ALBERT

Description: In 1998, Gasbarrini et al reported that in ITP cases with Helicobacter pylori (H.pylori) infection, elevation of platelet counts was observed by eradication of this bacterium. Since then, several reports from Italy and Japan confirmed the elevation of platelet counts after eradication. However, the characteristic background in the H.pylori positive ITP and eradication effects on platelet counts is unclear. On the other hand, reports from Spain, North Europe and USA could not show the evidence that eradication is effective on elevating platelet counts in H.pylori positive ITP. Therefore, we designed a nationwide retrospective study in Japan to evaluate the incidence of H.pylori positive ITP cases and the effects of eradication on platelet counts and to clear above problems. Four hundred and thirty-five ITP cases were enrolled over a period of one and half years (2002. 7~2003.12) from 12 hospitals. H. pylori infection was found in 300 cases(65%), who were significantly (P

Description: Glycoprotein V (GPV) is a subunit of the platelet GPIb-IX-V receptor for von Willebrand factor (vWF) and thrombin. GPV is cleaved from the platelet surface during activation by thrombin, but its role in platelet activation is still unknown. It is reported that the GPV knockout mouse did not suffer from a bleeding syndrome, and the GPV-deficient platelets showed normal GPIb expression and vWF-mediated adhesion. One report showed an increased sensitivity of these platelets to activation by low concentrations of thrombin, suggesting that GPV may act as a negative modulator of the thrombin response. Another showed that GPV-deficient platelets exhibited defective adhesion to a collagen-coated surface, suggesting GPV binds to collagen and participates in platelet adhesion and aggregation responses to this agonist. To define the signaling pathway through GPV, we searched for proteins that bind to the GPV. We preformed a yeast two-hybrid screening in a human placenta library, using the sequence for GPV cytoplasmic domain as a bait. As a result, we isolated 2 independent positive clones, and the both clones encoded a partial and a full length sequence of the same protein, named snapin (SNAPAP), which was previously reported as one of the SNARE (soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor)-associated proteins. By RT-PCR analysis, snapin mRNA was detected in megakaryocyte cell lines and peripheral blood platelets. When the snapin was expressed as a GFP-fusin protein in K562 cells, which expressed intrinsic GPIb-IX-V complex, the protein localized at perinuclear region and cytosol. By immunoprecipitation-western blotting study, GPV was co-immunoprecipitated with GFP-snapin from the lysates of these cells by anti-GFP antibody, whereas GPV was not detected in the precipitates by a control antibody. Previous reports showed that snapin is a binding protein of SNAP-25 (synaptosome-associated protein-25) and SNAP-23, and influences the release response of neurons and chromaffin cells by enhancing the synaptotagmin binding to the SNARE complex. Therefore, the novel interaction between GPV and snapin, identified in this study, might be involved in the intracellular membrane-fusion events and the granule release reaction during the platelet activation.

Description: Megakaryocytes and functional platelets were generated in vitro from murine embryonic stem (ES) cells with the use of a coculture system with stromal cells. Two morphologically distinctive megakaryocytes were observed sequentially. Small megakaryocytes rapidly produced proplatelets on day 8 of the differentiation, and large hyperploid megakaryocytes developed after day 12, suggesting primitive and definitive megakaryopoiesis. Two waves of platelet production were consistently observed in the culture medium. A larger number of platelets was produced in the second wave; 104 ES cells produced up to 108 platelets. By transmission electron microscopy, platelets from the first wave were relatively rounder with a limited number of granules, but platelets from the second wave were discoid shaped with well-developed granules that were indistinguishable from peripheral blood platelets. ES-derived platelets were functional since they bound fibrinogen, formed aggregates, expressed P-selectin upon stimulation, and fully spread on immobilized fibrinogen. These results show the potential utility of ES-derived platelets for clinical applications. Furthermore, production of gene-transferred platelets was achieved by differentiating ES cells that were transfected with genes of interest. Overexpression of the cytoplasmic domain of integrin β3 in the ES-derived platelets prevented the activation of αIIbβ3, demonstrating that this system will facilitate functional platelet studies. (Blood. 2003;102:4044-4051)