Document Detail

Development of a cryopreservation protocol for Leydig cells.
MedLine Citation:
PMID:  17596277     Owner:  NLM     Status:  MEDLINE    
BACKGROUND: In the present study, we describe a procedure to cryopreserve the postnatal members of the Leydig cell lineage, including progenitor (PLC), immature (ILC) and adult (ALC) Leydig cells from, respectively 21-, 35- and 90-day-old rats. METHODS: The cells were resuspended in a culture medium supplemented with 1% bovine serum albumin (Dulbecco's Modified Eagle's Medium [DMEM]/F12) to a final concentration of 2 x 10(6)cells/ml and the effects of varying concentrations of dimethylsulfoxide (DMSO) (5, 10, 15 or 20%) were assessed after freezing at -70 degrees C and then storing in liquid nitrogen. After 12 months of frozen storage, these cells were thawed rapidly at 37 degrees C and Trypan Blue exclusion staining and attachment to culture dishes were assessed as measures of viability. RESULTS: The trypan blue exclusion and attachment rates for Leydig cell stages were around 85% in the presence of 15% DMSO. After frozen storage, Leydig cell steroidogenic capacity in response to a range of LH doses, (0.01-100 ng/ml) was unchanged compared with freshly isolated control cells. Furthermore, the steady-state mRNA levels for Leydig cell specific transcripts were maintained. CONCLUSIONS: This study demonstrates that purified rat Leydig cells at a range of developmental stages can be frozen and that the cryopreserved cells retain normal function.
Guo-Rong Chen; Ren-Shan Ge; Han Lin; Lei Dong; Chantal M Sottas; Matthew P Hardy
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural     Date:  2007-06-27
Journal Detail:
Title:  Human reproduction (Oxford, England)     Volume:  22     ISSN:  0268-1161     ISO Abbreviation:  Hum. Reprod.     Publication Date:  2007 Aug 
Date Detail:
Created Date:  2007-07-27     Completed Date:  2007-10-12     Revised Date:  2007-12-03    
Medline Journal Info:
Nlm Unique ID:  8701199     Medline TA:  Hum Reprod     Country:  England    
Other Details:
Languages:  eng     Pagination:  2160-8     Citation Subset:  IM    
Department of Pathology of the 1st Affiliated Hospital, Wenzhou Medical College, Wenzhou, Zhejiang 325000, People's Republic of China.
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MeSH Terms
Androstane-3,17-diol / biosynthesis
Androsterone / biosynthesis
Cell Proliferation
Cell Survival
Cryopreservation / methods*
Dimethyl Sulfoxide / pharmacology
Leydig Cells / cytology*,  drug effects,  metabolism
Luteinizing Hormone / pharmacology
Rats, Sprague-Dawley
Testosterone / biosynthesis
Grant Support
Reg. No./Substance:
25126-76-5/Androstane-3,17-diol; 53-41-8/Androsterone; 58-22-0/Testosterone; 67-68-5/Dimethyl Sulfoxide; 9002-67-9/Luteinizing Hormone

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