High Sulfation and a High Molecular Weight Are Important for Anti-hepcidin Activity of Heparin

Michela Asperti,Paolo Arosio,Margherita Di Somma,Annamaria Naggi,Emiliano Esposito,Magdalena Gryzik,Maura Poli,Paola Ruzzenenti

doi:10.3389/fphar.2015.00316

Michela Asperti, Paolo Arosio + Show 6 more

Open Access

https://doi.org/10.3389/fphar.2015.00316

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Abstract

Heparins are efficient inhibitors of hepcidin expression even in vivo, where they induce an increase of systemic iron availability. Heparins seem to act by interfering with BMP6 signaling pathways that control the expression of liver hepcidin, causing the suppression of SMAD1/5/8 phosphorylation. The anti-hepcidin activity persists also when the heparin anticoagulant property is abolished or reduced by chemical reactions of oxidation/reduction (glycol-split, Gs-Heparins) or by high sulfation (SS-Heparins), but the structural characteristics needed to optimize this inhibitory activity have not been studied in detail. To this aim we analyzed three different heparins (Mucosal Heparin, the Glycol split RO-82, the partially desulfated glycol-split RO-68 and the oversulfated SSLMWH) and separated them in fractions of molecular weight in the range 4–16 kD. Since the distribution of the negative charges in heparins contributes to the activity, we produced 2-O- and 6-O-desulfated heparins. These derivatives were analyzed for the capacity to inhibit hepcidin expression in hepatic HepG2 cells and in mice. The two approaches produced consistent results and showed that the anti-hepcidin activity strongly decreases with molecular weight below 7 kD, with high N-acetylation and after 2-O and 6-O desulfation. The high sulfation and high molecular weight properties for efficient anti-hepcidin activity suggest that heparin is involved in multiple binding sites.

Highlights

Iron is essential and potentially toxic, its homeostasis needs to be tightly regulated
Abnormal regulation of hepcidin expression occurs in various pathologies: when too low it leads to iron excess as in hemochromatosis (Zhao et al, 2013), when hepcidin is too high it leads to iron restriction, as in Iron Refractory Iron Deficiency Anemia (IRIDA) and in the Anemia of Chronic Disease (ACD) (Poli et al, 2014c)
Different approaches are in study for treatments to control hepcidin (Ganz and Nemeth, 2011; Fung and Nemeth, 2013; Poli et al, 2014c) and among them we found that heparins strongly repress liver hepcidin expression in vitro and in vivo (Poli et al, 2011, 2014a) by interfering with BMP6/SMAD1/5/8 phosphorylation and signaling (Poli et al, 2014a)

Summary

Introduction

Iron is essential and potentially toxic, its homeostasis needs to be tightly regulated. Different approaches are in study for treatments to control hepcidin (Ganz and Nemeth, 2011; Fung and Nemeth, 2013; Poli et al, 2014c) and among them we found that heparins strongly repress liver hepcidin expression in vitro and in vivo (Poli et al, 2011, 2014a) by interfering with BMP6/SMAD1/5/8 phosphorylation and signaling (Poli et al, 2014a) The finding that this occurs with heparins without anticoagulant activity strongly facilitated their use in vivo. The results show that the anti-hepcidin activity is associated with molecular weight above 7 kD and with the 2-O and 6-O sulfation

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