Heme Iron Causes Cytotoxicity to Lung Microvascular Endothelial Cells through Oxidant Lipid Signaling: Protection by Novel Lipophilic Thiol Chelator and Antioxidant Drug, N,N'-bis-2-Mercaptoethyl Isophthalamide (NBMI) and Implications in Treatment of Sickle Cell Disease
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Date
2014-05
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The Ohio State University
Abstract
Hereditary hemoglobinopathies, mainly sickle cell disease (SCD) and β-thalassemia, contribute to iron overload through hemolysis and hemoglobin (Hb) degradation. Hemin (Fe3+-heme), released from the extracellular Hb, is identified as a critical player in hemoglobinopathies. Heme and iron released from hemoglobin during hemoglobinopathies lead to reactive oxygen species formation (ROS) and oxidant injury to cells and tissues, including the vascular endothelium. Pulmonary hypertension has been recognized as a serious complication in SCD and lung microvasculature appears to be a target to iron-induced lung microvascular endothelial dysfunction contributing to pulmonary hypertension in SCD. Currently available iron chelation therapies for the protection against iron-induced stress in SCD patients are limited and not efficacious. Therefore, here we hypothesized that iron overload would cause reactive oxygen species (ROS) generation and oxidative stress in SCD crisis triggering the redox-regulated activation of the membrane phospholipid-hydrolyzing and cellular lipid signaling enzyme, phospholipase D (PLD) in lung microvascular endothelium, which could be the underlying mechanism(s) of pulmonary hypertension in SCD patients. In our current investigation, we established the oxidant and thiol-redox mediation of iron-induced (hemin) cytotoxicity through PLD-mediated lipid signaling in the bovine lung microvascular endothelial cell (BLMVEC) model in vitro. Additionally, our study revealed that hemin caused cytotoxicity through ROS generation, oxidative stress, depletion of glutathione (GSH) redox status, membrane lipid peroxidation, and phospholipase D (PLD) activation, formation of PLD-generated bioactive lipid mediator, phosphatidic acid (PA), and lipid signaling in BLMVECs. The PLD-specific pharmacological inhibitor, 5-fluoro-2-indolyl des-chlorohalopemide hydrochloride hydrate (FIPI), not only attenuated hemin-induced PLD activation but also protected against cytotoxicity of hemin in BLMVECs suggesting the regulatory role of PLD and the PLD-generated bioactive lipid, PA in cytotoxicity of hemin in endothelial cells (ECs). Furthermore, our current study demonstrated that N,N'-bis-2-mercaptoethyl isophthalamide (NBMI), a novel synthetic, non-toxic, and lipophilic thiol-redox heavy metal chelator and antioxidant drug, offered a greater efficacy in attenuating the hemin-induced oxidative stress and PLD activation and protection against cytotoxicity of hemin in lung microvascular ECs as compared to the currently used iron and trace metal chelating drugs and thiol-protectants, thus offering promise for the treatment of iron overload and associated vascular EC injury during hemoglobinopathies including SCD.
Description
Honorable Mention Denman Undergraduate Research Forum 2014
2013 Mayers Undergraduate Research Scholarship
2013 Arts and Sciences Autumn Undergraduate Research Scholarship
2013 Mayers Undergraduate Research Scholarship
2013 Arts and Sciences Autumn Undergraduate Research Scholarship
Keywords
Sickle Cell Disease, Iron, Cytotoxicity, Lipid Signaling, NBMI, PLD, PA, Reactive Oxygen Species