137 ◽

Radiation Induced ◽

Total Body

Radiation damage due to total body irradiation (TBI) or targeted abdominal radiation can deplete ovarian follicles and accelerate reproductive aging. We characterized a mouse model of low-dose TBI to investigate how radiation affects the follicular and stromal compartments of the ovary. A single TBI dose of either 0.1 Gy or 1 Gy (Cesium-137 γ) was delivered to reproductively adult CD1 female mice, and sham-treated mice served as controls. Mice were euthanized either 2 weeks or 5 weeks post exposure, and ovarian tissue was harvested. To assess the ovarian reserve, we classified and counted the number of morphologically normal follicles in ovarian histologic sections for all experimental cohorts using an objective method based on immunohistochemistry for an oocyte-specific protein (MSY2). 0.1 Gy did not affect that total number of ovarian follicles, whereas 1 Gy resulted in a dramatic loss. At two weeks, there was a significant reduction in all preantral follicles, but early antral and antral follicles were still present. By five weeks, there was complete depletion of all follicle classes. We examined stromal quality using histologic stains to visualize ovarian architecture and fibrosis and by immunohistochemistry and quantitative microscopy to assess cell proliferation, cell death and vasculature. There were no differences in the ovarian stroma across cohorts with respect to these markers, indicating that this compartment is more radio-resistant relative to the germ cells. These findings have implications for reproductive health and the field of fertility preservation because the radiation doses we examined mimic scatter doses experienced in typical therapeutic regimens.

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Lower apoptosis rate in ovine preantral follicles from ovaries stored in supplemented preservation media

Zygote ◽

10.1017/s0967199414000665 ◽

2015 ◽

Vol 23 (6) ◽

pp. 943-950 ◽

Cited By ~ 4

Author(s):

R.J.S. Gonçalves ◽

A.Y.P. Cavalcante ◽

B.B. Gouveia ◽

T.L.B. Lins ◽

R.S. Barberino ◽

...

Keyword(s):

Ovarian Tissue ◽

Ovarian Follicles ◽

Terminal Deoxynucleotidyl Transferase ◽

Oocyte Diameter ◽

Minimal Essential Medium ◽

Preantral Follicles ◽

Apoptosis Rate ◽

Physiological Values

SummaryThe aim of this study was to investigate the effect of ovarian tissue transportation conditions (medium and period of time) on the morphology, apoptosis and development of ovine preantral follicles cultured in vitro. Each ovarian pair was cut into nine slices, with one fragment being fixed immediately (fresh control). The remaining fragments were placed individually in cryotubes containing conservation medium (minimal essential medium (MEM) without supplementation or MEM+ – with supplementation) and stored at 35ºC for 6 or 12 h without (non-cultured) or with subsequent culture for 5 days. Then, the fragments were processed for histological and terminal deoxynucleotidyl transferase (TdT) mediated dUTP nick-end labelling (TUNEL) examination. Preservation of ovarian slices in MEM or MEM+ (non-cultured) resulted in similar percentages of normal follicles when compared with the fresh control. Nevertheless, compared with the fresh control, a decrease in the percentage of normal follicles was observed in tissues cultured for 5 days. Only for tissues preserved in supplemented medium (MEM+) for 6 h, the percentage of TUNEL positive cells was similar between non-cultured tissues and tissues cultured for 5 days. Follicular activation and growth (follicular and oocyte diameter) were higher in cultured tissues than in fresh control or non-cultured tissues, except those from fragments preserved for 6 h in MEM and then cultured for 5 days in which no growth was observed. In conclusion, ovine ovarian tissue was successfully preserved in supplemented medium (MEM+) at a temperature close to physiological values (35°C) for up to 6 h without affecting apoptosis in the ovarian follicles and their ability to develop in vitro.

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Quantitative variations in ovarian follicles of female Sprague Dawley rats after exposure to low dose gamma radiation

Journal of Applied and Natural Science ◽

10.31018/jans.v6i1.375 ◽

2014 ◽

Vol 6 (1) ◽

pp. 57-61

Author(s):

S. Singh ◽

J. Sharma ◽

P. K. Goyal

Keyword(s):

Dose Rate ◽

Low Dose ◽

Ovarian Tissue ◽

Ovarian Follicles ◽

High Dose Rate ◽

Whole Body ◽

High Dose ◽

Sprague Dawley ◽

Irreversible Damage ◽

Sprague Dawley Rats

In the present experiment, an attempt was made to assess the genetic risk of low dose radiations in mammals. For this purpose female Sprague Dawley rats, 11-12 weeks old, were irradiated with whole body Co 60 gamma rays in three fractionated doses of 0.10 Gy (cumulative dose 0.30 Gy) given at an interval of one month at two different dose-rates (0.00368 Gy/min. and 0.0589 Gy/min.). Ovaries were studied for quantitative evaluation of follicles at 1, 4, 12, 28 and 52 weeks after last fractionated exposure. Quantitation revealed lower number of ovarian follicles in irradiated animals than in controls. The follicular number decreased with the advancement of time after last exposure (i.e. 2 months) and reached a peak level on 28 weeks. After that the recovery was evident but the number remained below 25% of total follicles even at 52 weeks autopsy interval, which indicated an irreversible damage in ovarian tissue. Primary follicles were found to be the most radiosensitive among the various types of follicles. The highest loss in these follicles was noted at 12 weeks after exposure with the high dose-rate, where only 10.61% of them were scored. Dose-rate exhibited an inverse relationship with the number of surviving follicles. At the higher dose-rate (0.0589 Gy / min.), depletion in the total follicular number was significantly higher than at the low dose-rate (0.00368 Gy/min.) used.

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