Adhésion à l endothélium dans la drépanocytose
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1 Adhésion à l endothélium dans la drépanocytose Jacques Elion Départment de Génétique et INSERM UMR 763 Hôpital Robert Debré, Paris et CHU de Pointe à Pitre, Les Abymes Centre de Référence Francilien pour les Syndromes Drépanocytaires et Centre de Référence des Antilles et de la Guyane CGRF Marseille 15/17 Octobre 2007
2 1980s: abnormal adhesion of SS-RBC to the endothelium Mohandas & Evans, Blood 1984 Hoover R, Rubin R, Wise G, Warren R Adhesion of normal and sickle erythrocytes to endothelial monolayer cultures. Blood 1979, 54: Hebbel RP, Yamada O, Moldow CF, Jacob HS, White JG, Eaton JW Abnormal adherence of sickle erythrocytes to cultured vascular endothelium: possible mechanism for microvascular occlusion in sickle cell disease. J Clin Invest 1980, 65: U763
3 RBC adhesion in SCD Three questions: - which vascular territory? - which red cells? - what molecular mechanism(s)? U763
4 RBC adhesion in SCD - post capillary veinules - young RBC / activated endothelium - molecular mechanism(s): Elion & Labie Hématologie stress reticulocytes express adhesion molecules normally found on erythroid precursors in the bone marrow (VLA-4 = 4β1 integrin) - VCAM-1 is expressed only on an activated endothelium - plasma proteins proinflammatory cytokines U763
5 a very needful discovery because: in SCD, the primary physiopatholgical event is insufficient to explain vasoocclusion since the delay time to HbS polymerization is longer than the time required in the circulation to reoxigenate the blood thus all mechanisms reducing blood flow may constitute a triggering factor Steinberg M.H., N Engl J Med 1999; 340: cell adhesion. vascular tone U763
6 since then the initial scheme has become notably more complex from JP Cartron SCADHESION project C Le Van Kim, PL Tharaux, J Elion hallmarks: - all the cells present in the vessel are involved - cell activation is a major contributor UMR763
7 Activated endothelial cells are major contributors to the pathophysiology of SCD Conran & Costa, Clin Biochem, 2009,
8 RBC are activable cells and activation participates to RBC adhesion to the endothelium El Nemer, Le Van Kim, Colin Blood Transfus Clin Biol Transfus Clin Biol UMR763 modifié d après MC Durpes
9 RBC are activable cells and activation participates to RBC adhesion to the endothelium RBC adhesion to the endothelium as a therapeutic target? Blocking specific interactions: antibodies, RGD peptides Vascular "lubricant": ReothRx Does hydroxycarbamide (hydroxyurea) affect cellular adhesion to the endothelium? UMR763
10 HYDROXYCARBAMIDE (HC) is the only effective drug in SCD FDA 1998 (adults) DROXIA Bristol-Myers Squibb EMEA 2007 (adults and children) SIKLOS AddMedica Initial expectation from HC: increased HbF expression decreased HbS polymerisation reduced vaso-occlusion β S β 6 Val β S β S β S β S β S deoxyhbs polymer β 85 Phe β 88 Leu β S β S β S β S γ Glu Heterotetramer 2β S γ Glu γ Glu γ HbF 2γ2 UMR763
11 HYDROXYCARBAMIDE & Sickle Cell Disease but Oral HU : proven efficacy in clinical trials (adults & children) (Charache S et al, N Engl J Med 1995) Early clinical response Delayed and highly variable Hb F increase UMR763
12 Could HC affect cell adhesion? from JP Cartron SCADHESION project C Le Van Kim, PL Tharaux, J Elion - expression of adhesion molecules on RBCs? - endothelial cells? UMR763
13 in vivo, HC reduces the expression of the 4β1 integrin (and CD36) on reticulocytes % of reticulocytes with overexpression of 4β β1 Controls (n=8) HU (n=19) VOC (n=18) NonVOC (n=23) Before HU On HU Number of 4β1/ reticulocyte U Controls (n=20) HU (n=20) VOC (n=18) Non VOC (n=23) Before HU On HU Odièvre et al, Haematologica, 2008
14 But paradoxically, in vivo, HC enhances the expression of BCAM/Lu Number of Lu / reticulocyte Reticulocytes Controls (n=20) HU (n=20) VOC (n=18) NonVOC (n=23) Number of Lu / reticulocyte Before HU On HU RBCs 2500 Number of Lu / RBC U Controls (n=20) HU (n=20) VOC (n=18) NonVOC (n=23) Number of Lu / RBC Before HU On HU Odièvre et al, Haematologica, 2007 in press
15 in vivo results are mimicked in vitro on erythroid progenitors in culture 4β1 integrin (VLA-4) Number of VLA-4 / GPA+ cell at J BCAM/Lu No HU With HU Number of Lu / GPA+ cell at J No HU With HU U763 Odièvre et al, Haematologica, 2008
16 Could HC affect cell adhesion? from JP Cartron SCADHESION project C Le Van Kim, PL Tharaux, J Elion - expression of adhesion molecules on RBCs - endothelial cells? UMR763
17 Targeted approach: HC decreases VCAM-1 expression at the surface of endothelial cells in culture (flow cytometry) mvcam-1 Mean fluorescence index /3 decrease ** Control HU Cyto HU+Cyto HC = inflammatory conditions TrHBMEC cells Incubation time : 24h Brun et al, 2003, Pharmacogenomics J. UMR_S 763
18 Could HC affect cell adhesion? from JP Cartron SCADHESION project C Le Van Kim, PL Tharaux, J Elion - expression of adhesion molecules on RBCs - endothelial cells UMR763
19 Global approach: transcriptome analysis 405 cdna Immunology hematology genes oligonucleotides 2,448 genes are modulated confirms the previous data Down-regulation: VCAM-1 (and also ET-1) and extends them to other adhesion molecules Down-regulation: TSP-1, vwf, PECAM-1 UMR 763
20 Yes, HC can affect cell adhesion from JP Cartron SCADHESION project C Le Van Kim, PL Tharaux, J Elion - expression of adhesion molecules on RBCs - effect on endothelial cells UMR763
21 Conclusions Cell adhesion to the endothelium is a major element for the initiation vasoocclusion in SCD Therapeutic target? HC decreases cellular adhesion to endothelium - proadhesive RBCs - adhesion properties of ECs UMR 763
22 Claudine Lapouméroulie Manuel Brun Marie-Hélène Odièvre Sandrine Laurance Emmanuelle Verger Paris Hôpital Robert Debré Guadeloupe Pointe-à-Pitre UMR 763 Marie-Dominique Hardy-Dessources Monique Decastel Marie-Claude Durpes Collaboration U665 Wassim El Nemer Caroline Le Van Kim Yves Colin Jean-Pierre Cartron Institut national de la santé et de la recherche médicale Physiopathology and Pharmacogenomics of Therapeutics for Sickle Cell Disease
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