Nitric oxide reduces nontransferrin-bound iron transport in HepG2 cells

Nitric oxide (NO) donors S-nitroso-N-acetylpenicillamine (SNAP) and sodium nitroprusside (SNP) modulate iron regulatory protein (IRP) activity and may , therefore, affect iron uptake through transferrin receptor expression. Ho wever, iron also enters the cell as nontransferrin-bound iron (NTBI), and t he aim of this study was to evaluate the effects of NO donors on NTBI trans port in HepG2 cells, a model of liver physiology, Incubation with SNP and S NAP led to a time- and concentration-dependent reduction in Fe3+ and Fe2+ u ptake, thus indicating an effect on the transporter rather than on the redu ctase. In terms of Fe2+ uptake, no variations in the Michaelis-Menten const ant (K-m) and a reduction in maximum uptake (V-max) (50, 33, and 16.6 fmol/ mu g protein/min in control, SNP-, and SNAP-treated cells, respectively) we re detected, which suggested a decrease in the number of putative NTBI tran sport protein(s). Gel shift assays showed that IRP activity was reduced by SNP and slightly increased by SNAP. Northern blot analysis of transferrin r eceptor messenger RNA (mRNA) levels showed variations similar to those obse rved for IRPs, but both NO donors increased L-ferritin mRNA levels and had no effect on the stimulator of Fe transport (SFT) mRNA, In conclusion, NO d onors significantly reduce NTBI transport in HepG2 cells, an effect that se ems to be IRP and SFT independent. Moreover, the reduction in NTBI uptake a fter NO treatment suggests that this form of iron may play a minor role in the increased hepatic iron stores observed in inflammation or that other li ver cells are more involved in this pathological condition.