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Fig. 4. Regulation of Ltbp-1 expression by the AhR. (A) AhR–/– MEFs were treated for 3 and 6 hours with 1.5 µM actinomycin D (Act D) or 175 µM cycloheximide (CHX). Ltbp-1 and ß-actin mRNA expression were analyzed by northern blot using 10 µg total RNA. (B) AhR+/+ MEF were treated with the solvent DMSO (–) or 10 nM of the AhR ligand TCDD (+) and Ltbp-1 expression analyzed by RT-PCR. (C) AhR+/+ MEF were treated with 10 ng/ml recombinant TGFß protein for 1.5 or 3 hours. AhR and Ltbp-1 expression were determined by northern blot using 10 µg total RNA. (D) AhR–/– MEF were treated with 10 ng/ml recombinant TGFß protein for the indicated times and Ltbp-1 expression measured by northern blot using 10 µg total RNA. (E) AhR–/– MEF were treated for 24 hours with 1 µg/ml neutralizing anti-TGFß antibody and Ltbp-1 expression analyzed by northern blot using 10 µg total RNA. The levels of the 28S rRNA shown in panels C, D and E were obtained from ethidium bromide-stained agarose gels and were used to confirm equal loading. The experiments were done in duplicate in two MEF cultures.
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