The Heme Scavenger Hemopexin Reverts Heme-Driven Pro-Inflammatory Phenotypic Switching of Macrophages in Sickle Cell Disease

Hemolytic diseases, such as sickle cell anemia and thalassemia, are characterized by enhanced release of hemoglobin and heme into the circulation, heme-iron loading of reticulo-endothelial system macrophages as well as chronic inflammation. Here we demonstrate that in addition to activating the vascular endothelium, hemoglobin and heme alter the macrophage phenotype in sickle cell disease.

We show that exposure of cultured macrophages to hemolytic aged red blood cells, heme or iron causes their functional phenotypic change towards a pro-inflammatory phenotype, with increased expression levels of inflammatory markers such as IL-6 (P<0.001), TNFα (P<0.0001), IL-1β (P<0.001) MHCII (P<0.001), CD86 (P<0.05) and CD14 (P<0.0001) and decreased expression levels of anti-inflammatory markers such as IL-10 (P<0.0001), CD206 (P<0.0001), Arginase-1 and Ym1. Interestingly, hemolysis and macrophage heme/iron accumulation in a mouse model of sickle cell disease triggers similar pro-inflammatory phenotypic alterations in hepatic macrophages, increasing the expression levels of IL-6 (P<0.0001), MHCII (P<0.05), CD86 (P<0.0001) and iNOS (P<0.01). On the mechanistic level, heme-induced pro-inflammatory phenotype switching of macrophages critically depends on ROS production and activation of the TLR4 signaling pathway.

We further demonstrate that the heme scavenger hemopexin protects reticulo-endothelial macrophages from heme overload in heme-loaded Hx-null mice (P<0.05) and reduces production of cytokines (IL-6, TNFα: P<0.01) and reactive oxygen species (P<0.001). Importantly, in sickle mice the administration of human exogenous hemopexin attenuates the inflammatory phenotype of macrophages, by decreasing the expression of IL-6 (P<0.05), MHCII (P<0.01), CD86 (P<0.001) and iNOS (P<0.05).

Taken together, our data suggest that therapeutical administration of hemopexin is beneficial to counteract heme-driven macrophage-mediated inflammation in sickle cell disease.