Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress

Abstract

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.

Data demonstrating the anti-oxidant role of hemopexin in the heart

“…Data in Fig. 5 show that the heart of Hx -/- mice, despite of heme accumulation and elevated ROS [1], did not show sign of fibrosis and inflammation apart a slight increase in the level of Tumor Necrosis Factor (TNF)α and Interleukin (IL)-6 mRNAs.

Fig. 1Heme promotes ROS formation in isolated adult rat cardiomyocytes.

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“…Primary adult rat cardiomyocytes were treated with 5 µM heme or vehicle for 15 min. Heme and Hx-heme complex were prepared as described [1].…”

“…For Western blotting, tissue and cell proteins were extracted as reported [1]. Fifty µg total protein or 0.25 µL mouse serum were separated on SDS-PAGE and immunoblotted using antibodies against HO-1 (dilution 1:300, Enzo Life Sciences), Hx (1:1000) [8], N-Tyr (1:1000, Merck Millipore).…”

“…The data presented in this article are related to the research article entitled Hemopexin counteracts systolic dysfunction induced by heme-driven oxidative stress (G. Ingoglia, C. M. Sag, N. Rex, L. De Franceschi, F. Vinchi, J. Cimino, S. Petrillo, S. Wagner, K. Kreitmeier, L. Silengo, F. Altruda, L. S. Maier, E. Hirsch, A. Ghigo and E. Tolosano, 2017) [1]. Data show that heme induces reactive oxygen species (ROS) production in primary cardiomyocytes.…”

Hydrogen Medicine Therapy: An Effective and Promising Novel Treatment for Multiple Organ Dysfunction Syndrome (MODS) Induced by Influenza and Other Viral Infections Diseases?

“…As auto-oxidation of catecholamine occurs, a large number of free radicals can be produced, accelerating formation of oxidative stress [1]. Meanwhile, oxidative stress activates the complement system, producing a variety of chemotactic substances, such as C3 fragments, leukotrienes etc, which attract and activate neutrophils ultimately [29]. Thus, inflammatory infiltrates develops in many corresponding organs.…”