Abstract
Heme is a prosthetic group of oxygen carrier hemoglobin (Hb). During hemolytic conditions, such as sickle cell disease or malaria, heme can be released from cell-free Hb triggering toxicity through oxidative reactions. The role of heme in inflammatory processes has been extensively studied but remains not completely elucidated. Although activation of TLR4 and downstream inflammatory signaling pathways can be triggered by free heme, heme-catabolism through hemoxygenase 1 (Hmox-1) has been reported to downregulate some components of inflammation and immunity. In our laboratory, we could demonstrate that heme does not induce inflammation, suggesting that previous reports of pro-inflammatory heme-activity might have been influenced by the difficult biophysical properties of purified (protein-free) heme. In contrast, we have identified heme as a potent suppressor of inflammation in macrophages. Vascular endothelial cells, as sentinels for homeostasis in the blood stream, are among the first cells exposed to cell-free Hb and heme during hemolytic diseases.
To define the role of heme in the endothelial cell inflammatory response, we performed a number of in vitro and in vivo studies. In vitro, heme-treated endothelial cells were stimulated by IL-1β. In vivo, heme-treated mice were challenged with lipopolysaccharide (LPS) or with an agonistic anti-CD40 antibody, which induces a systemic inflammatory response with necrotizing hepatitis.
In the first part of the present study, we found that heme treatment suppresses the inflammatory signaling to IL-1β in human umbilical vein endothelial cells (HUVEC). The most interesting finding was that heme challenge suppresses the expression of the adhesion molecule VCAM-1, causing decreasing monocyte adhesion. Along the same line, in vivo heme challenge blunts the endothelial response in the context of a LPS-induced systemic inflammatory response syndrome. In a murine model of anti-CD40-induced necroinflammatory liver disease, heme-treatment rescues almost completely the phenotype of the disease.
In the second part of the study, we dissected the possible mechanisms for heme-induced hypoinflammation. Using a Hmox-1 knock-out mouse model, we could demonstrate that the hypoinflammation was related to heme itself and not to Hmox-1 activity, the main enzyme of heme-breakdown. Further, we identified Nrf2 activation as a possible candidate for heme-induced hypo-inflammation in endothelial cells. The heme gene signature of HUVEC is dominated by the expression of Nrf2 target genes. The transcription factor Nrf2 is involved in upregulation of anti-oxidant genes upon increase of oxidative stress in the cell. Recently, Kobayashi et al. described a novel property of Nrf2 in suppression of pro-inflammatory NFκB response genes transcription in macrophages.
In conclusion, we demonstrated a novel anti-inflammatory effect of heme on endothelial cells and suggested the Nrf2 anti-inflammatory pathway as a possible mechanism involved in this process.
Valkova, Kristyna