INFORME DE LA ADMINISTRACIÓN – De Enero a Diciembre de 2015
10) PREMIOS Y RECONOCIMIENTOS
monocyte function. Recently, Mitsuhata et al. reported that isoflurane inhibits the endotoxin- induced TNF-α and IL-1β secretion (7). Because isoflurane alters the activation of adhesion receptors on neutrophils (9), we investigated the effect of isoflurane on monocyte selectin and β2-integrin activation.
In the present study we used an in vitro whole blood model and two-colour flow cytometry to clarify if isoflurane alters the expression of monocyte adhesion receptors. Expression of monocyte adhesion receptors was measured in unstimulated cells and following activation with the bacterial peptide FMLP, which is a physiological agonist for the FMLP receptor on the monocyte cell surface. Recently, we have shown that our experimental setting allows rapid equilibration of the isoflurane concentration between the gas and fluid phase (12). However, due to the slightly smaller partition coefficient of isoflurane in diluted whole blood (12), the dissolved amount of isoflurane in our setting is 15% lower compared with fresh whole blood (13).
PSGL-1 is the essential ligand mediating rolling on endothelial cells, which is the first step in transendothelial leucocyte recruitment (2). Activation of leucocytes decreases surface expression of PSGL-1 by an as yet unidentified mechanism and decreases binding to endo- thelial P-selectin under conditions of flow (14). The data presented in this paper demonstrate that isoflurane reduces surface PSGL-1 expression in unstimulated and FMLP-stimulated monocytes. The underlying mechanism could be a general activation of the monocyte or a direct effect on the removal of PSGL-1 from the cell surface by isoflurane without activating other cell functions. As no increase in the CD11b expression or decrease in surface L-selectin expression was detected in this study, we conclude that direct activation of monocytes by isoflurane, comparable to that following FMLP or platelet activating factor (PAF), is unlikely to be the reason for the removal of PSGL-1 from the monocyte cell surface. In vitro studies showed that moderate decreases in PSGL-1 surface expression dramatically reduced bind- ing to immobilized P-selectin under flow conditions (2,14). Since isoflurane reduced surface PSGL-1 expression even in unstimulated monocytes, we suggest that isoflurane may inhibit the initial contact between monocytes and endothelial cells. However, the in vivo functional significance of this alteration on monocyte recruitment remains to be determined.
Following leucocyte rolling on endothelial cells, tight adhesion and transmigration is mediated by binding of the β2-integrins CD11a and CD11b to endothelial ICAM-1. Recently, Shang et al. (15) investigated the role of both β2-integrins on monocyte migration through
human umbilical vein endothelial cells (HUVEC). Blocking CD11a or CD11b alone by using monoclonal antibodies did not inhibit monocyte transmigration, but blockade of both integrins partially inhibited monocyte migration across HUVECs. CD11a is not constitutively adhesive and external stimulation with FMLP, cytokines or chemokines is a prerequisite for receptor activation (4). To enable binding of CD11a to ICAM-1, CD11a is redistributed after activation over the monocyte cell surface to form high avidity clusters with ligand binding
activity. In the present study, stimulation with FMLP increased the mean fluorescence inten- sity of CD11a on the monocyte surface, representing activation of the CD11a ligand binding activity. This activation process was significantly inhibited following exposure of monocytes to 1 MAC isoflurane, but isoflurane had no significant effect on monocyte CD11b expression. Accordingly, we suggest that the inhibition of the activation of CD11a by isoflurane alone might have no impact on monocyte transmigration through endo- thelial cells. In contrast, we have previously shown that isoflurane inhibits the activation of both β2-integrins on neutrophils (9), which could partially explain the inhibiting effect of isoflurane on neutrophil accumulation during ischaemia-reperfusion injury.
In conclusion, this study demonstrated that isoflurane in clinically used concentrations increases the removal of the selectin PSGL-1 from the monocyte surface. Since PSGL-1 is im- portant during the initial step of monocyte adhesion to endothelial P-selectin, the decrease in monocyte surface PSGL-1 may have profound effects on monocyte–endothelial interac- tions. Furthermore, the effects of isoflurane on monocyte adhesion molecule expression are different from those reported for neutrophils.
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1. Von Andrian UH, Chambers JD, McEvoy LM, Bargatze RF, Arfors KE, Butcher EC. Two-step model of leukocyte–endothelial interaction in inflammation: distinct roles for LECAM-1 and the leukocyte β2 integrins in vivo. Proc Natl Acad Sci U S A 1991; 88: 7538–42.
2. Lim YC, Snapp K, Kansas GS, Camphausen R, Ding H, Luscinskas FW. Important contributions of P- selectin glycoprotein-1-mediated secondary capture to human monocyte adhesion to P-selectin, E-selectin, and TNF-α-activated endothelium under flow in vitro. J Immunol 1998; 161: 2501–8. 3. Norman KE, Katopodis AG, Thoma G, Kolibinger F, Hicks AE, Cotter MJ et al. P-selectin glycoprotein
ligand-1 supports rolling on E- and P-selectin in vivo. Blood 2000; 96: 3585–91.
4. Van Kooyk Y, Figdor CG. Avidity regulation of integrins: the driving force in leukocyte adhesion. Curr Opin Cell Biol 2000; 12: 542–7.
5. Vestweber D, Blanks JE. Mechanisms that regulate the function of the selectins and their ligands. Physiol Rev 1999; 79: 181–213.
6. Rezzonico R, Imbert V, Chicheportiche R, Dayer JM. Ligation of CD11b and CD11c β2 integrins by antibodies or soluble CD23 induces macrophage inflammatory protein 1α (MIP-1α) and MIP-1β production in primary human monocytes through a pathway dependent on nuclear factor-κB. Blood 2001; 97: 2932–40.
7. Mitsuhata H, Shimizu R, Yokoyama MM. Suppressive effects of volatile anesthetics on cytokine release in human peripheral blood mononuclear cells. Int J Immunopharmacol 1995; 17: 529–34. 8. Moudgil GC, Gordon J, Forrest JB. Comparative effects of volatile anaesthetic agents and nitrous
oxide on human leukocyte chemotaxis in vitro. Can Anaesth Soc J 1984; 3: 631–7.
9. de Rossi LW, Horn NA, Buhre W, Gass F, Hutschenreuter G, Rossaint R. Effect of isoflurane on neutrophil selectin and β2-integrin activation in vitro. Anesth Analg 2002; 95: 583–7.
10. Collard CD, Gelman S. Pathophysiology, clinical manifestations, and prevention of ischemia- reperfusion injury. Anesthesiology 2001; 94: 1133–8.
11. Möbert J, Zahler S, Becker BF, Conzen PF. Inhibition of neutrophil activation by volatile anesthet- ics decreases adhesion to cultured human endothelial cells. Anesthesiology 1999; 90: 1372–81. 12. de Rossi LW, Horn NA, Hecker KE, Robitzsch T, Hutschenreuter G, Rossaint R. Effect of halothane
and isoflurane on binding of ADP- and TRAP-6 activated platelets to leukocytes in whole blood. Anesthesiology 2002; 96: 117–24.
13. YuR-G, Zhou J-X, Liu J. Prediction of volatile anaesthetic solubility in blood and priming fluids for extracorporeal circulation. Br J Anaesth 2001; 86: 338–44.
14. Davenpeck KL, Brummet ME, Hudson SA, Mayer RJ, Bochner BS. Activation of human leukocytes reduces surface P-selectin glycoprotein ligand-1 (PSGL-1, CD162) and adhesion to P-selectin in vitro. J Immunol 2000; 165: 2764–72.
15. Shang XZ, Issekutz AC. Contribution of CD11a/CD18, CD11b/CD18, ICAM-1 (CD54) and – 2 (CD102) to human monocyte migration through endothelium and connective tissue fibroblast barriers. Eur J Immunol 1998; 28: 1970–9.