Interleukin-6 and Lipopolysaccharide Modulate Hepcidin mRNA Expression by Hepg2 Cells

Abstract

Iron homeostasis is controlled by hepcidin (Hpc) as well as other ways. Hpc expression is regulated by iron (Fe) storage and by inflammation, but the joint effect of both stimuli remains unclear. We studied the modulatory role of inflammatory agents (IL6 and LPS) over Hpc and DMT1 mRNA expression in HepG2 cells preloaded with Fe. HepG2 cells were preloaded with different Fe concentrations (holo-Tf or Fe-NTA) and then incubated with IL6 or LPS. We measured intracellular Fe levels by AAS with graphite furnace, transferrin receptor (TfR) by ELISA and mRNA relative abundance of Hpc and DMT1 by qRT-PCR. The maximum effect on Fe uptake was observed in cells incubated with 30ng/ml IL6 (p < 0.01) and 500ng/ml LPS (p < 0.05). In HepG2 cells preloaded with holo-Tf or Fe-NTA and challenged with IL6 and LPS, we observed a decreased: (a) Hpc mRNA relative abundance (two-way ANOVA: p < 0.05 and p < 0.001, respectively), (b) DMT1 mRNA relative abundance and TfR1 protein levels (two-way ANOVA: p < 0.001), and (c) intracellular Fe concentration (two-way ANOVA: p < 0.001 and p < 0.01, respectively) compared to control cells incubated only with Fe (holo-Tf or Fe-NTA). Our results support the idea that Fe storage and inflammation act together to regulate Fe homeostasis and suggest a negative regulation in this hepatic cellular model to prevent excessive increases in Hpc.