Boretti FS, Buehler PW, D’Agnillo F, et al. . J Clin Invest. 2009;119:2271-80.
Sequestration of extracellular hemoglobin within a haptoglobin complex decreases its hypertensive and oxidative effects in dogs and guinea pigs
 

The red blood cell packages hemoglobin and has evolved as the ideal oxygen delivery van. Its anti-oxidant environment, rich in glutathione, catalase, superoxide dismutase, etc., maintains iron in a reduced state to carry oxygen and prevents oxidation of globin and lipids. However, if hemoglobin is released, such as during intravascular hemolysis, naked hemoglobin scavenges the potent vasodilator nitric oxide and is readily oxidized to methemoglobin, thereby catalyzing oxidative damage to the vasculature and organs through reactive iron. These mechanisms underlie pathophysiologies common to pulmonary hypertension in sickle cell disease, toxicities of hemoglobinbased oxygen carriers, and mismatched transfusion reactions.1 Therapies including inhaled nitric oxide or phosphodiesterase inhibitors have been utilized to increase bio-available nitric oxide in attempts to ameliorate these toxicities. Fortunately, evolution has provided mechanisms to clear free hemoglobin and heme, including haptoglobin and hemopexin. Haptoglobin binds hemoglobin very tightly and delivers the complex to CD163 on macrophages for subsequent induction of heme oxygenase-1 and detoxification. Free heme binds hemopexin and is cleared by CD91, also inducing heme oxygenase-1 (Figure). Heme oxygenase detoxifies hemoglobin in cells and through the enzymatic process provides cytoprotectants, such as carbon monoxide, biliverdin/bilirubin, and ferritin. In chronic hemolytic diseases, haptoglobin and hemopexin are rapidly cleared from the plasma, leaving the vasculature vulnerable to hemoglobin’s toxic ways. Boretti et al. in Dominik Schaer’s laboratory in Zurich, along with investigators at the FDA, have gone back to the body’s natural defense against free hemoglobin toxicity, haptoglobin, to modify hemoglobin’s hypertensive and oxidative effects. The investigators infused experimental animals with stroma-free hemoglobin (SFHb), which induced arterial hypertension. In guinea pigs (which are similar to humans in blood and tissue antioxidant profiles) this was associated with hemoglobinuria and accumulation of iron and oxidized proteins in the kidneys. However, if SFHb was given with a 1:1 mixture of human haptoglobin, there was marked blunting of the hypertensive response, more rapid clearance of hemoglobin from the circulation, prevention of hemoglobinuria and renal iron deposition, and less oxidized protein in the kidneys. In beagles, there were no changes in mean arterial pressure or systemic vascular resistance after SFHb infusion if the animals were first treated with prednisone (4 mg/kg twice daily for 3 days), which increased haptoglobin levels 4-fold. The protective effects were dependent on highaffinity hemoglobin-haptoglobin complex formation. Interestingly, hemoglobin within the complexes was still able to bind and release oxygen and neither NO binding/oxidation rate constants nor auto-oxidation were effected.2 These data suggest that haptoglobin blunting of SFHb-hypertensive effect may be due to mechanisms other than NO inactivation.

A Model of Intravascular Hemolysis: Pathological and Adaptive Responses. Free hemoglobin can lead to pathological vasoconstriction and vasculopathy by limiting NO bioavailability and potentiating oxidative stress. An adaptive response occurs when hemoglobin is bound to haptoglobin and cleared by CD163 or heme binds to hemopexin cleared by CD91with induction of cytoprotectants including heme oxygenase-1. In turn, an anti-inflammatory/anti-oxidant response protects the vasculature.2 Copyright 2009 by American Society for Clinical Investigation. Reproduced with permission of American Society for Clinical Investigation.
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A Model of Intravascular Hemolysis: Pathological and Adaptive Responses. Free hemoglobin can lead to pathological vasoconstriction and vasculopathy by limiting NO bioavailability and potentiating oxidative stress. An adaptive response occurs when hemoglobin is bound to haptoglobin and cleared by CD163 or heme binds to hemopexin cleared by CD91with induction of cytoprotectants including heme oxygenase-1. In turn, an anti-inflammatory/anti-oxidant response protects the vasculature.2 Copyright 2009 by American Society for Clinical Investigation. Reproduced with permission of American Society for Clinical Investigation.

A Model of Intravascular Hemolysis: Pathological and Adaptive Responses. Free hemoglobin can lead to pathological vasoconstriction and vasculopathy by limiting NO bioavailability and potentiating oxidative stress. An adaptive response occurs when hemoglobin is bound to haptoglobin and cleared by CD163 or heme binds to hemopexin cleared by CD91with induction of cytoprotectants including heme oxygenase-1. In turn, an anti-inflammatory/anti-oxidant response protects the vasculature.2 Copyright 2009 by American Society for Clinical Investigation. Reproduced with permission of American Society for Clinical Investigation.
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A Model of Intravascular Hemolysis: Pathological and Adaptive Responses. Free hemoglobin can lead to pathological vasoconstriction and vasculopathy by limiting NO bioavailability and potentiating oxidative stress. An adaptive response occurs when hemoglobin is bound to haptoglobin and cleared by CD163 or heme binds to hemopexin cleared by CD91with induction of cytoprotectants including heme oxygenase-1. In turn, an anti-inflammatory/anti-oxidant response protects the vasculature.2 Copyright 2009 by American Society for Clinical Investigation. Reproduced with permission of American Society for Clinical Investigation.

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These studies aid in understanding toxicities due to free plasma hemoglobin and support potential therapeutic applications for haptoglobin in preventing them. As nicely pointed out by Kato2 in an accompanying commentary to this article (Figure), corticosteroids may improve hemolysis by enhancing clearance of hemoglobin through induction of haptoglobin and CD163. Furthermore, in sickle cell disease, haptoglobin levels are low. Would increased levels modulate sickle vasculopathy? In turn, if hemoglobin/haptoglobin complexes induce heme oxygenase-1, another adaptive protective stratagem would be triggered in hemolytic disease.3  Patients with intravascular hemolysis and vascular damage in the brain and kidney due to thrombotic thrombocytopenic purpura benefit from plasma exchange not only by removing ADAMTS13 antibodies and replacing ADAMTS enzyme, but also by adding fresh haptoglobin, preventing hemoglobin and heme catalyzed oxidative stress. Possibly, studies on haptoglobin polymorphisms in vascular diseases may reflect these underlying pathophysiologies. For now, there are few pharmacologic mechanisms to increase plasma haptoglobin, but future research should focus on this important mop for hemoglobin.

2.
Kato GJ. Haptoglobin halts hemoglobin's havoc. J Clin Invest. 2009;119:2140-42.
3.
Belcher JD, Beckman JD, Balla G, et al. Heme Degradation and Vascular Injury. Antioxid Redox Signal. 2009. [Epub ahead of print]

Competing Interests

Dr. Vercellotti indicated no relevant conflicts of interest.