Macrophage metabolic rewiring improves heme-suppressed efferocytosis and tissue damage in sickle cell disease.

Sickle cell disease (SCD) is hallmarked by an underlying chronic inflammatory condition, which is contributed by heme-activated proinflammatory macrophages. Although previous studies addressed heme ability to stimulate macrophage inflammatory skewing through Toll-like receptor4 (TLR4)/reactive oxygen species signaling, how heme alters cell functional properties remains unexplored. Macrophage-mediated immune cell recruitment and apoptotic cell (AC) clearance are relevant in the context of SCD, in which tissue damage, cell apoptosis, and inflammation occur owing to vaso-occlusive episodes, hypoxia, and ischemic injury. Here we show that heme strongly alters macrophage functional response to AC damage by exacerbating immune cell recruitment and impairing cell efferocytic capacity. In SCD, heme-driven excessive leukocyte influx and defective efferocytosis contribute to exacerbated tissue damage and sustained inflammation. Mechanistically, these events depend on heme-mediated activation of TLR4 signaling and suppression of the transcription factor proliferator-activated receptor γ (PPARγ) and its coactivator peroxisome proliferator-activated receptor γ coactivator 1α (PGC1α). These changes reduce efferocytic receptor expression and promote mitochondrial remodeling, resulting in a coordinated functional and metabolic reprogramming of macrophages. Overall, this results in limited AC engulfment, impaired metabolic shift to mitochondrial fatty acid ß-oxidation, and, ultimately, reduced secretion of the antiinflammatory cytokines interleukin-4 (IL-4) and IL-10, with consequent inhibition of continual efferocytosis, resolution of inflammation, and tissue repair. We further demonstrate that impaired phagocytic capacity is recapitulated by macrophage exposure to plasma of patients with SCD and improved by hemopexin-mediated heme scavenging, PPARγ agonists, or IL-4 exposure through functional and metabolic macrophage rewiring. Our data indicate that therapeutic improvement of heme-altered macrophage functional properties via heme scavenging or PGC1α/PPARγ modulation significantly ameliorates tissue damage associated with SCD pathophysiology.

Vasculo-toxic and pro-inflammatory action of unbound haemoglobin, haem and iron in transfusion-dependent patients with haemolytic anaemias.

Increasing evidence suggests that free haem and iron exert vasculo-toxic and pro-inflammatory effects by activating endothelial and immune cells. In the present retrospective study, we compared serum samples from transfusion-dependent patients with ß-thalassaemia major and intermedia, hereditary spherocytosis and sickle cell disease (SCD). Haemolysis, transfusions and ineffective erythropoiesis contribute to haem and iron overload in haemolytic patients. In all cohorts we observed increased systemic haem and iron levels associated with scavenger depletion and toxic 'free' species formation. Endothelial dysfunction, oxidative stress and inflammation markers were significantly increased compared to healthy donors. In multivariable logistic regression analysis, oxidative stress markers remained significantly associated with both haem- and iron-related parameters, while soluble vascular cell adhesion molecule 1 (sVCAM-1), soluble endothelial selectin (sE-selectin) and tumour necrosis factor α (TNFα) showed the strongest association with haem-related parameters and soluble intercellular adhesion molecule 1 (sICAM-1), sVCAM-1, interleukin 6 (IL-6) and vascular endothelial growth factor (VEGF) with iron-related parameters. While hereditary spherocytosis was associated with the highest IL-6 and TNFα levels, ß-thalassaemia major showed limited inflammation compared to SCD. The sVCAM1 increase was significantly lower in patients with SCD receiving exchange compared to simple transfusions. The present results support the involvement of free haem/iron species in the pathogenesis of vascular dysfunction and sterile inflammation in haemolytic diseases, irrespective of the underlying haemolytic mechanism, and highlight the potential therapeutic benefit of iron/haem scavenging therapies in these conditions.