An efficient and economic high-throughput cell screening model targeting the glucocorticoid receptor

Abstract

Aim: To discover compounds or proteins that can efficiently bind the glucocorticoid receptor (GR) and trigger the transcription of target genes, resulting in clinical improvement of diseases such as rheumatoid arthritis, asthma, inflammatory bowel disease, a high-throughput drug screening cell model using green fluorescent protein 4 (GFP4) as a marker expressed in response to GR activation has been established and evaluated. Methods: Eight repeats of the glucocorticoid response element (GRE) were cloned into the Peak12SxSynGFP4 vector, and the resulting recombinant plasmid Peak12GRE8 × SxSynGFP4 was stably transfected into the 293E cells. The stable and sensitive cell line 293E/GRE8 × /GFP4 was selected by dexamethasone (DEX) using fluorescent microscopy and fluorescence-activated cell sorting. DEX induction and phorbol myristate acetate (PMA) inhibition of the green fluorescence intensity of the cell line were tested. Results: The expression of GFP4 in the cell line was under the control of GRE, up-regulated by DEX treatment and down-regulated by phorbol myristate acetate (PMA). The up-regulation of the GFP4 expression was DEX concentration-dependent, with an EC₅₀ at approximately 5 × 10^{− 8} M. The down-regulation of the GFP4 expression was phorbol myristate acetate (PMA) concentration-dependent, with an IC₅₀ at approximately 3 × 10^{− 6} gl − 1. The expression of GFP4 was effectively activated when cells were treated with triamcinolone acetonide. Conclusion: This drug screening cell line can be used for GR-targeted high-throughput drug screening for the treatment of inflammatory diseases.

Keywords: :

• Glucocorticoid receptor
• high-throughput drug screening
• cell model