Serotonin Signaling in Mitral Valve Homeostasis, Maintenance and Restoration

 描述(申请人提供)：粘液瘤性二尖瓣(MV)病(MMVD)影响全球超过1.4亿患者。这项应用旨在通过研究影响MMVD病理生理的5-羟色胺(5-HT)机制来维持和恢复MMVD患者的MV稳态。5-羟色胺是一种神经递质，如果水平升高，可能会导致心脏瓣膜病。我们的研究表明，MMVD的进展是由通过5HTR受体(5HTR)2A和2B的5HT信号和启动子区域(全长启动子)的特异性5HT转运体(SERT)多态所介导的，呈孟德尔分布。因此，MMVD似乎是一种由于5HTR水平增加而导致5HTR反应性增加的疾病，并因LL-SERT基因导致SERT最高表达而加剧。5-HTSERT后下游加工导致活性氧(ROS)和转化生长因子-β-1信号的增加。具体目的1：探讨LL-SERT基因多态性与MMVD进展和手术修复后复发及5-羟色胺反应性的关系。1.a)假设：具有LL-SERT多态的患者MMVD进展更快的风险更高，需要在更年轻的年龄进行手术；此外，手术后，由于持续的5HT增强机制，患有MMVD的这种基因可能有更高的再次手术风险。我们将根据SERT多态、SERT SNP变异和影像研究，进行遗传学和生物信息学分析，以确定MVD患者的发生率、进展率和临床复发。1.b)假设：来自LL-SERT多态患者的VIC对5-羟色胺刺激更敏感，导致MVICs激活和组织重塑。从MMVD患者和对照组分离出LL、LS和SS来源的VIC和MV组织，并暴露于5HT刺激。终点将是MVICs表型、ROS产生和ECM重塑。特异性目的2：探讨转化生长因子-β-1相关信号通路在3种不同SERT基因型人MV细胞中的信号转导机制。2.a)假设：抑制人源性MV细胞和组织中5HTR2A和2B的表达控制细胞表型和细胞外基质。这个子目标将使用已建立的针对5HTR2A/2B的siRNA构建来研究它们在5HT剂量范围内的影响，终点包括细胞增殖和细胞外基质重塑。该方法还将分别研究SERT基因型SS、LL和LS。2.b)假说：与5HTR信号有关的下游转化生长因子-β-1参与了微血管病变的进展。这些研究将探讨5HTR-转化生长因子-β-1相互作用的影响，包括转化生长因子-β-1受体的表达、转化生长因子-β-1的分泌、正则和非正则信号以及5HTR活性产生的转化生长因子-β-1对ROS和细胞外基质的影响。

 DESCRIPTION (provided by applicant): Myxomatous mitral valve (MV) disease (MMVD) affects over 140 million patients worldwide. This application is concerned with maintaining and restoring MV homeostasis in MMVD patients through investigations of serotonin (5HT) mechanisms that affect MMVD pathophysiology. 5HT is a neurotransmitter, which if present at increased levels can cause valvulopathies. Our research indicates that progression of MMVD is mediated by 5HT signaling via 5HT receptors (5HTR) 2A and 2B and by a specific 5HT transporter (SERT) polymorphism in the promoter region (LL-SERT, the full-length promoter) with a Mendelian distribution. Thus, MMVD appears to be a disorder with increased 5HTR responsiveness due to augmented levels of 5HTRs, and is exacerbated by maximal expression of SERT due to the LL-SERT genotype. Downstream processing post-SERT of 5HT results in increased reactive oxygen species (ROS) and TGF-β1 signaling. Specific Aim 1: To determine the impact of LL-SERT polymorphism in MMVD progression and recurrence after surgical repair and 5HT responsiveness. 1. a) Hypothesis: Patients with LL-SERT polymorphism are at higher risk of more rapid progression of MMVD, requiring surgery at a younger age; furthermore post-surgery this genotype with MMVD may have an increased risk for re-operation due to continuing 5HT-enhanced mechanisms. We will conduct genetic and bioinformatics analyses to determine occurrence, progression rate, and clinical recurrence of MVD patients based on SERT polymorphisms, SERT SNP variations, and imaging studies. 1. b) Hypothesis: VIC derived from patients with LL-SERT polymorphism are more responsive to 5HT stimulation resulting in MVICs activation and tissue remodeling. LL-, LS-, and SS-derived VICs and MV tissues will be isolated from MMVD patients and controls and exposed to 5HT stimulation. Endpoints will be MVICs phenotype, ROS production, and ECM remodeling. Specific Aim 2: To investigate the mechanisms of 5HTR signaling in human derived MV cells from all 3 SERT genotypes, focusing on TGF-β1 related pathways. 2. a) Hypothesis: Suppression of 5HTR2A & 2B expression in human derived MV cells and tissues controls cell phenotype and ECM. This subaim will use established siRNA constructs specific for 5HTR2A/2B to investigate their effects over a 5HT dose range, with endpoints including cell proliferation and ECM remodeling. This approach will also study SERT genotypes, SS, LL, and LS separately. 2. b) Hypothesis: Downstream TGF-β1 events related to 5HTR signaling mediate the progression of MMVD. These studies will investigate the effect of 5HTR-TGF-β1 interactions including studies of TGF- β1-receptor expression, TGF-β1 excretion, canonical and non-canonical signaling and the effects of TGF-β1 arising from 5HTR activity on ROS and ECM.