Asperti Michela, Naggi Annamaria, Esposito Emiliano, Ruzzenenti Paola, Di Somma Margherita, Gryzik Magdalena, Arosio Paolo, Poli Maura
Molecular Biology Laboratory, Department of Molecular and Translational Medicine, University of Brescia Brescia, Italy.
G. Ronzoni Institute for Chemical and Biochemical Research Milan, Italy.
Front Pharmacol. 2016 Jan 11;6:316. doi: 10.3389/fphar.2015.00316. eCollection 2015.
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.
肝素即使在体内也是高效的铁调素表达抑制剂，在体内可促使全身铁利用率增加。肝素似乎是通过干扰控制肝脏铁调素表达的骨形态发生蛋白6（BMP6）信号通路发挥作用，导致SMAD1/5/8磷酸化受到抑制。当肝素的抗凝特性通过氧化/还原化学反应（二醇裂解，Gs-肝素）或高硫酸化（SS-肝素）被消除或降低时，其抗铁调素活性依然存在，但尚未对优化这种抑制活性所需的结构特征进行详细研究。为此，我们分析了三种不同的肝素（黏膜肝素、二醇裂解的RO-82、部分去硫酸化的二醇裂解的RO-68和过度硫酸化的SSLMWH），并将它们分离成分子量在4-16kD范围内的级分。由于肝素中负电荷的分布有助于其活性，我们制备了2-O-和6-O-去硫酸化肝素。对这些衍生物在肝HepG2细胞和小鼠中抑制铁调素表达的能力进行了分析。这两种方法得出了一致的结果，表明当分子量低于7kD、具有高N-乙酰化以及2-O和6-O去硫酸化后，抗铁调素活性会大幅降低。高效抗铁调素活性所需的高硫酸化和高分子量特性表明肝素涉及多个结合位点。
