Abstract
The disruption of androgen receptor (AR) mediated androgen signaling played very important roles in several androgen related diseases and symptoms. In order to screen the androgen modulating potency of chemicals, we developed stable AR-GreenS cell lines to access AR mediated transcriptional activation and optimized the protocol. Stable AR-GreenS cell line was stably transfected with pGL4-MMTV/Hygro, which is a firefly luciferase reporter vector bearing androgen responsive element (ARE), in 22RV1 cell line which is a human prostate cancer cells contained functional AR. AR-GreenS cell line was characterized the expression of hormone receptors. In this stable cell line, 5α-Dihydrotestosterone (DHT) was dose-dependently induced the luciferase activity and the activity was significantly increased at 1.0 x 10-10M and stated to reach to plateu 10-8M with maximum about 15 folds compared with vehicle control. This DHT induced luciferase activity was inhibited by the treatment of AR antagonist, bicalutamide. AR-GreenS cells have maintained their growth rate, morphology and responsiveness to DHT until now 85 passages cultured for over 13 months. The inter variation of assay was relatively small with about 5.1± 1.3 mean value of CV (the coefficient of variation). Using AR-GreenS cells, we established the 3 days test protocol and optimized the testing condition for AR transcriptional activation assay. We validated the stable cell line using 20 compounds among 78 substances which are recommended substances for validation of in vitro androgen receptor transcriptional activation assays by ICCVAM.
*This research was support by a grant (12162KFDA737) from Korea Food & Drug Administration in 2012
