Placenta Growth Factor (PlGF), a Novel Inducer of Plasminogen Activator Inhibitor-1 (PAI-1) in Sickle Cell Disease (SCD)

Sickle cell disease (SCD) is characterized by a prothrombotic state. Plasminogen activator inhibitor-1 (PAI-1) is known to modulate fibrinolysis, lung injury/fibrosis, and angiogenesis. However, its role in SCD is less understood, and the molecular mechanisms underlying increased PAI-1 are unknown. Herein, we show a novel link between PAI-1 and sickle erythropoiesis. Plasma PAI-1 levels were high in SCD patients at steady state and in two humanized sickle mouse models, with increased PAI-1 immunolabeling in sickle mouse lung, bronchial epithelial cells, alveolar macrophages, and pulmonary microvascular endothelial cells. Placenta growth factor (PlGF), released at high levels by sickle erythroblasts, induced PAI-1 expression in primary human pulmonary microvascular endothelial cells and monocytes through activation of c-Jun N-terminal kinase (JNK), NADPH oxidase, and hypoxia-inducible factor-1α (HIF-1α). Analysis of the human PAI-1 promoter revealed this induction was mediated by hypoxia-response element (HRE)-1, HRE-2, and distal activator protein (AP-1) sites. We also identify the involvement of c-Jun, c-Jun/c-Fos, and JunD, but not JunB, in binding with AP-1 sites of the PAI-1 promoter upon PlGF induction. Consistent with these findings, levels of PAI-1 were low in PlGF knock-out mice and sickle-PlGF knock-out mice; overexpression of PlGF in normal mice increased circulating PAI-1. In conclusion, we identify a novel mechanism of PAI-1 elevation in SCD.     

Iron Regulatory Protein-1 Protects against Mitoferrin-1-deficient Porphyria

Mitochondrial iron is essential for the biosynthesis of heme and iron-sulfur ([Fe-S]) clusters in mammalian cells. In developing erythrocytes, iron is imported into the mitochondria by MFRN1 (mitoferrin-1, SLC25A37). Although loss of MFRN1 in zebrafish and mice leads to profound anemia, mutant animals showed no overt signs of porphyria, suggesting that mitochondrial iron deficiency does not result in an accumulation of protoporphyrins. Here, we developed a gene trap model to provide in vitro and in vivo evidence that iron regulatory protein-1 (IRP1) inhibits protoporphyrin accumulation. Mfrn1^+/gt;Irp1^−/− erythroid cells exhibit a significant increase in protoporphyrin levels. IRP1 attenuates protoporphyrin biosynthesis by binding to the 5′-iron response element (IRE) of alas2 mRNA, inhibiting its translation. Ectopic expression of alas2 harboring a mutant IRE, preventing IRP1 binding, in Mfrn1^gt/gt cells mimics Irp1 deficiency. Together, our data support a model whereby impaired mitochondrial [Fe-S] cluster biogenesis in Mfrn1^gt/gt cells results in elevated IRP1 RNA-binding that attenuates ALAS2 mRNA translation and protoporphyrin accumulation.