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: 8535813
• 负责人: Linda L. Demer
• 金额: $ 36.65万
• 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/8535813
• 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

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 在钙化性主动脉瓣疾病中的作用 钙化性主动脉瓣疾病 (CAVD) 在流行病学上与高脂血症相关，其生物学效应似乎是通过连接氧化脂质、细胞因子和氧化应激的炎症、正反馈回路发生的。尽管它是一种迅速致命的疾病，但治疗选择极其有限。其临床严重程度是由于小叶僵硬，限制了瓣膜的打开，增加了流出阻力 和需氧量，同时损害心脏灌注和氧气供应。因此，更好地了解钙化和硬化的机制对于早期诊断和药物治疗的发展至关重要。 CAVD 越来越被认为是一个受调节的过程，涉及瓣膜细胞的骨软骨分化。现在的证据表明，转化生长因子 (TGF-) 超家族成员的下游靶标（称为“针对十肢瘫痪的抑制性小母体”（I-SMAD；SMAD6 和 7））调节瓣膜胚胎发生，并可能作为缓解因子在 CAVD 中。它们的上游效应子 BMP-2 和 TGF-在 CAVD 中表达，并且均诱导瓣膜细胞钙化。它们通过受体相关 SMAD（R-SMAD）发出信号来诱导骨软骨形成因子。尽管 I-SMAD 最初被认为是负调节因子，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，BMP-2 和 TGF-β 也可能促进小叶硬化，因为它们能够诱导瓣膜间质细胞聚集和收缩，形成类似于 CAVD 中小叶上的结节。在初步研究中，我们发现1）高脂血症小鼠主动脉瓣中I-SMAD表达上调，2）TNF-诱导主动脉平滑肌细胞中I-SMAD表达，3）蛋白激酶A（PKA）激活诱导I- SMAD 表达并抑制体外细胞结节形成。我们假设 I-SMAD 是由促炎因子诱导的，并且它们通过负调节 TGF 超家族诱导的 CAVD 来减轻 CAVD。为了检验这一新颖的假设，我们提出了三个具体目标。在目标 1 中，我们将建立促炎因子对 CAVD 中 I-SMAD 诱导的影响机制。我们将使用体外（小鼠瓣膜间质细胞；mVIC）、离体（小鼠瓣膜器官培养；mVOC）和 体内（PontgLdlr-/-Apob100 和 p75Tnfr-/-Ldlr-/-Apob100 小鼠）方法。在目标 2 中，我们将测试 I-SMAD 是否调节 CAVD 以响应促炎因子。我们将使用 Smad6-/- 小鼠及其 I-SMAD 缺陷型 mVIC 和 mVOC 采用体外、离体和体内方法。氧化应激和炎症将在体外由氧化脂质和TNF-α诱导，而在体内则由腺嘌呤饮食诱导。在目标 3 中，我们将测试 I-SMAD 是否介导 PKA 对瓣膜结节形成和收缩的抑制作用。我们将使用 I-SMAD 基因沉默的胶原蛋白凝胶收缩和结节形成测定。拟议工作的结果将利用 I-SMAD 的监管权力来制定 CAVD 的预防措施和医疗方法。 （摘要结束）