Short-term culture and hormone secretion analysis of prepubertal testicular tissue

This study aimed to investigate the ex vivo properties of prepubertal testicular biopsy samples collected for fertility preservation, with particular focus on how patient-specific factors and tissue handling may affect the hormone-secreting capacity of testicular somatic cells. Testicular biopsy samples from 59 boys aged 0.7 to 14.3 years, obtained before or during treatment for malignant or haematological diseases within a fertility preservation research program, were cultured short-term using an organotypic culture system. Culture medium was collected periodically and analysed for secretion of anti-Müllerian hormone, inhibin B, and testosterone. Hormone secretion was evaluated in relation to age at biopsy, spermatogonial density, previous chemotherapy exposure, exposure to alkylating agents, and biopsy transportation time. The tissues retained the ability to secrete hormones outside the body after transportation and, in some cases, previous chemotherapy exposure. Hormone secretion was not significantly associated with spermatogonial density, suggesting that somatic cell function may be preserved even when germ cells are depleted. Age at biopsy and exposure to alkylating agent chemotherapy were significant predictors of hormone secretion in culture. These findings indicate that short-term hormone secretion assays can be used to assess the functional capacity of somatic cell populations in prepubertal testicular tissue collected for fertility preservation, and that both developmental stage and previous treatment exposure should be considered when interpreting tissue function. Lay Summary Some boys with cancer or blood diseases may have a small piece of testicular tissue frozen before they start treatment, with the hope of preserving future fertility. In this study, testicular tissue from 59 boys aged 0.7 to 14.3 years was kept in a laboratory dish for a short time after biopsy. We measured whether the tissue could still release testosterone and other hormones that support germ cells that will make sperm in adulthood. The tissue of was able to release hormones after being transported from the operating room to the laboratory premises and, in some cases, even after previous chemotherapy. Hormone release was linked to the child’s age and to previous treatment with a type of chemotherapy known to damage fertility. It was not clearly linked to the number of germ cells in the tissue. This suggests that hormone producing testicular cells may remain active even when germ cells are reduced. These tests may help assess tissue quality before future fertility treatments.