Biological response of human aortic endothelial cells exposed to acellular hemoglobin solutions developed as potential blood substitutes M. Toussaint-Hacquard a, * , Y. Devaux b , D. Longrois b , B. Faivre-Fiorina a , S. Muller c , J.F. Stoltz c , C. Vigneron a , P. Menu a a Laboratoire d’He ´matologie-Physiologie, UPRES EA 3452, Faculte ´ de Pharmacie, 54000 Nancy, France b UPRES EA 3447 Le ´sions-re ´parations remodelage cardiaque et arte ´riel, Faculte ´ de Me ´decine, 54500 Vandoeuvre les Nancy, France c Laboratoire de Me ´canique et Inge ´nierie Cellulaire et Tissulaire, LEMTA UMR CNRS 7563 et IFR 111 Bioinge ´nierie, Faculte ´ de Me ´decine, 54500 Vandoeuvre les Nancy, France Received 6 August 2002; accepted 11 September 2002 Abstract The cardiovascular effects of hemoglobin-based oxygen carriers (HBOCs) are mainly related to their nitric oxide (NO) scavenging properties but other effects such as the impact of these hemoglobins on the endothelial cell (EC) biology are not well understood. We hypothesized that HBOCs could modify EC functions by altering gene expression, in particular the endothelial NO synthase (NOS3) and/or by activating EC. Cultured human aortic endothelial cells (HAEC) were incubated for 3 hours with purified cell-free Hb, Dex-BTC-Hb or aa-Hb (16 g/L). Expression of NOS3 mRNA and protein were assessed by semi-quantitative RT-PCR and Western blot respectively immediately after and 24 hours after incubation. The expression and localization of the adhesion molecule ICAM-1 were detected by fluorescence microscopy. None of the solutions tested modified NOS3 mRNA and protein expression despite adequate controls that up- or down-regulate NOS3 expression. The expression and the localization of ICAM-1 on the cell membrane were modified after 3 hours of incubation with all the hemoglobin solutions tested in a manner similar to tumor necrosis factor-a. In conclusion, HAEC incubation with clinically relevant concentrations of HBOCs induced changes in the pattern of ICAM-1 expression consistent with cell activation/cell signaling mechanisms. However, HBOCs did not alter NOS3 gene expression. D 2002 Elsevier Science Inc. All rights reserved. Keywords: Cell-free hemoglobin; Endothelial cell; Nitric oxide synthase; ICAM-1; Blood substitute 0024-3205/02/$ - see front matter D 2002 Elsevier Science Inc. All rights reserved. PII:S0024-3205(02)02368-8 * Corresponding author. Department of Hematology-Physiology, Faculty of Pharmacy, 54001 Nancy Cedex, France. Tel.: +33-383-68-23-22; fax: +33-383-68-23-19. E-mail address: marie.toussaint@pharma.uhp-nancy.fr (M. Toussaint-Hacquard). www.elsevier.com/locate/lifescie Life Sciences 72 (2003) 1143– 1157