Role of Inhibitory SMADs in Calcific Aortic Valve Disease

抑制性 SMAD 在钙化主动脉瓣疾病中的作用

• 批准号: 8352180
• 负责人: Linda L. Demer
• 金额: $ 38.5万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-23 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8352180
• 关键词: 3-Dimensional; Adenine; Alkaline Phosphatase; Animals; Aortic Valve Stenosis; Biological; Biological Assay; Blood Vessels; Calcified; Cardiac; Cartilage; Cells; Collagen; Collagen Type X; Congestive Heart Failure; Cyclic AMP-Dependent Protein Kinases; Development; Diet; Dietary Fats; Disease; Early Diagnosis; Early treatment; Embryo; Embryonic Development; Fatty acid glycerol esters; Feedback; Fibrosis; Fluorochrome; Gel; Gene Silencing; Homeostasis; Human; Hyperlipidemia; Image; In Vitro; Inflammation; Inflammatory; Inflammatory Bowel Diseases; Knowledge; Lead; Lesion; Link; MADH6 gene; MADH7 gene; Measures; Mediating; Mediator of activation protein; Medical; Modality; Molecular; Molecular Target; Morbidity - disease rate; Mothers; Mus; Myocardial; Myocardial perfusion; Nodule; Organ Culture Techniques; Osteocalcin; Outcome; Oxygen; Pathway interactions; Patients; Perfusion; Play; Preventive; Process; Public Health; Resistance; Role; Scleroderma; Severities; Signal Transduction; Signaling Molecule; Smooth Muscle Myocytes; Symptoms; Syncope; TNF gene; Testing; Therapeutic; Therapeutic Intervention; Time; Tissues; Transforming Growth Factors; Up-Regulation; Ventricular Arrhythmia; Work; abstracting; aortic valve; aortic valve disorder; base; bone morphogenetic protein 2; calcification; cartilage cell; cell type; cytokine; feeding; in vivo; interstitial cell; member; mortality; novel; oxidant stress; oxidized lipid; receptor; response; tumorigenesis; valve replacement; ventricular hypertrophy; 3-Dimensional; Adenine; Alkaline Phosphatase; Animals; Aortic Valve Stenosis; Biological; Biological Assay; Blood Vessels; Calcified; Cardiac; Cartilage; Cells; Collagen; Collagen Type X; Congestive Heart Failure; Cyclic AMP-Dependent Protein Kinases; Development; Diet; Dietary Fats; Disease; Early Diagnosis; Early treatment; Embryo; Embryonic Development; Fatty acid glycerol esters; Feedback; Fibrosis; Fluorochrome; Gel; Gene Silencing; Homeostasis; Human; Hyperlipidemia; Image; In Vitro; Inflammation; Inflammatory; Inflammatory Bowel Diseases; Knowledge; Lead; Lesion; Link; MADH6 gene; MADH7 gene; Measures; Mediating; Mediator of activation protein; Medical; Modality; Molecular; Molecular Target; Morbidity - disease rate; Mothers; Mus; Myocardial; Myocardial perfusion; Nodule; Organ Culture Techniques; Osteocalcin; Outcome; Oxygen; Pathway interactions; Patients; Perfusion; Play; Preventive; Process; Public Health; Resistance; Role; Scleroderma; Severities; Signal Transduction; Signaling Molecule; Smooth Muscle Myocytes; Symptoms; Syncope; TNF gene; Testing; Therapeutic; Therapeutic Intervention; Time; Tissues; Transforming Growth Factors; Up-Regulation; Ventricular Arrhythmia; Work; abstracting; aortic valve; aortic valve disorder; base; bone morphogenetic protein 2; calcification; cartilage cell; cell type; cytokine; feeding; in vivo; interstitial cell; member; mortality; novel; oxidant stress; oxidized lipid; receptor; response; tumorigenesis; valve replacement; ventricular hypertrophy

DESCRIPTION (provided by applicant): Role of Inhibitory SMADs in Calcific Aortic Valve Disease Calcific aortic valve disease (CAVD) is associated epidemiologically with hyperlipidemia, the biological effects of which appear to occur through an inflammatory, positive feedback loop linking oxidized lipids, cytokines, and oxidant stress. Although it is a rapidly fatal disorder, therapeutic options are extremely limited. Its cliical severity is due to leaflet stiffening, which restricts valve opening, increasing outflow resistance and oxygen demand, while impairing cardiac perfusion and oxygen supply. Thus, a better understanding of the mechanisms underlying calcification and stiffening is critical for the development of early diagnosis and medical therapy. CAVD is increasingly acknowledged as a regulated process, involving osteochondrogenic differentiation of valvular cells. Evidence now suggests that the downstream targets of members of the transforming growth factor- (TGF-) superfamily, known as "inhibitory small mothers against decapentaplegic" (I-SMADs; SMAD6 and 7), regulate valve embryogenesis and may serve as a mitigating factor in CAVD. Their upstream effectors, BMP-2 and TGF-, are expressed in CAVD, and both induce valvular cell calcification. They signal through receptor-associated SMADs (R-SMADs) to induce osteochondrogenic factors. Although I-SMADs were originally identified as negative regulators, under some conditions, they have been shown to promote TGF- superfamily signaling. In addition, BMP-2 and TGF- may also promote leaflet stiffening, given their ability to induce aggregation and contraction of valvular interstitial cells into nodules resembling those on leaflet in CAVD. In preliminary studies, we found that 1) I-SMAD expression is upregulated in aortic valves in hyperlipidemic mice, 2) TNF- induces I-SMAD expression in aortic smooth muscle cells, and 3) protein kinase A (PKA) activation induces I-SMAD expression and inhibits cellular nodule formation in vitro. We hypothesize that I-SMADs are induced by pro-inflammatory factors and that they mitigate CAVD by negatively regulating TGF- superfamily-induced CAVD. To test this novel hypothesis, we propose three Specific Aims. In Aim 1, we will establish the mechanism of effects of pro-inflammatory factors on I-SMAD induction in CAVD. We will use in vitro (murine valvular interstitial cells; mVIC), ex vivo (murine valvular organ culture; mVOC) and in vivo (PontgLdlr-/-Apob100 and p75Tnfr-/-Ldlr-/-Apob100 mice) approaches. In Aim 2, we will test whether I-SMADs regulate CAVD in response to pro-inflammatory factors. We will employ in vitro, ex vivo, and in vivo approaches using Smad6-/- mice and their I-SMAD-deficient mVIC and mVOC. Oxidant stress and inflammation will be induced by oxidized lipids and TNF- in vitro and by an adenine diet in vivo. In Aim 3, we will test whether I-SMADs mediate inhibitory effects of PKA on formation and contraction of valvular nodules. We will use collagen-gel contraction and nodule formation assays with I- SMAD gene silencing. Results of the proposed work will harness the regulatory power of I-SMADs for development of preventive measures and medical treatments for CAVD. (End Abstract)

描述（由申请人提供）： 抑制性SMAD在钙化主动脉瓣疾病钙化主动脉瓣疾病（CAVD）中的作用在流行病学上与高脂血症有关，其生物学作用似乎是通过连接氧化的脂质，细胞因子和氧化胁迫的炎症，阳性反馈循环而发生的。尽管这是一种迅速致命的疾病，但治疗选择极为有限。它的静态严重程度是由于传单僵硬造成的，这限制了阀门的开口，增加了流出阻力 和氧气需求，同时损害心脏灌注和氧气供应。因此，更好地了解钙化和僵硬的机制对于早期诊断和医疗疗法的发展至关重要。越来越多地认为CAVD是一个受调节的过程，涉及瓣膜细胞的骨软骨分化。现在有证据表明，转化生长因子（TGF-）超家族成员的下游靶标被称为“抑制性小母亲对脱皮术的抑制小母亲”（I-SMADS; SMAD6和7），调节瓣膜胚胎发生，并可以用作CAVD中的缓解因子。它们的上游效应子BMP-2和TGF-在CAVD中表达，都诱导瓣膜细胞钙化。它们通过与受体相关的SMAD（R-SMADS）发出信号，以诱导骨软骨因子。尽管I-SMAD最初被鉴定为负调节剂，但在某些条件下，它们已被证明可以促进TGF-超家族信号传导。此外，鉴于它们能够诱导瓣膜间质细胞聚集和收缩成类似于CAVD的传单的结节，BMP-2和TGF-也可能会促进传单僵硬。在初步研究中，我们发现1）I-SMAD表达在高脂血症小鼠的主动脉瓣中上调，2）TNF- TNF-诱导主动脉平滑肌细胞中的I-SMAD表达，而3）蛋白激酶A（PKA）激活诱导I-SMAD表达并诱导细胞的表达，并抑制细胞的结节形成。我们假设I-SMAD是由促炎性因素诱导的，并且它们通过负调节TGF-超家族引起的CAVD来减轻CAVD。为了检验这个新的假设，我们提出了三个特定目标。在AIM 1中，我们将建立促炎性因素对CAVD I-SMAD诱导的影响的机理。我们将使用体外（鼠瓣膜间质细胞； MVIC），离体（鼠瓣膜器官培养； MVOC）和 invivo（pontgldlr - / - apob100和p75tnfr - / - ldlr - / - apob100小鼠）接近。在AIM 2中，我们将测试I-Smads是否对促炎性因素进行调节CAVD。我们将使用Smad6 - / - 小鼠及其I-SMAD缺乏的MVIC和MVOC使用体外，体内和体内方法。氧化应激和炎症将由氧化的脂质和TNF-体外以及体内腺嘌呤饮食诱导。在AIM 3中，我们将测试I-SMADS是否介导PKA对瓣膜结节的形成和收缩的抑制作用。我们将使用I-Smad基因沉默使用胶原凝胶收缩和结节形成测定。拟议工作的结果将利用I-SMADS的监管能力来制定CAVD的预防措施和医疗治疗。 （结尾摘要）