Genetic Determinants of Vasorelaxation to beta-Agonist

描述(由申请人提供)： 血管扩张剂对β受体激动剂的钝化反应与高血压的发病机制有关。血管扩张反应也因种族而异，遗传基础尚不清楚。这项建议的总体目标是确定导致血管扩张剂对β-激动剂反应的变异性的基因变异。在具体目标1中，我们将测试以下假设：高血压患者对β-激动剂的血管扩张反应缺陷与一氧化氮减少的内皮介导的静脉扩张有关，以及编码β-激动剂靶标的基因位点的等位基因变异，如β-肾上腺素能受体(例如，β2-AR)和受体后信号转导(例如，鸟嘌呤核苷酸结合(G)蛋白Gsα和G蛋白偶联受体激酶Bark)，合成β-激动剂肾上腺素的酶(例如，苯乙醇胺N-甲基转移酶，PNMT)，而β-激动剂效应因子(例如，内皮一氧化氮合酶，eNOS)是β-激动剂血管扩张反应的基础和预测因素。单核苷酸多态(SNPs)将通过对候选基因座进行重新测序和基因分型来确定。将根据SNPs和/或单倍型(给定染色体上的SNPs阵列)比较正常血压患者和高血压患者的局部手部静脉对β-激动剂异丙肾上腺素(ISO)的反应。到目前为止的初步数据表明，高血压患者的静脉扩张反应受损，并与内皮NO产生功能障碍有关，并发现β2-AR单倍型和PNMT启动子区域的共同SNP(-353G/A)与血压调节有关。在特定的目标2中，我们将测试假设，即β2-AR的等位基因变异是淋巴细胞对β-激动剂反应的基础，以及与ISO的静脉扩张反应相关的等位基因变异在体外和体外影响基因功能。将分离淋巴细胞，并根据单个Beta2-AR SNP(例如，Arg16GIy和G1n27G1u)和/或单倍型来测量它们对ISO的反应(例如，密度、敏感性和脱敏)。我们将把重要的等位基因变异(例如，PNMT启动子上的-353G/A)与体内基因功能的生物标记物(如PNMT酶活性和血浆肾上腺素水平)相关联，并分离或产生这些遗传变异，然后评估它们对培养细胞蛋白质表达和功能的影响。这项研究应该确定遗传因素，这些因素不仅对高血压的发展起作用，而且对β-激动剂和β-拮抗剂的药效学个体间差异也有贡献。

DESCRIPTION (provided by applicant): Blunted vasodilator responses to beta agonist have been implicated in the pathogenesis of hypertension. The vasodilator responses also vary by race, and the genetic basis is not clear. The overall goal of this proposal is to determine genetic variants that underlie the variability in vasodilator response to beta-agonists. In Specific Aim 1, we will test the hypotheses that defective venodilatory response to beta-agonists in hypertension is associated with reduced endothelium-mediated venodilation by nitric oxide, and that allelic variants at gene loci encoding the targets of beta-agonists such as beta-adrenergic receptors (e.g., beta2-AR) and post-receptor signaling (e.g., guanine-nucleotide-binding (G) protein Gs alpha and G-protein-coupled receptor kinases BARK), the enzymes for the synthesis of the beta-agonist epinephrine (e.g., Phenylethanolamine N-methyltransferase, PNMT), and of the beta-agonist effector (e.g., endothelial nitric oxide synthesis, eNOS) underlie and predict the venodilatory response to beta-agonists. Single nucleotide polymorphisms (SNPs) will be determined by resequencing and genotyping the candidate gene loci. The local hand venous response to beta-agonist isoproterenol (ISO) will be compared between normotensives and hypertensives and according to SNPs and/or haplotypes (the array of SNPs at a given chromosome). The preliminary data to date demonstrated that the venodilatory response in hypertension was impaired and related to dysfunction in endothelial NO production, and found that Beta2-AR haplotypes and a common SNP at PNMT promoter region (-353G/A) were associated with blood pressure regulation. In Specific Aim 2, we will test the hypotheses that allelic variants at beta2-AR underlie the lymphocyte response to beta-agonists, and that allelic variant associated with venodilatory response to ISO affect gene function ex vivo and in vitro. Lymphocytes will be isolated and their responses to ISO (e.g., density, sensitivity, and desensitization) will be measured according to individual Beta2-AR SNPs (e.g., Arg16GIy and G1n27G1u) and/or haplotypes. We will correlate the significant allelic variants (e.g., -353 G/A at PNMT promoter) with the biomarkers for the function of genes in vivo such as PNMT enzymatic activity and plasma epinephrine levels, and will isolate or generate these genetic variants and then assess their impact on protein expression and function in cultured cells. This study should define the genetic factors that contribute not only to the development of hypertension but also to the inter-individual variations in pharmacodynamics for both beta-agonists and beta-antagonists.