Heparanase Overexpression Reduces Hepcidin Expression, Affects Iron Homeostasis and Alters the Response to Inflammation
Fig 2
HepG2 clones overexpressing heparanase showed a reduction of hepcidin expression and indices of iron loading.
Two stable clones of HepG2 cells transfected with pcDNA3.1-HPA (HPA3 and HPA6) were analyzed for hepcidin expression, BMP/SMAD signaling and indices of iron status. (A) qPCR was assessed to analyze the level of HPA mRNA. (B) Western blot for HPA shows the level of the latent (65 kDa) and active (50 kDa) forms. Densitometry quantification of the two forms was performed in relation to actin as calibrator. (C) qPCR was performed to analyze the level of hepcidin mRNA, (D) and the level of Id1 mRNA in relation to Hprt1. (E) WB of phosphorylated SMAD5 and their densitometry quantification referred to actin and WB of total SMAD5, (F) WB of transferrin Receptor 1, Tfr1, and its densitometry, (G) WB of ferritin light chain, FTL and its densitometry; (H) WB of Ferroportin (FPN) and its densitometry. The values are expressed as–dCt (for HPA mRNA) or as fold change over the control (MOCK) (for hepcidin and Id1 mRNA). The images are representative from three different analyses
