Isolation of germ cells from human testicular tissue for low temperature storage and autotransplantation.
OBJECTIVE
To develop a new protocol for conserving fertile potential in men undergoing sterilizing chemotherapy by low temperature banking of germ cells which can be returned to the patient's testes after thawing.


METHODS
Isolation of human and murine germ cells for comparing cellular viability after cooling to liquid nitrogen temperatures by the use of different cryoprotective agents and for infusion into the testis.


METHODS
Laboratory research environment.


METHODS
Men undergoing routine surgery in a urology department.


METHODS
Testicular biopsy.


METHODS
Cellular viability and infusion of seminiferous tubules.


RESULTS
After isolation using a two-step enzymatic disaggregation protocol, 66% to 87% of germ cells from human and murine specimens, respectively, were still viable. Cell survival was similar in four commonly used cryoprotective agents after cooling to liquid nitrogen temperatures. Seminiferous tubules infused by back flow with dye solution via the rete testis were filled with an efficiency of 55%.


CONCLUSIONS
Judging from the high viability of unfractionated germ cells, it is feasible to isolate germ cells from testicular biopsies for low temperature banking with the aim of attempting to restore fertility after iatrogenic sterilization.
Brook
P F
PF
Centre for Reproduction, Growth, and Development, Leeds General Infirmary, University of Leeds, United Kingdom.
Radford
J A
JA
Shalet
S M
SM
Joyce
A D
AD
Gosden
R G
RG
D000818:Animals; D002469:Cell Separation; D017690:Cell Transplantation; D015925:Cryopreservation; D003451:Cryoprotective Agents; D006801:Humans; D007248:Infertility, Male / Q000139:chemically induced / Q000601:surgery; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D012671:Seminiferous Tubules; D013094:Spermatozoa / Q000166:cytology*; D013737:Testis / Q000166:cytology*
D003451:Cryoprotective Agents