Heme triggers TLR4 signaling leading to endothelial cell activation and vaso-occlusion in murine sickle cell disease

Belcher, John D.; Chen, Chunsheng; Nguyen, Julia; Milbauer, Liming; Abdulla, Fuad; Alayash, Abdu I.; Smith, Ann; Nath, Karl A.; Hebbel, Robert P.; Vercellotti, Gregory M.

doi:10.1182/blood-2013-04-495887

Cited by 599 publications

(869 citation statements)

References 55 publications

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“…Recent studies have highlighted the potential therapeutic role of exogenous Hx in mouse models of sickle cell disease. In sickle cell disease mice, exogenous Hx markedly reduces inflammatory vasculopathy, prevents the development of severe acute vaso occlusive events and improves cardiac function [9,23,24]. Given these data, we asked whether the beneficial effect of Hx on the cardiovascular system might be due not only to preservation of vascular homeostasis but also to specific effects on the heart.…”

Section: Introductionmentioning

confidence: 99%

Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress

Ingoglia

Sag

Rex

et al. 2017

Free Radical Biology and Medicine

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Heart failure is a leading cause of morbidity and mortality in patients affected by different disorders associated to intravascular hemolysis. The leading factor is the presence of pathologic amount of pro-oxidant free heme in the bloodstream, due to the exhaustion of the natural heme scavenger Hemopexin (Hx). Here, we evaluated whether free heme directly affects cardiac function, and tested the therapeutic potential of replenishing serum Hx for increasing serum heme buffering capacity.The effect of heme on cardiac function was assessed in vitro, on primary cardiomyocytes and H9c2 myoblast cell line, and in vivo, in Hx -/-mice and in genetic and acquired mouse models of intravascular hemolysis. Purified Hx or anti-oxidants N-Acetyl-L-cysteine and α-tocopherol were used to counteract heme cardiotoxicity.In mice, Hx loss/depletion resulted in heme accumulation and enhanced reactive oxygen species (ROS) production in the heart, which ultimately led to severe systolic dysfunction. Similarly, high ROS reduced systolic Ca 2+ transient amplitudes and fractional shortening in primary cardiomyocytes exposed to free heme. In keeping with these Ca 2+ handling alterations, oxidation and CaMKIIdependent phosphorylation of Ryanodine Receptor 2 were higher in Hx -/-hearts than in controls.Administration of anti-oxidants prevented systolic failure both in vitro and in vivo. Intriguingly, Hx rescued contraction defects of heme-treated cardiomyocytes and preserved cardiac function in hemolytic mice.We show that heme-mediated oxidative stress perturbs cardiac Ca 2+ homeostasis and promotes contractile dysfunction. Scavenging heme, Hx counteracts cardiac heme toxicity and preserves left ventricular function. Our data generate the rationale to consider the therapeutic use of Hx to limit the cardiotoxicity of free heme in hemolytic disorders.

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Section: Introductionmentioning

confidence: 99%

Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress

Ingoglia

Sag

Rex

et al. 2017

Free Radical Biology and Medicine

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“…Our biochemical results also suggest that anti-oxidant therapy might be beneficial to reduce the stress caused by autoxidation, hemin loss, globin precipitation, and red cell lysis. Free hemin is itself quite toxic, generating free radicals, promoting lipid oxidation, and causing inflammation by binding to TLR4-like receptors on macrophages (45)(46)(47).…”

Section: Journal Of Biological Chemistry 2549mentioning

confidence: 99%

Hemoglobin Kirklareli (α H58L), a New Variant Associated with Iron Deficiency and Increased CO Binding

Bissé

Schaeffer-Reiss

Dorsselaer

et al. 2017

Journal of Biological Chemistry

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Edited by F. Peter GuengerichMutations in hemoglobin can cause a wide range of phenotypic outcomes, including anemia due to protein instability and red cell lysis. Uncovering the biochemical basis for these phenotypes can provide new insights into hemoglobin structure and function as well as identify new therapeutic opportunities. We report here a new hemoglobin ␣ chain variant in a female patient with mild anemia, whose father also carries the trait and is from the Turkish city of Kirklareli. Both the patient and her father had a His-58(E7) 3 Leu mutation in ␣1. Surprisingly, the patient's father is not anemic, but he is a smoker with high levels of HbCO (ϳ16%). To understand these phenotypes, we examined recombinant human Hb (rHb) Kirklareli containing the ␣ H58L replacement. Mutant ␣ subunits containing Leu-58(E7) autoxidize ϳ8 times and lose hemin ϳ200 times more rapidly than native ␣ subunits, causing the oxygenated form of rHb Kirklareli to denature very rapidly under physiological conditions. The crystal structure of rHb Kirklareli shows that the ␣ H58L replacement creates a completely apolar active site, which prevents electrostatic stabilization of bound O 2 , promotes autoxidation, and enhances hemin dissociation by inhibiting water coordination to the Fe(III) atom. At the same time, the mutant ␣ subunit has an ϳ80,000-fold higher affinity for CO than O 2 , causing it to rapidly take up and retain carbon monoxide, which prevents denaturation both in vitro and in vivo and explains the phenotypic differences between the father, who is a smoker, and his daughter.

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“…Coinfusion of haptoglobin (protein scavenger) and hemopexin (heme scavenger) were effective in reversing these inflammatory responses 8. More recently, microparticles laden with Hb were found to be a source for heme released into the microvasculature 9.…”

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confidence: 99%

Differential heme release from various hemoglobin redox states and the upregulation of cellular heme oxygenase‐1

et al. 2016

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Despite advances in our understanding of the oxidative pathways mediated by free hemoglobin (Hb), the precise contribution of its highly reactive redox forms to tissue and organ toxicities remains ambiguous. Heme, a key degradation byproduct of Hb oxidation, has recently been recognized as a damage‐associated molecular pattern (DAMP) molecule, able to trigger inflammatory responses. Equally damaging is the interaction of the highly redox active forms of Hb with other biological molecules. We determined the kinetics of heme loss from individual Hb redox states—ferrous (Fe2+), ferric (Fe3+), and ferryl (Fe4+)—using two different heme receptor proteins: hemopexin (Hxp), a naturally occurring heme scavenger in plasma, and a double mutant (H64Y/V86F), apomyoglobin (ApoMb), which avidly binds heme released from Hb. We show for the first time that ferric Hb (Fe3+) loses heme at rates substantially higher than that of ferryl Hb (Fe4+). This was also supported by a higher expression of heme oxygenase‐1 (HO‐1) when ferric Hb was added to cultured lung alveolar epithelial cells (E10). The reported cytotoxicity of Hb may therefore be attributed to a combination of accelerated heme loss from the ferric form and protein radical formation associated with ferryl Hb. Targeted therapeutic interventions can therefore be designed to curb specific oxidative pathways of Hb in hemolytic anemias and when Hb is used as an oxygen‐carrying therapeutic.

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Heme triggers TLR4 signaling leading to endothelial cell activation and vaso-occlusion in murine sickle cell disease

Abstract: Key Points Heme, released from hemoglobin, elicits vaso-occlusion in transgenic sickle mice via endothelial TLR4 signaling. Heme/TLR4 signaling activates NF-κB and triggers vaso-occlusion through Weibel-Palade body degranulation and adhesion molecule expression.

Cited by 599 publications

References 55 publications

Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress

Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress

Hemoglobin Kirklareli (α H58L), a New Variant Associated with Iron Deficiency and Increased CO Binding

Differential heme release from various hemoglobin redox states and the upregulation of cellular heme oxygenase‐1

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