Haematologica 2004;89: supplement no. 6 - Supplements ...

VIII Congress of the Italian Society of Experimental Hematology, Pavia, September 14-16, 200429remained uPAR-negative. Western blot analysis witha polyclonal anti-uPAR antibody confirmed a progressiveincrease of uPAR expression in all donorsduring G-CSF stimulation and showed that PBMNCexpressed only the intact form of uPAR. G-CSF-treatmentalso induced increased serum levels of solubleuPAR (suPAR). In almost all cases, cell surface uPARexpression on CD33 + and CD14 + cells and serumsuPAR levels increased to the maximum extent atdays 3-5 of G-CSF stimulations, when CD34 + HSCalso peaked into the circulation. Western blot analysisshowed that after G-CSF treatment there was notonly increase of the intact form of suPAR, but alsoappearance or strong increase of cleaved forms ofsuPAR (c-suPAR) in all analyzed sera. c-suPAR wasable to chemoattract CD34 + KG1 leukemia cells andbone marrow (BM) CD34 + HSC, as documented by invitro migratory response of these cells toward achemotactic suPAR-derived peptide (uPAR84-95).uPAR84-95 induced CD34 + KG1 and BM CD34 + HSCmigration by activating the high-affinity formylmethylpeptide (fMLP) receptor (FPR). In addition,uPAR84-95 inhibited CD34 + KG1 and CD34 + HSCmigration toward the stromal derived factor 1(SDF1), thus suggesting a heterologous desensitationof its receptor CXCR4.Finally, we studied theeffect of uPAR84-95 on the output of clonogenicprogenitors from long-term culture (LTC) of highlypurified BM CD34 + cells from normal donors. Nonadherentcells, weekly removed after a 2 h treatmentwith uPAR84-95, yielded a significant higher numberof clonogenic progenitors as compared to thoseobtained from non-adherent cells of LTC treated withthe scrambled version of uPAR84-95.All together,our data document that G-CSF-induced up-regulationof uPAR on circulating CD33 + and CD14 + cells isassociated with increased suPAR shedding, whichleads to the appearance of serum c-suPAR. c-suPARcould contribute to HSC mobilization by promotingtheir FPR-mediated migration and by inducingCXCR4 desensitation. Our findings suggest a potentialutility of the cleaved form of suPAR, or its derivedchemotactic peptide, in the strategies to optimizeHSC mobilization, especially in G-CSF poor mobilizers.Oral CommunicationsNON-MALIGNANT HEMATOLOGYCO-17AN ASSOCIATION OF PLATELET GLYCOPROTEIN GENE HAPLO-TYPES AND BLEEDING SEVERITY IN PATIENTS WITH PREVIOUS-LY DIAGNOSED VON WILLEBRAND DISEASE (VWD) TYPE 1:RESULTS OF A PILOT STUDY IN 14 ITALIAN FAMILIESFederici AB,* Baronciani L,* Canciani MT,* Cozzi G,*Mistretta C,* Gianniello F,* Peake IR,° Salomon DR,^Kunicki TJ^*Angelo Bianchi Bonomi Hemophilia and ThrombosisCenter, IRCCS Maggiore Hospital and Universityof Milan; °Division of Genomic Medicine, Royal HallamshireHospital, Sheffield, UK, ^The ScrippsResearch Institute, La Jolla, CA, USAVon Willebrand Disease (VWD) type 1 is difficult todiagnose because of bleeding variability and lowheritability of Von Willebrand Factor (VWF) levels.Secondary gene effects that increase risk for bleedingmay modify the phenotype, and platelet adhesionreceptors are prime candidates. We compared a bleedingseverity score and bleeding times to candidategene haplotypes within pedigrees of fourteen indexcases of previously diagnosed VWD type 1, using avariance component model. The 14 families fromMilan have been already enrolled the European studyentitled Molecular and Clinical Markers for Diagnosisand Management of Type 1 von Willebrand Disease(Scientific coordinator: I. R. Peake). VWD type 1patients were classified according to the previousdefinitions of the Scientific Standardization Committee(SSC) on VWF of the International Society onThrombosis and Haemostasis (ISTH) such as individualscharacterized by reduced levels of normal VWFand positive personal-family bleeding history. A bleedinghistory was derived from detailed questionnairesadministered to all the affected and non-affectedmembers (including index cases), and the severityof bleeding was ranked from 0 to 3 in each of 11bleeding categories. Donors were genotyped usingprimer extension method, and nine candidate geneswere selected for this analysis. With respect to bleedingseverity score, ITGA2 haplotype 2 (807C), GP6haplotype b (Pro219) and ITGA2B haplotype 1 (Baka)were found to be associated with increased bleeding(p=0.001, 0.05, and 0.002, respectively). No haplotypeswere associated with bleeding times, and noassociation was observed with six other candidategenes, GP1BA, ITGB3, VWF, FGB, IL6 or TXA2R. Asso-haematologica vol. 89[suppl. n. 6]:september 2004