17 beta-Estradiol inhibits expression of human interleukin-6 promoter-reporter constructs by a receptor-dependent mechanism.

We previously reported that 17,8-estradiol inhibits cytokine- stimulated bioassayable IL-6 and the steady-state level of IL-6 mRNA. To determine the molecular basis of this effect, the transient expression of chloramphenicol acetyltransferase (CAT) reporter plasmid driven by the human IL-6 promoter was studied here in HeLa or murine bone marrow stromal cells (MBA 13.2). 1713-estradiol (10-8 M) completely suppressed stimulated CAT expression in HeLa cells cotransfected with IL-6/CAT constructs and a human estrogen receptor (hER) expression plasmid; but had no effect on reporter expression in HeLa cells not transfected with hER. 17fl-estradiol also inhib- ited stimulated expression in MBA 13.2 cells (which express the estrogen receptor constitutively) without the requirement of cotransfection of the hER plasmid. The hormonal effects were indistinguishable between constructs containing a 1.2-kb fragment of the 5' flanking region of the IL-6 gene or only the proximal 225-bp fragment. However, yeast-derived recombi- nant hER did not bind to the 225-bp segment in DNA band shift assays, nor did the 225-bp fragment compete for binding of an estrogen response element oligonucleotide to yeast-derived es- trogen receptor. These data suggest that 17,6-estradiol inhibits the stimulated expression of the human IL-6 gene through an estrogen receptor mediated indirect effect on the transcriptional activity of the proximal 225-bp sequence of the promoter. (J. Clin. Invest. 1994 . 93:944-950.) Key words: cytokines * estrogen * stromal cells -osteoporosis * receptors MD) (specific activity = 1.8 X 107 U/mg). Recombinant murine TNF-a was purchased from Boehringer Mannheim Corp. (Indianapolis, IN) (specific activity = 6 X 107 U/mg). '251-16-a-iodo-3,170estradiol (2,200 Ci/mmol) was purchased from New England Nuclear 1.