Document Detail


Establishment of type II 5alpha-reductase over-expressing cell line as an inhibitor screening model.
MedLine Citation:
PMID:  17646096     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
Dihydrotestosterone (DHT) is the most potent male hormone that causes androgenetic alopecia. The type II 5alpha-reductase is an enzyme that catalyzes the conversion of testosterone (T) to DHT, therefore it can be expected that specific inhibitors for type II 5alpha-reductase may improve the pathophysiologic status of androgenetic alopecia. In this study, we attempted to establish the reliable and convenient screening model for type II 5alpha-reductase inhibitors. After transfection of human cDNA for type II 5alpha-reductase into HEK293 cells, the type II 5alpha-reductase over-expressing stable cells were selected by G418 treatment. RT-PCR and Western blot analyses confirmed that type II 5alpha-reductase gene was expressed in the stable cells. In in vitro enzymatic assay, 10 microg of stable cell extract completely converted 1 microCi (approximately 0.015 nmol) of T into DHT. The type II 5alpha-reductase activity was inhibited by finasteride in a dose-dependent manner, confirming the reliability of screening system. In cell culture condition, 2 x 10(5) of stable cells completely converted all the input T (approximately 0.03 nmol) into DHT by 4h incubation, demonstrating that the stable cell line can be used as a cell-based assay system. Using this system, we selected the extracts of Curcumae longae rhizoma and Mori ramulus as the potential inhibitors for type II 5alpha-reductase. These results demonstrate that the type II 5alpha-reductase over-expressing stable cell line is a convenient and reliable model for screening and evaluation of inhibitors.
Authors:
Sunhyae Jang; Young Lee; Seong-Lok Hwang; Min-Ho Lee; Su Jin Park; In Ho Lee; Sangjin Kang; Seok-Seon Roh; Young-Joon Seo; Jang-Kyu Park; Jeung-Hoon Lee; Chang Deok Kim
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2007-06-22
Journal Detail:
Title:  The Journal of steroid biochemistry and molecular biology     Volume:  107     ISSN:  0960-0760     ISO Abbreviation:  J. Steroid Biochem. Mol. Biol.     Publication Date:    2007 Nov-Dec
Date Detail:
Created Date:  2007-09-17     Completed Date:  2007-11-27     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9015483     Medline TA:  J Steroid Biochem Mol Biol     Country:  England    
Other Details:
Languages:  eng     Pagination:  245-52     Citation Subset:  IM    
Affiliation:
Department of Dermatology and Research Institute for Medical Sciences, School of Medicine, Chungnam National University, Daejeon, Republic of Korea.
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MeSH Terms
Descriptor/Qualifier:
Base Sequence
Blotting, Western
Cell Line
Cholestenone 5 alpha-Reductase / antagonists & inhibitors,  metabolism*
DNA Primers
Enzyme Inhibitors / pharmacology*
Humans
Models, Theoretical
Reverse Transcriptase Polymerase Chain Reaction
Chemical
Reg. No./Substance:
0/DNA Primers; 0/Enzyme Inhibitors; EC 1.3.1.22/Cholestenone 5 alpha-Reductase

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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