BACKGROUND Female cancer patients are offered 'banking' of gametes before starting fertility-threatening cancer therapy. Transplants of fresh and frozen ovarian tissue between healthy fertile and infertile women have demonstrated the utility of the tissue banked for restoration of endocrine and fertility function. Additional methods, like follicle culture and isolated follicle transplantation, are in development. METHODS Specialist reproductive medicine scientists and clinicians with complementary expertise in ovarian tissue culture and transplantation presented relevant published literature in their field of expertise and also unpublished promising data for discussion. As the major aims were to identify the current gaps prohibiting advancement, to share technical experience and to orient new research, contributors were allowed to provide their opinioned expert views on future research. RESULTS Normal healthy children have been born in cancer survivors after orthotopic transplantation of their cryopreserved ovarian tissue. Longevity of the graft might be optimized by using new vitrification techniques and by promoting rapid revascularization of the graft. For the in vitro culture of follicles, a successive battery of culture methods including the use of defined media, growth factors and three-dimensional extracellular matrix support might overcome growth arrest of the follicles. Molecular methods and immunoassay can evaluate stage of maturation and guide adequate differentiation. Large animals, including non-human primates, are essential working models. CONCLUSIONS Experiments on ovarian tissue from non-human primate models and from consenting fertile and infertile patients benefit from a multidisciplinary approach. The new discipline of oncofertility requires professionalization, multidisciplinarity and mobilization of funding for basic and translational research.
The objective of this study was to investigate the effect of follicle stimulating hormone (FSH) priming on the in-vitro maturation (IVM) of human oocytes from healthy ovaries using a chemically defined culture system. Seventeen patients donating oocytes for research received a truncated course of 600 IU FSH over 5 days and a further control group of nine patients received no FSH treatment. Mid-follicular phase cumulus-enclosed oocytes (n = 160) were aspirated from follicles < or =4 mm diameter under transvaginal ultrasound guidance and were cultured for 48 h in microdrops of medium containing 10 mIU/ml FSH and 100 mIU/ ml human chorionic gonadotrophin (HCG). The results demonstrated that human oocytes will efficiently undergo IVM under serum-free conditions. After mild FSH stimulation, a greater number of cumulus-enclosed oocytes was collected, and following culture, a lower rate of degeneration was observed. Significantly more oocytes completed nuclear maturation to metaphase II following FSH stimulation (71.1 versus 43.5%). In conclusion, a truncated course of FSH stimulation in vivo improved the oocyte maturation rate in vitro, giving a mean of 4.8+/-0.7 metaphase II oocytes per patient compared with only 2.1+/-0.7 from control patients, thus yielding more mature oocytes for future IVF treatment.
A culture system has been designed in which enzymatically isolated oocyte-granulosa cell complexes from fresh and frozen-thawed ovine ovarian tissue can be grown to antral size in vitro. Oocyte-granulosa complexes ranging from 100 to 240 microns in diameter were dissected from stromal tissue and grown individually in serum-free medium for 30 days. Complexes < 190 microns generally excluded their oocytes or lost three-dimensional structure early in the culture period. In contrast, complexes isolated from fresh or frozen-thawed tissue and measuring 190-240 microns on the day of isolation formed antral cavities in 25 +/- 9% and 18 +/- 6% (mean +/- SEM) of cases, respectively. The effect of gonadotrophin supplementation to the culture medium was tested on frozen-thawed oocyte-granulosa cell complexes only. In cultures supplemented with both FSH and LH or FSH alone, there was no significant difference in the number of oocyte-granulosa cell complexes that formed antral cavities (18 +/- 7%). However, antrum formation was significantly less frequent in cultures lacking gonadotrophin stimulation (7 +/- 4%). All oocyte-granulosa cell complexes maintained a three-dimensional structure throughout culture and developed a functional P450 aromatase enzyme complex, as revealed by the induction of oestradiol production during 8 days of culture after antrum formation in serum-free medium containing testosterone. Oocytes recovered after 30 days of culture were viable and had increased in diameter from 78 +/- 2 microns on the day of isolation, to 131 +/- 3 microns at the end of culture. These results show that oocyte-granulosa cell complexes isolated from cryopreserved ovarian tissue can be grown to antral size in vitro with similar efficiency to those isolated from fresh tissue.
The Foot and Mouth Disease (FMD) epidemic of 2001 clearly illustrated the fragility of the UK's farm animal genetic resources. In particular, millions of sheep were killed by the disease and by the ‘stamping out’ policy chosen for disease control. Loss of genetic resources was not evenly spread throughout the UK, nor throughout the many different sheep breeds that are native to the UK and for which the UK has a formal responsibility for protection to the United Nations. In fact, the FMD epidemic demonstrated for the first time that sheep breeds comprising large numbers of individuals which are commercially farmed, can nevertheless be at considerable risk of extinction. The breeds most affected were those restricted to geographical regions of the UK into which the FMD spread. These regionally important breeds are adapted to their particular regional environments, represent an important living heritage for the UK and are a key component in sustaining the rural economies of sheep farming communities.The events of 2001 provided clear proof that there are two components of the UK's farm animal genetic resources demanding protection. One component is already recognised as a priority and is composed of the numerically rare breeds of all domesticated species: these are already under the protection of the Rare Breeds Survival Trust (RBST). The second component has not previously been recognised as a priority for protection. The FMD crisis proved that sheep breeds could exist as large numbers of individuals, but nevertheless face extinction due to their regional location. Urgent attention must be focussed on our Heritage Breeds of sheep. The UK has one of the greatest number of native sheep breeds of any country in the world. The Heritage Breeds provide potentially valuable genetic resources for environmental, low-input farming systems.Heritage GeneBank was founded during the FMD epidemic specifically to protect sheep breeds at threat of extinction from the disease. A group of academic research scientists established a genetic salvage programme: collecting semen and embryos for protection in a gene bank. Germplasm from seven breeds is in long-term storage. Following the crisis, the scientists involved in the gene bank made a commitment to continue their conservation work in recognition that the Heritage Breeds of sheep in the UK continue to require protection.This paper describes: (1) the work of Heritage GeneBank (HGB); (2) the threefold mission of The Sheep Trust, the new national charity that evolved from HGB (http://www.thesheeptrust.org); and (3) the ongoing urgent need for conservation of the UK's Heritage Breeds of sheep threatened by genetic erosion.
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