Spermatogonial survival after cryopreservation and short-term orthotopic immature human cryptorchid testicular tissue grafting to immunodeficient mice

Abstract

Through our orthotopic xenografting model, we have demonstrated the survival and proliferative activity of spermatogonia and Sertoli cells in cryopreserved immature human cryptorchid tissue. Testicular tissue banking may thus prove to be a promising technique for the preservation of fertility in prepubertal boys undergoing oncological treatments. As the stem cell niche is maintained, the cryopreserved tissue can potentially be used for future autotransplantation. In addition, whole tissue freezing does not exclude alternative clinical uses, including isolated cell transplantation after dissociation, selection and enrichment. However, as this work was done on cryptorchid tissue, studies on normal immature testicular tissue, involving longer grafting periods, are needed to demonstrate a differentiation capacity before clinical implementation. Ethical and safety issues should also be addressed.

Cryopreservation of Testicular Tissue

“…Shinohara et al (2002) reported the birth of mouse offspring from sperm retrieved from cryopreserved pre-pubertal testis tissue with DMSO after transplantation under tunica albuginea of the recipient testes (Shinohara et al, 2002). Similar results were obtained using primate testis tissue, where 1.4M (but not 0.7M) DMSO was able to protect some of the developmental potential of grafts from rhesus monkeys but the 0.7M DMSO protocol was successful for cryopreservation of human testis tissue (Wyns et al, 2007) at one age/developmental stage but not others (Wyns et al, 2008;Keros et al, 2005Keros et al, , 2007. Somewhat different from reports in other species, and after an extensive study of several strategies for cryopreservation of immature testis tissue, we concluded that glycerol was a better cryoprotectant for pig tissues (Abrishami et al, 2010a).…”

“…Mouse testes have considerably less connective tissue content than most other species; therefore, tissue fragment size is especially a concern for testis tissues from species with higher interstitial tissue density. For cryopreservation of (cryptorchid) testes from prepubertal boys, fragments sizes of 2-9 mm 3 were used successfully (Wyns et al, 2007). We also reported that immature porcine testis tissues undergoing the same cryopreservation treatments were not affected by the original size of the testis tissue fragment (5, 15, 20, or 30 mg) (Abrishami et al, 2010a).…”

“…Transmitted electron microscopy, although not widely used, can be invaluable in the examination of subcellular components most likely to be affected by testis tissue cryopreservation, including cytoplasm integrity, nuclear membrane, and various organelles (Keros et al, 2007). Other valuable morphological analyses may include assessment of cell-specific changes, for example, using double-staining of proliferation markers (e.g., Ki67) and MAGE-AH, vimentin, or CD34 for identification of spermatogonia, Sertoli cells, or peritubular cells, respectively (Keros et al, 2007;Wyns, et al, 2007).…”

“…Post-thawing in vitro organotypic culture of the cryopreserved testis tissue has allowed assessment of its survival in the shortterm (Curaba et al 2011;Keros et al, 2007) and measurement of its hormone release into culture media (Gouk et al, 2011). Perhaps more robust examination is provided by grafting, where the survival and developmental competence (both in terms of germ cell differentiation and androgen release) of the cryopreserved tissue in vivo as grafts allows a longer-term functional assessment (Abrishami et al, 2010a;Jahnukainen et al, 2007;Wyns et al, 2007).…”

New Approaches of Ovarian Tissue Cryopreservation from Domestic Animal Species

“…It is the only available solution for pre-pubertal boys who must receive a gonadotoxic treatment (eg. cancer therapy; (Keros et al, 2005;Keros et al, 2007;Wyns et al, 2007;Wyns et al, 2008;Wyns et al, 2010).…”