Leoncini Giuliana, Bruzzese Debora, Signorello Maria Grazia
Department of Experimental Medicine, Biochemistry Section, University of Genoa, Viale Benedetto XV 1, 16132 Genova, Italy.
Biochem Pharmacol. 2005 Jan 15;69(2):289-96. doi: 10.1016/j.bcp.2004.09.022. Epub 2004 Nov 19.
Nitric oxide production, L-arginine transport and intracellular [Ca2+] changes in human platelets stimulated without stirring by low doses of collagen have been evaluated. Collagen decreased in a dose-dependent manner the nitric oxide formation. A reduction of about 30% of the basal level was produced by 5 microg/mL. Aspirin did not change the collagen effect. The inhibition was reversed by EGTA. Moreover collagen reduced L-arginine uptake. The exposure of platelets to 5 microg/mL collagen diminished of about 30% L-arginine transport. The specific involvement of the system y+ is suggested. In addition in FURA 2-loaded platelets collagen induced a dose-dependent slow sustained [Ca2+] rise that was almost completely cancelled by EGTA. Finally the treatment of whole platelets with collagen affected in a dose-dependent manner the maximal nitric oxide formation, suggesting a direct effect at the level of nitric oxide synthase enzyme. The phosphorylation of specific serine/threonine residues regulated by protein kinase C could be involved. In conclusion during the early phases of platelet stimulation with collagen nitric oxide formation is diminished. This reduction can be due to a lower availability of L-arginine for cytosolic nitric oxide synthase and/or to a decreased activity related to modifications of the enzyme.
已评估了低剂量胶原蛋白在不搅拌情况下刺激人血小板时一氧化氮的产生、L-精氨酸转运及细胞内[Ca2+]变化。胶原蛋白以剂量依赖方式降低一氧化氮的生成。5微克/毫升可使基础水平降低约30%。阿司匹林不改变胶原蛋白的作用。EGTA可逆转这种抑制作用。此外，胶原蛋白减少L-精氨酸摄取。血小板暴露于5微克/毫升胶原蛋白时，L-精氨酸转运减少约30%。提示系统y+有特定参与。另外，在负载FURA 2的血小板中，胶原蛋白诱导剂量依赖性的缓慢持续[Ca2+]升高，几乎完全被EGTA消除。最后，用胶原蛋白处理全血小板以剂量依赖方式影响最大一氧化氮生成，提示在一氧化氮合酶水平有直接作用。可能涉及蛋白激酶C调节的特定丝氨酸/苏氨酸残基的磷酸化。总之，在胶原蛋白刺激血小板的早期阶段，一氧化氮生成减少。这种减少可能是由于细胞溶质一氧化氮合酶的L-精氨酸可用性降低和/或与酶修饰相关的活性降低。
