Divergent Role OF Heme Oxygenase Inhibition in the Pathogenesis of Sepsis

Abstract

The reduction of neutrophil migration to an infectious focus is associated with a high mortality in severe sepsis. Previously, we showed that heme oxygenase (HO) products downregulate neutrophil recruitment in a noninfectious inflammatory model. The present study was designed to determine the role of HO in sepsis induced by cecal ligation and puncture (CLP) model. We demonstrated that pretreatment, but not the combination of pretreatment plus posttreatment with zinc protoporphyrin IX (ZnPP IX), an HO inhibitor, prevented the reduction of CXCR2 on circulating neutrophils and the failure of intraperitoneal neutrophil migration to the site of infection. Consequently, bacterial dissemination, systemic inflammatory response, and organ injury were prevented. In addition, pretreatment with the HO inhibitor avoided hypotension and consequently increased survival. Moreover, in mice subjected to severe CLP, the pretreatment, but not the combination of pretreatment plus posttreatment with ZnPP IX, prevented the increase of plasmatic free heme observed in nontreated severe CLP. The administration of exogenous hemin to mice subjected to moderate sepsis consistently increased the mortality rate. Furthermore, hemin resulted in a reduction of neutrophil migration both in vivo and in vitro. Altogether, our results demonstrated that pretreatment with the HO inhibitor prevents the pathological findings in severe CLP. However, the combination of pretreatment plus posttreatment with ZnPP IX enhances sepsis severity because of an increase in circulating levels of heme, which is deleterious to the host tissues and also inhibits neutrophil migration.

Heme Oxygenase Inhibition Enhances Neutrophil Migration Into the Bronchoalveolar Spaces and Improves the Outcome of Murine Pneumonia-Induced Sepsis

“…In recent years, numerous studies have demonstrated that HO, together with its end products CO and BVD, is able to modulate the inflammatory response. Heme oxygenase 1 activity is capable of suppressing neutrophil rolling, adhesion, and migration, thereby preventing the entry of neutrophils into the site of inflammation (11,13). The anti-inflammatory properties of HO-1 are due to the direct inhibition of adhesion molecule expression and monocyte chemotactic protein 1 and macrophage colonystimulating factor secretion by endothelial cells (24).…”

“…Mice were pretreated with ZnPPIX 30 min before sepsis induction, as described in our previous work (13). Six hours after sepsis induction, we evaluated neutrophil migration to the bronchoalveolar spaces.…”

“…In fact, inhibitors of HO increased the rolling and adhesion of leukocytes on endothelial cells and the transmigration of neutrophils to inflammatory sites (11), whereas HO-1 overexpression, CO donors, and BVD decreased these parameters (11,12). Moreover, the administration of HO inhibitors during CLP-induced sepsis prevents the failure of neutrophils migration (13), attenuates liver damage (14), and, as consequence, reduced the mortality rate.…”

Toll-like receptor 9 activation in neutrophils impairs chemotaxis and reduces sepsis outcome*

“…GRK-2 staining by immunofluorescence was performed as previously described (16). Briefly, purified neutrophils on glass slides were fixed, permeabilized, and blocked with 1% bovine serum albumin (Sigma-Aldrich) in phosphate buffered saline containing normal goat serum (1:50).…”

“…GRKs phosphorylate serine/threonine residues on G-protein-coupled receptors, leading to receptor internalization and intracellular sorting, which results in either recycling or degradation of those receptors (10). Furthermore, the cell surface expression of CXCR2 is downregulated in severe sepsis by mechanisms that are dependent on the activation of toll-like receptors (TLRs) 2 and 4 (11)(12)(13), nitric oxide (14), tumor necrosis factor-α (15), and heme oxygenase products (16), all of which collectively induce the expression of GRK-2.…”