Heme induces significant neutrophil adhesion in vitro via an NF kappa B and reactive oxygen species-dependent pathway

Author(s):Show less -

Miguel, Lediana I. ^[1] ; Leonardo, Flavia C. ^[1] ; Torres, Lidiane S. ^[1] ; Garcia, Flavia ^[1] ; Mendonca, Rafaela ^[1] ; Ferreira, Jr., Wilson A. ^[1] ; Gotardo, Erica M. F. ^[1] ; Fabris, Fernanda C. Z. ; Brito, Pamela L. ^[1] ; Costa, Fernando F. ^[1] ; Conran, Nicola ^[1]

Affiliation

^[1] Cidade Univ, Hematol & Transfus Ctr, Rua Carlos Chagas, 480, Barao Geraldo, BR-13083970 Campinas, SP - Brazil

• Document type: Journal article
• Source: Molecular and Cellular Biochemistry; v. 476, n. 11 JUN 2021.
• Web of Science Citations: 0

Abstract

Intravascular hemolysis, a major manifestation of sickle cell disease (SCD) and other diseases, incurs the release of hemoglobin and heme from red blood cells, in turn triggering inflammatory processes. This study investigated the in vitro effects of heme, a major inflammatory DAMP, on the adhesive properties of isolated human neutrophils. Heme (20 and 50 mu M) significantly increased the adhesion of neutrophils to fibronectin and to recombinant ICAM-1, under static conditions, even more efficiently than the potent pro-inflammatory cytokine, tumor necrosis factor-alpha (TNF); a microfluidic assay confirmed that heme stimulated neutrophil adhesion under conditions of shear stress. Heme-induced neutrophil adhesion was associated with the increased activities, but not expressions, of the Mac-1 and LFA-1 integrin subunits, CD11b and CD11a, on the cell surface. Notably, heme (50 mu M) significantly induced NF kappa B translocation in neutrophils, and inhibition of NF kappa B activity with the BAY11-7082 molecule abolished heme-induced cell adhesion to fibronectin and significantly decreased CD11a activity. Flow cytometric analysis demonstrated major reactive oxygen species (ROS) generation in neutrophils following heme stimulation that could be inhibited by the antioxidant, alpha-tocopherol, and by BAY11-7082. Furthermore, co-incubation with alpha-tocopherol abrogated both heme-stimulated neutrophil adhesion and CD11a/CD11b activation. Thus, our data indicate that heme, at clinically relevant concentrations, is a potent activator of neutrophil adhesion, increasing the ligand affinity of the beta 2 integrins via a mechanism that may be partially mediated by an NFkB-dependent pathway and the generation of ROS. Given the fundamental role that the adhesion of neutrophils to the vascular wall plays in SCD vaso-occlusion and other vascular inflammatory processes, our findings provide further evidence that cell-free heme is a major therapeutic target in the hemolytic diseases. (AU)

• FAPESP's process: 18/08010-9 - Characterization of the inflammatory effects of intravascular hemolytic processes, in vitro and in vivo
• Grantee: Nicola Amanda Conran Zorzetto
• Support Opportunities: Regular Research Grants