Cryobiology in human assisted reproductive technology. Would Hippocrates approve?

Recent advances in human cryobiology have been substantially greater than the first slow step from freezing spermatozoa in animals in Italy, published in 1776 to observing motility in frozen-thawed human sperm in 1938(1). Reports on cryopreservation of rabbit oocytes (1947)(1) and births from fertilised frozen-thawed mice oocytes in 1977(1) were soon followed by the first human pregnancy (1983)(1) and birth (1984)1 following transfer of frozen-thawed embryos after in-vitro fertilisation (IVF). Whereas cryopreservation of human sperm and embryos in tertiary level fertility centres is now commonplace, the full clinical, scientific and sociological consequences of progress in this rapidly moving field are to be determined. These include pregnancy with frozen-thawed human mature, oocytes after conventional IVF (1986)4, intracytoplasmic sperm injection (ICSI)(5) (1996), pregnancies following use of frozen-thawed mature (1995)(5,6) and immature oocytes (1999)(7), ovarian tissue banking (8) and possible autografting (1999)(9) as well as repeated freeze-thawing of male gametes and of embryos (10,11). Cryopreservation of female and male gametes instead of embryos offer solutions of obvious religious, ethical, legal and clinical problems. In addition, there may be benefits in reducing the cost of infertility treatment, improving the safety of fertility treatment with respect to ovarian hyperstimulation syndrome and repeated treatment with controlled ovarian hyperstimulation, prevention of diseases such as sexually transmitted diseases and hereditary disorders and preventing infertility by possible long-term storage of gametes, gonadal tissue and even embryos. The benefits of cryopreservation of sperm, oocytes and embryos in the management of subfertile couples, many being self-evident to some, bear emphasis. Cryopreservation of sperm offers substantial organisational, cost and social advantages in IVF/ICSI treatment, in that it is no longer necessary for both partners to be present at the time of oocyte retrieval, or to have the sperm retrieval done simultaneously, as frozen-thawed sperm (ejaculatory, epididymal or testicular) can be used. This strategy permits men in the latter two categories to be able to support their partners at the time of oocyte retrieval, with the knowledge that their sperm surgically obtained some time previously, is available. It is now clear that, in men with obstructive azoospermia, the use of fresh or frozen-thawed sperm will yield equivalent fertilisation rates following ICSI. In men with non-obstructive azoospermia, with a 60% chance only of obtaining sperm from the testicular aspiration or biopsy, the option could be cryopreservation of the sperm harvested first and later controlled ovarian hyperstimulation of the female partner, to use thawed sperm which will lead to equivalent fertilisation rates using fresh sperm. Thus, one may avoid cost of treatment of the female in those couples who do not wish to use donor sperm as a back-up in the 40% of men from whom sperm is not obtained. Important consequences of cryopreservation of gametes and gonadal tissue are likely to be in the area of prevention of hereditary and familial diseases, as cryopreservation of oocytes, sperm, embryos and blastocysts is exploited fully in pre-implantation genetic diagnosis (PGD) strategies12. Embryo biopsy now permits screening to identify normal embryos from couples who are carriers of known single gene defects and hereditary disorders and the list of these conditions is expanding rapidly. PGD is feasible on frozen-thawed blastomeres even if cells have lysed after thawing, providing information relevant for surviving blastomeres or blastocysts. But what of the gene probes which will soon deluge us on the completion of the Human Genome Project? Can we anticipate benefits and consider proposing that couples with familial disorders, whether degenerative e.g. Type 2 Diabetes, or malignant conditions such as cancer of the ovary, breast and colon? Should we cryopreserve oocytes/sperm/embryos for the purposes of PGD once the markers are available? Cryobiology indeed provides hope now for women and men with neoplastic diseases, who are about to receive oncotherapy for malignancies which inevitably will render them sterile. Men may now freeze epididymal, testicular as well as ejaculatory sperm as ICSI has revolutionalised the treatment of male infertility. It might be likely that testicular tissue from prepubertal boys can be cryopreserved with a reasonable expectation that techniques will soon be developed to effect maturation of spermatogonia in-vivo or in-vitro13. The greatest advance is likely to be for women suffering from reproductive cancer, who may now consider mature and immature oocytes being frozen or vitrified with a reasonable chance of fertilisation by ICSI later, as well as the cryopreservation and storage of ovarian cortex tissue biopsies. Work is proceeding still to refine techniques of in-vitro maturation of frozen-thawed immature oocytes, and the frozen-thawed ovarian cortex tissue slices. The potential benefits will not only be to female fertility for the latter conditions but endocrine disorders as well as by autotransplantation (1999)9. Currently, ovarian tissue banking8 is being considered by women undergoing procedures or treatment which could destroy ovarian function with quite realistic but cautious expectations of preserving ovarian function, but tomorrow women may consider banking ovarian tissue as insurance against childlessness because of the risk of disorders in the reproductive tract (endometriosis, simple recurrent ovarian cysts) and even advancing years. For those who have conceived with surplus oocytes cryopreserved, anonymous oocyte donation is a possibility for the solution of ethical and legal problems. All over Europe, the age of women having their first child is dramatically increasing now being in their late twenties, with likely significant implications in the need to fertility treatment in the Millennium. Society has always been excited but understandably cautious about the prospect of whole body cryopreservation. Hippocrates would have argued that Society could separate medicine and its advances from religious views, dogma and prejudice and, on the present evidence, would probably have looked upon human cryobiology favourably. Human cryobiology is here to stay and society as well as the profession is addressing its relevance. There are clear signs that this technology can and will alleviate suffering by preventing genetic and familial diseases, infections and infertility as well as lowering the cost and social consequences of the treatment. For these reasons, further research in this field should be welcomed and supported.