O-12: Clinical Grade Isolation and Cryopreservation of Human Testicular Cells

Background: More than a decade work in preservation of male fertility by spermatogonial stem cell (SSC) transplantation in animal models as well as new discoveries about the nature of human SSCs has prompted an investigation into the possibility of an effective clinicalgrade procedure for isolation and cryopreservation of human testicular cells. Materials and Methods: Clinical-grade reagents, validated equipment, and protocols consistent with cGTP/ cGMP standards were used in developing a procedure suitable for the safe and effective isolation and cryopreservation of human testicular cells. These procedures were designed to be compliant with the relevant FDA regulations. Flow Cytometry in conjunction with specific markers for germ cells (VASA), SSCs (SSEA-4) and testicular somatic cells (LHR) were used to determine the cellular composition before and after cryopreservation. Results: The procedure proved to effectively isolate and cryopreserve testicular cells from hormone-treated adult patients undergoing sexual reassignment. Our results clearly showed that important cell populations are not only preserved but enriched by the cryopreservation process. Sterility tests show that the cryopreserved cells and the thawed cells processed by our protocols under cGTP and some cGMP conditions were free from any microbial contamination. Conclusion: Cryopreservation of testicular cells isolated from fresh tissue is favorable to cryopreservation of testicular tissue because it accommodates cell analysis, quality control, and sterility testing. Further studies are needed to determine whether these findings in adult testicular tissue from sexual reassignment patients can be generalized to young male patients prior cancer therapy.