"Testicular cell freezing for fertility preservation in pre-pubertal boys undergoing gonadotoxic therapy"

Student thesis: Doctoral Thesis


Fertility preservation prior to gonadotoxic therapies is an important emerging field. This technology seeks to safeguard future chances of fatherhood in prepubertal patients facing fertility compromising regimens (e.g. chemo- or radiotherapy) by cryopreserving spermatogonial stem cell (SSC)- containing testicular tissue (TT). This procedure is currently offered as an innovative experimental method. This thesis started with an overview of the efficiency of existing TT and testicular cell suspension (TCS) cryopreservation protocols. Efficient cryopreservation protocols for human TT include DMSO, human serum albumin and sucrose, and these are currently applied in the clinic. For TCSs, cryopreservation and cell functionality remained suboptimal. We optimized the cryopreservation protocol for TCSs with immature mouse TCSs aiming for a better cell recovery and fertility restoration after SSC transplantation. The use of an insulate-controlled freezing device and DMEM supplemented with 1.5M DMSO, 10% FCS and 60µM of an anti-apoptotic factor (Z-VAD(Oe)-FMK) as cryomedium resulted in 49% survival of SSCs. For human primary testicular cells, upon freezing, the highest recovery of viable cells was achieved by freezing cells using DMEM supplemented with 1M DMSO, 20% HSA, 30 µM Z-VAD(Oe)-FMK and 200 mM of trehalose in an insulate-controlled slow freezing device. Controlled-rate thawing improved the viability even more. Finally, we compared the ability to restore fertility by SSCT after cryopreserving TCSs with that after cryopreserving TT. Our results highlighted the superiority of TT cryopreservation, as currently applied, to preserve functional SSCs.
Date of Award27 May 2019
Original languageEnglish
Awarding Institution
  • Vrije Universiteit Brussel
  • University of Amsterdam
SupervisorEllen Goossens (Promotor), M.M. van Pelt Ans (Promotor), Olle Söder (Jury), Mariëtte Goddijn (Jury), Willem Ombelet (Jury), Eline Menu (Jury), Herman Tournaye (Jury) & Hilde Van De Velde (Jury)


  • Fertility
  • Gonadotoxic therapy

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