טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentAvraham Masiah
SubjectCryopreservation of Testicular Tissue to Maintain the
Fertility Potential
DepartmentDepartment of Medicine
Supervisors Mr. Lightman Abraham
Professor Emeritus Itskovitz Joseph


Abstract

In light of the long-term damage to male gonads by various chemotherapy treatments, and the need to preserve their fertility potential, we have conducted this study with the objective of finding the optimal procedure for cryopreservation of testicular tissue.  Two cryogenic protocols were examined. The first consisted of slow-rate freezing, concomitant with “seeding”, until reaching temperature of 196÷C. The second protocol consisted of fast-rate freezing. Additional variables which were examined included 3 cryoprotective agents: Ethylene Glycol, Dimethylsulfoxide, and Glycerol. For evaluation and comparison between these protocols we assessed several morphological indices of testicular tissue viability: 1. Tubular structure of the testicular tissue following freezing and thawing, 2. Survival rates of germinal cell line and Sertoli cells, 3. Survival of transplanted tissue following the cryopreservation procedure, 4. Viability staining of frozen/thawed testicular cells.

Testicular tissue was obtained from mature rats and cryopreserved according to the aforementioned protocols. Histological examination enabled semi-quantitative morphological assessment of the seminiferous tubules, which retained their normal structure. Differences were observed in the degree of tissue damage when comparing the different cryopreservation protocols. The structural assessment of the seminiferous tubules, germ cells survival rate, and viability staining, showed that slow freezing is superior to fast freezing for testicular cryopreservation. Of the cryoprotective agents examined, Ethylene glycol demonstrated the highest protective capability while Glycerol showed the lowest cryoprotective effect. Transplantation of cryopreserved testicular tissue to a mature and immature mouse recipient showed the best results when using the slow freezing protocol with Ethylene glycol as cryoprotectant.