Neuroprotection of brain-permeable iron chelator VK-28 against intracerebral hemorrhage in mice

Iron overload plays a key role in the secondary brain damage that develops after intracerebral hemorrhage (ICH). The significant increase in iron deposition is associated with the generation of reactive oxygen species (ROS), which leads to oxidative brain damage. In this study, we examined the protective effects of VK-28, a brain-permeable iron chelator, against hemoglobin toxicity in an ex vivo organotypic hippocampal slice culture (OHSC) model and in middle-aged mice subjected to an in vivo, collagenase-induced ICH model. We found that the effects of VK-28 were similar to those of deferoxamine (DFX), a well-studied iron chelator. Both decreased cell death and ROS production in OHSCs and in vivo, decreased iron-deposition and microglial activation around hematoma in vivo, and improved neurologic function. Moreover, compared with DFX, VK-28 polarized microglia to an M2-like phenotype, reduced brain water content, deceased white matter injury, improved neurobehavioral performance, and reduced overall death rate after ICH. The protection of VK-28 was confirmed in a blood-injection ICH model and in aged-male and young female mice. Our findings indicate that VK-28 is protective against iron toxicity after ICH and that, at the dosage tested, it has better efficacy and less toxicity than DFX does.

Iron Chelation by Deferoxamine Induces Autophagy

Deferoxamine (DFO) is a high affinity Fe (III) chelator produced by Streptomyces pilosus that is used clinically to remove systemic iron in secondary iron overload disorders. DFO cannot be absorbed through the intestine and must be injected. As shown previously, De Domenico et al. EMBO J (2006), expression of Ferroportin (Fpn), the only mammalian iron exporter, can deplete cells of ferritin by lowering cytosolic iron and by exporting iron from cells. Fpn-mediated iron loss induces ferritin degradation by the proteosome. In this study we show that permeable iron chelators, desferirax or deferriprone also induce the proteosomal degradation of ferritin. In contrast, DFO-mediated iron chelation at clinically useful concentrations, leads to ferritin degradation in lysosomes. Immunochemical analysis revealed that DFO-treated cells show increased levels of LC3B, a protein required for autophagy, suggesting that DFO induces autophagy. Treatment of cells with desferasirox or deferriprone did not lead to accumulation of LC3B. Studies using high molecular weight conjugates of DFO or inhibitors of endocytosis showed that the presence of DFO in lysosomes was responsible for the induction of autophagy. Incubation of DFO-treated cells with 3-methyladenine, an autophagy inhibitor, does not, however, prevent ferritin loss suggesting there may be an alternate route for ferritin degradation. This hypothesis was confirmed by examining the effect of the proteosome inhibitor, MG132, on DFOinduced autophagy in cells treated with DFO and 3-methyladenine. Addition of MG132 to 3-methyladenine treated cells prevents ferritin degradation. These results indicate that ferritin degradation occurs by two routes: a DFO-induced entry of ferritin into lysosomes and a cytosolic route in which iron is extracted from ferritin prior to degradation by the proteosome.

Analysis of yggX and gshA Mutants Provides Insights into the Labile Iron Pool in Salmonella enterica

ABSTRACT Strains of Salmonella enterica lacking YggX and the cellular reductant glutathione exhibit defects similar to those resulting from iron deficiency and oxidative stress. Mutant strains are sensitive to hydrogen peroxide and superoxide, deregulate the expression of the Fur-regulated gene entB, and fail to grow on succinate medium. Suppression of some yggX gshA mutant phenotypes by the cell-permeable iron chelator deferoxamine allowed the conclusion that increased levels of cellular Fenton chemistry played a role in the growth defects. The data presented are consistent with a scenario in which glutathione acts as a physiological chelator of the labile iron pool and in which YggX acts upstream of the labile iron pool by preventing superoxide toxicity.