Vitrification with microinjection of single seminiferous tubules: an efficient cryopreservation approach for limited testicular tissue

Sha Han; Liangyu Zhao; Chao Yang; Junwei Xu; Chencheng Yao; Chuan Huang; Hongbin Zhang; Zhiyong Ji; Jiaqiang Luo; Yingying Guo; Yan Hong; Feng Liu; Xinli Zhou; Zhi Zhou; Yonghua Huang; Zheng Li

doi:10.1016/j.rbmo.2021.06.026

Vitrification with microinjection of single seminiferous tubules: an efficient cryopreservation approach for limited testicular tissue

Reprod Biomed Online. 2021 Oct;43(4):687-699. doi: 10.1016/j.rbmo.2021.06.026. Epub 2021 Jul 3.

Authors

Sha Han¹, Liangyu Zhao², Chao Yang³, Junwei Xu³, Chencheng Yao³, Chuan Huang³, Hongbin Zhang⁴, Zhiyong Ji³, Jiaqiang Luo³, Yingying Guo⁵, Yan Hong³, Feng Liu³, Xinli Zhou⁵, Zhi Zhou⁶, Yonghua Huang⁷, Zheng Li⁸

Affiliations

¹ Department of Andrology, the Center for Men's Health, Urologic Medical Center, Shanghai Key Laboratory of Reproductive Medicine, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, China; Institute of Refrigeration and Cryogenics, Shanghai Jiao Tong University, Shanghai 200240, China.
² Department of Andrology, the Center for Men's Health, Urologic Medical Center, Shanghai Key Laboratory of Reproductive Medicine, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, China; Department of Urology, The Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, Guangdong Province 519000, China; Guangdong Provincial Key Laboratory of Biomedical Imaging, The Fifth Affiliated Hospital, Sun Yat-sen University, Zhuhai, Guangdong Province 519000, China.
³ Department of Andrology, the Center for Men's Health, Urologic Medical Center, Shanghai Key Laboratory of Reproductive Medicine, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, China.
⁴ Institute of Refrigeration and Cryogenics, Shanghai Jiao Tong University, Shanghai 200240, China.
⁵ University of Shanghai for Science and Technology, Shanghai 200000, China.
⁶ School of Life Science and Technology, Shanghai Tech University Shanghai 200120, China. Electronic address: [email protected].
⁷ Institute of Refrigeration and Cryogenics, Shanghai Jiao Tong University, Shanghai 200240, China. Electronic address: [email protected].
⁸ Department of Andrology, the Center for Men's Health, Urologic Medical Center, Shanghai Key Laboratory of Reproductive Medicine, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, China. Electronic address: [email protected].

PMID: 34556414
DOI: 10.1016/j.rbmo.2021.06.026

Abstract

Research question: Is vitrification with microinjection of single seminiferous tubules an efficient cryopreservation approach for limited testicular tissue?

Design: Testicular tissue from 10 patients with normal spermatogenesis were assigned to a fresh control group or one of the following cryopreservation procedures: uncontrolled slow freezing (USF) using either 1.5 or 2.1 M DMSO combined with sucrose and vitrification with or without single seminiferous tubules microinjection.

Results: Single seminiferous tubules microinjected with cryoprotective agents (CPA) enhanced the penetration of CPA compared with CPA-treated testicular tissue fragments. Microinjection of seminiferous tubules (VLP) maintained tubule structural integrity and germ cell numbers, and reduced spermatogonial apoptosis after cryopreservation compared with vitrification without microinjection (apoptosis rate: VLP versus vitrification without microinjection, P = 0.047; VLP versus USF, P= 0.049). Freezing of single seminiferous tubules using 0.25-ml straws and traditional sperm freezing methods protected sperm retrieval and recovery rates, and the progressive motility index.

Conclusions: Vitrification of single seminiferous tubule with microinjection of low CPA concentration is an effective approach to testicular cryopreservation.

Keywords: Cryopreservation; Microinjection; Seminiferous tubule; Slow cooling; Vitrification.

Publication types

Controlled Clinical Trial

MeSH terms

Adult Germline Stem Cells*
Cryopreservation / methods*
Cryoprotective Agents / administration & dosage*
Humans
Male
Microinjections
Seminiferous Tubules*
Spermatogonia*
Vitrification

Substances

Cryoprotective Agents