睪丸玻璃化冷凍保存

癌症兒童化療前將未成熟睪丸組織玻璃化冷凍保存，

解凍後再植入於裸鼠睪丸，睪丸組織之精原幹細胞可進一步成熟成精子

http://humrep.oxfordjournals.org/content/28/3/578.abstract

Vitrification preserves proliferation capacity in human spermatogonia

1. Jonathan Poels1,2, 
2. Anne Van Langendonckt1,2, 
3. Marie-Christine Many3,
4. François-Xavier Wese4 and 
5. Christine Wyns1,2,*

+Author Affiliations

1. ¹Gynecology Unit, Medical School, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, Avenue Mounier, 52, 1200 Brussels, Belgium
2. ²Department of Gynecology-Andrology, Cliniques Universitaires Saint-Luc, Avenue Hippocrate, 10, 1200 Brussels, Belgium
3. ³Experimental Morphology Unit, Medical School, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, 1200 Brussels, Belgium
4. ⁴Urology Unit, Medical School, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, 1200 Brussels, Belgium

1. ↵*Correspondence address. Tel: +32-2-764-95-01; Fax: +32-2-764-95-07; E-mail: christine.wyns@uclouvain.be

• Received October 25, 2012.
• Revision received November 28, 2012.
• Accepted December 10, 2012.

Abstract

STUDY QUESTION Does vitrification of human immature testicular tissue (ITT) have potential benefits for future fertility preservation? Does vitrification of human ITT have potential benefits in an in vivo murine xenotransplantation model?

SUMMARY ANSWER Vitrification is able to maintain proliferation capacity in spermatogonial cells after 6 months of xenografting.

WHAT IS KNOWN ALREADY Controlled slow-freezing is the procedure currently applied for ITT cryobanking in clinical practice. Vitrification has been proposed as a promising technique for long-term storage of ITT, with a view to preserving spermatogonial stem cells (SSCs) for future fertility restoration in young boys suffering from cancer. After vitrification of ITT, in vitro survival of SSCs was demonstrated, but their functionality was not evaluated.

STUDY DESIGN, SIZE, DURATION Ten ITT pieces issuing from 10 patients aged 2–12 years were used. Fragments of fresh tissue (serving as controls) and fresh, frozen-thawed and vitrified-warmed testicular pieces xenografted to the scrotum of nude mice for 6 months were compared.

MATERIALS, SETTING, METHODS Upon graft removal, histological and immunohistochemical analyses were performed to evaluate spermatogonia (SG) (MAGE-A4), intratubular proliferation (Ki67), proliferating SG and Leydig cells (3β-HSD). The entire piece of grafted tissue was assessed in each case.

MAIN RESULTS AND THE ROLE OF CHANCE Seminiferous tubules showed good integrity after cryopreservation and xenografting for 6 months in all three groups. Survival of SG and their ability to proliferate was observed by immunohistochemistry in all grafted groups. SG were able to initiate spermatogenesis, but blockage at the pachytene stage was observed. The recovery rate of SG was 3.4 ± 3.8, 4.1 ± 7.3 and 7.3 ± 6.3%, respectively, for fresh, slow-frozen and vitrified-warmed tissue after 6 months of xenografting.

LIMITATIONS, REASONS FOR CAUTION The study is limited by the low availability of ITT samples of human origin. The mouse xenotransplantation model needs to be refined to study human spermatogenesis.

WIDER IMPLICATIONS OF THE FINDINGS The findings of the present study have potential implications for cryobanking of ITT and fertility preservation. Spermatogonial loss recorded after fresh ITT transplantation indicates that the avascular grafting technique needs to be optimized. There are so far no convincing data justifying modification of current clinical practice for ITT storage with slow-freezing, but this study demonstrates that it is worth pursuing optimization of ITT vitrification as an alternative for preservation of SSCs.