Cryosurvival and spermatogenesis after allografting prepubertal mouse tissue: comparison of two cryopreservation protocols.

Although childhood cancer treatments are yielding higher survival rates, sterility remains one of the major side effects. For prepubertal boys there are currently no options to preserve fertility. Testicular tissue banking together with subsequent grafting may become a possible strategy in the future. In the present study, we compared two cryopreservation protocols using prepubertal murine testicular tissue. Fresh and cryopreserved testicular tissue was grafted subcutaneously on the back of immune-deficient mice for at least 3 months. Prepubertal murine tissue recovered well after cryopreservation with both ethylene glycol (EG) and dimethylsulfoxide (DMSO). While in fresh murine allografts, spermatozoa were observed in 23% of the tubules; in both the DMSO and the EG groups, 32% of the seminiferous tubules contained spermatozoa. However, with DMSO the structure of the seminiferous tubules was better preserved.

Evaluation of in vivo conception after testicular stem cell transplantation in a mouse model shows altered post-implantation development.

BACKGROUND: Apart from research applications, testicular stem cell transplantation (TSCT) may one day also have valuable clinical applications. Therefore, it is important to investigate whether this technique is a safe method to have progeny. This controlled study aims at evaluating the fetuses and the live born offspring obtained after TSCT in male mice. METHODS: Male mice were mated with wild-type (WT) females after TSCT to produce offspring. First, fetuses were evaluated on the 17th gestational day. The length, weight and morphological age were compared to those of control mouse fetuses. The live born offspring were then investigated for their reproductive potential over three generations. RESULTS: The litter sizes after TSCT were decreased compared to controls. Fetuses showed developmental retardation of a quarter of a day, but no major external abnormalities were observed. The live born pups were able to produce normal litter sizes, at least until the third generation. CONCLUSIONS: Transplanted animals are able to reproduce naturally. Although litter sizes are lower and development is retarded, no major morphological or procreative abnormalities were observed.

Spermatogonial survival after grafting human testicular tissue to immunodeficient mice.

BACKGROUND: The xenografting of pre-pubertal human testicular tissue to an immunodeficient mouse is a theoretical strategy for restoring fertility in childhood cancer patients, while circumventing the risk of malignant recurrence. This study aimed at comparing the grafting of pre-pubertal and adult murine testicular tissue, as well as that of human adult testicular tissue, to two immunodeficient recipients, i.e. Swiss Nude mice and SCID-NOD mice. MATERIALS AND METHODS: In this study, we evaluated the survival of pre-pubertal and adult murine testicular tissues, and that of adult human testicular tissue after subcutaneous grafting to immunodeficient mice. RESULTS: After allografting pre-pubertal testicular tissue pieces, meiotic cells were observed in 69.1% of the grafts, while complete spermatogenesis was observed in 30.9%. All grafts of adult murine testicular tissue and 59.5% of the adult human testicular grafts showed sclerosis. However, in 21.6% of the adult human testicular grafts, spermatogonia were still observed, with increasing sclerosis in time. No significant differences were observed between the two mouse models under evaluation. CONCLUSION: After xenografting human adult testicular tissue to a recipient mouse, spermatogonia were maintained over a period of >195 days. However, in order to prove xenografting as a method for external germ line storage, the transplants should have a more immature developmental stage. Moreover, not only the developmental status of the tissue at the time-point of grafting, but also the structural organisation of the seminiferous epithelium, might influence the development of the testicular tissue.

Preserving the reproductive potential of men and boys with cancer: current concepts and future prospects.

The introduction of ICSI has totally changed the reproductive prospects for boys and men who are treated for cancer. With post-pubertal boys and adult men, semen cryopreservation should be offered to every patient undergoing a cancer treatment since preservation of fertility cannot be guaranteed for an individual patient and treatment may shift to a more sterilizing regimen. In the ICSI era, all semen samples, even those containing only a few motile sperm, should be accepted for cryopreservation. Patients who are azoospermic at the time cancer is diagnosed may be offered testicular sperm extraction and cryopreservation of testicular tissue. With pre-pubertal boys, no prevention of sterility by sperm banking is possible since no active spermatogenesis is present. However, in the next decade, prevention of sterility in childhood cancer survivors will become a major challenge for reproductive medicine. In theory, testicular stem cell banking is the only way of preserving the future fertility of boys undergoing a sterilizing chemotherapy. In animal models, testicular stem cell transplantation has proved to be effective; however, it remains to be shown that this technique is clinically efficient as well, especially when frozen-thawed cells are to be transplanted. Malignancy recurrence prevention is an important prerequisite for any clinical application of testicular stem cell transplantation. Although still at the experimental stage, cryobanking of testicular tissue from pre-pubertal boys may now be considered an acceptable strategy.