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From: Kuch
Date: 5/14/2002
Time: 1:38:06 PM
Remote Name: 66.88.192.59
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Abstract TITLE: Regulation of androgen activity by receptor gene inhibition through specific antisense oligonucleotides in human sebocytes AUTHORS (ALL): Fimmel, Sabine 1; Saborowski, Antje 1; Beutler, Claudia 1; Zouboulis, Christos C.1 INSTITUTIONS (ALL): 1. Department of Dermatology, University Medical Center Benjamin Franklin, The Free University of Berlin, Berlin, , DE;
ABSTRACT BODY: Abstract Body: Target cell responsiveness to hormonal signals are product of both cellular concentrations of the ligand and the corresponding receptor. The effects of androgens in the skin are influenced by the number of nuclear androgen receptor (AR) molecules available in skin cells. Androgens are the best-known stimulators of sebaceous gland activity, enhancing lipogenesis, proliferation, and terminal differentiation of human sebocytes in vitro and in vivo. In this study we applied antisense technology, which has already proven to be useful both as an experimental tool in functional genomics and as source of novel therapeutics. Here we describe an approach to inactivate the AR mRNA in SZ95 sebocytes by thioat- and ribosyl-antisense oligonucleotides. To transfer oligonucleotides into human SZ95 sebocytes we developed a cationic liposome- mediated transfection system with DOTAP. The transfection efficiency, proved by detection of the cellular uptake of fluorescein-labelled random oligonucleotides with fluorescence microscopy, increased within the nucleus of the sebocytes. The expression of AR on mRNA and protein levels was investigated by quantitative PCR (TaqMan) and Western blotting. The biological effect of androgens (5-DHT and testosterone) was evaluated by a proliferation assay. The most successful transient control of AR expression was detected with 1.0 µM ribosyl-antisense and revealed a 87% inhibition of AR expression compared to native cells. After a recovery time longer than 24 h the AR protein content reached again the level of untreated cells. The functional interaction between the diminished number of AR molecules and its ligands was demonstrated by an inhibition of proliferation of AR antisense-transfected SZ95 sebocytes after stimulation with androgens (1 nmol). The biological gene regulation of the AR with antisense oligonucleotides in vitro is the first step to influence the AR expression in the skin by topical antisense application.
KUCH
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Date: 16 Apr 2007
Time: 21:23:38
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