镰状细胞贫血患者在其精氨酸新陈代谢途径中血小板转录体暴露的变化增强表达的特征

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Amplified Expression Profiling of Platelet Transcriptome Reveals Changes in Arginine Metabolic Pathways in Patients With Sickle Cell Disease
Background— In sickle cell disease, ischemia-reperfusion injury and intravascular hemolysis produce endothelial dysfunction and vasculopathy characterized by reduced nitric oxide and arginine bioavailability. Recent functional studies of platelets in patients with sickle cell disease reveal a basally activated state, which suggests that pathological platelet activation may contribute to sickle cell disease vasculopathy.
Methods and Results— Studies were therefore undertaken to examine transcriptional signaling pathways in platelets that may be dysregulated in sickle cell disease. We demonstrate and validate in the present study the feasibility of comparative platelet transcriptome studies on clinical samples from single donors by the application of RNA amplification followed by microarray-based analysis of 54 000 probe sets. Data mining an existing microarray database, we identified 220 highly abundant genes in platelets and a subset of 72 relatively platelet-specific genes, defined by >10-fold increased expression compared with the median of other cell types in the database with amplified transcripts. The highly abundant platelet transcripts found in the present study included 82% or 70% of platelet-abundant genes identified in 2 previous gene expression studies on nonamplified mRNA from pooled or apheresis samples, respectively. On comparing the platelet gene expression profiles in 18 patients with sickle cell disease in steady state to those of 12 black control subjects, at a 3-fold cutoff and 5% false-discovery rate, we identified 100 differentially expressed genes, including multiple genes involved in arginine metabolism and redox homeostasis. Further characterization of these pathways with real-time polymerase chain reaction and biochemical assays revealed increased arginase II expression and activity and decreased platelet polyamine levels.
Conclusions— The present studies suggest a potential pathogenic role for platelet arginase and altered arginine and polyamine metabolism in sickle cell disease and provide a novel framework for the study of disease-specific platelet biology.
镰状细胞贫血患者在其精氨酸新陈代谢途径中血小板转录体暴露的变化增强表达的特征
背景--在镰状细胞贫血病，局部缺血、再灌注损伤和血管内溶血导致了以一氧化氮和精氨酸生物有效性减低为特征的内皮细胞损伤和血管病变。在有镰状细胞贫血病的病人中最新功能研究显露血小板处于基本被激活的状态，考虑病理性血小板活化作用也许能产生镰状细胞贫血病的血管病变。
方法和结果---接下来的研究检测血小板在镰状细胞贫血病的转录信号途径也许是不规则的。 我们在本研究中展示并且确认从唯一自愿者临床样品由RNA放大作用应用对54 000个探针集合的基于微阵列的分析以比较血小板转录体研究可行性。数据采集有一个现有的微阵列数据库，我们在血小板上辨认了220个高度丰富的基因，并且72个相对地血小板特异基因的一个子集，定义为在数据库增加了>10倍表示比较其他细胞类型中点有被增强。 在本研究中发现的高度丰富的血小板纪录包括82%或70%在2项早先基因表达研究中辨认出的血小板丰富的基因，分别从合并的或分离的样品中研究未增强的 mRNA而来。在比较18例镰状细胞贫血病患者血小板基因表达外形特征在12个黑控制主题比较稳定，有3倍切除和5%错误发现率。我们辨认了100个不同表达的基因，包括在精氨酸新陈代谢途径和氧化还原作用中介入的多个基因。这些途径特性以实时聚合酶链式反应和生物化学的分析的进一步描述显露了增加的精氨酸II表现和活动并且减少了血小板多聚水平。
结论--本研究建议在镰状细胞贫血病血小板和精氨酸为一个潜在的致病性作用，改变精氨酸和多胺的新陈代谢为特异性血小板生物学疾病的研究提供一个新颖的框架。