Functional constituents of a local serotonergic system, intrinsic to the human coronary artery smooth muscle cells.
Functional constituents of a local serotonergic system, intrinsic to the human coronary artery smooth muscle cells.
复制标题
局部血清素能系统的功能成分，是人类冠状动脉平滑肌细胞固有的。
DOI：
10.1007/s11033-015-3874-x
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发表时间：
2015
影响因子：
2.8
通讯作者：
Agrawal,DevendraK
中科院分区：
文献类型：
--
作者：
Baskar,Kannan;Sur,Swastika;Selvaraj,Vithyalakshmi;Agrawal,DevendraK
人冠状动脉平滑肌细胞（HCASMCs）在冠状动脉粥样硬化和冠状动脉疾病（CAD）的发病机制中起重要作用。血清素是一种介导血管平滑肌细胞有丝分裂和冠状动脉粥样硬化的物质。我们假设HCASMC具有血清素能系统的某些功能成分，如：色氨酸羟化酶和血清素转运体。我们的目的是检测功能性色氨酸羟化酶-1 （TPH1）和血清素转运蛋白（SERT）在HCASMCs中的存在。qPCR检测TPH1 mRNA转录本，Western blot检测TPH1和SERT蛋白表达。通过DNA凝胶电泳和qPCR产物测序验证引物的特异性和准确性。使用基于荧光染料的血清素转运蛋白检测SERT的功能。采用超高效液相色谱法测定TPH的酶活性。HCASMCs同时表达SERT和TPH的mRNA转录物和蛋白。qPCR结果显示，两个转录本均为单个熔融曲线峰，序列分析显示扩增子与各自的基因一致。在HCASMCs中存在SERT和TPH酶活性。综上所述，TPH和SERT在HCASMCs中都有功能表达。这些发现是新颖的，代表了检验HCASMCs中血清素能系统的临床相关性及其在冠状动脉粥样硬化和CAD发病机制中的作用的第一步。
Human coronary artery smooth muscle cells (HCASMCs) play an important role in the pathogenesis of coronary atherosclerosis and coronary artery diseases (CAD). Serotonin is a mediator known to produce vascular smooth muscle cell mitogenesis and contribute to coronary atherosclerosis. We hypothesize that the HCASMC possesses certain functional constituents of the serotonergic system such as: tryptophan hydroxylase and serotonin transporter. Our aim was to examine the presence of functional tryptophan hydroxylase-1 (TPH1) and serotonin transporter (SERT) in HCASMCs. The mRNA transcripts by qPCR and protein expression by Western blot of TPH1 and SERT were examined. The specificity and accuracy of the primers were verified using DNA gel electrophoresis and sequencing of qPCR products. The functionality of SERT was examined using a fluorescence dye-based serotonin transporter assay. The enzymatic activity of TPH was evaluated using UPLC. The HCASMCs expressed both mRNA transcripts and protein of SERT and TPH. The qPCR showed a single melt curve peak for both transcripts and in sequence analysis the amplicons were aligned with the respective genes. SERT and TPH enzymatic activity was present in the HCASMCs. Taken together, both TPH and SERT are functionally expressed in HCASMCs. These findings are novel and represent an initial step in examining the clinical relevance of the serotonergic system in HCASMCs and its role in the pathogenesis of coronary atherosclerosis and CAD.