Figure 3

AMP kinase agonists inhibit profibrotic gene expression. Confluent dermal fibroblasts in two-dimensional monolayer cultures were incubated with the indicated concentrations of (A) AICAR or (B) metformin for 24 hours. Total RNA was subjected to real-time qPCR. The results represent the means ± SEM of triplicate determinations. C. Fibroblasts were immunostained with antibodies to Type I collagen (red), or stained with DAPI (blue). Representative immunofluorescence photomicrographs. Original magnification ×60. D. Fibroblasts transiently transfected with (SBE)₄-luc were pre-treated with AICAR or metformin (1 mM) for 24 hours, followed by TGF-β (10 ng/ml) for 24 hours. Cell lysates were assayed for their luciferase activities. The results represent the means ± SEM of triplicate determinations. AICAR, 5-amino-1-β-D-ribofuranosyl-imidazole-4-carboxamide; DAPI, 4',6-diamidino-2-phenylindole; SEM, standard error of the mean; TGF-β, transforming growth factor-β.