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)

Public Health Relevance

TO PUBLIC HEALTH Calcific aortic stenosis is a rapidly fatal disease with few therapeutic options, largely due to limited knowledge of its causes. Growing evidence pinpoints the signaling molecules, I-SMADs, as having a key role in the valvular calcification. This study will determine how I-SMADs regulate CAVD, and results may lead to new treatment modalities.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
1R01HL114709-01
Application #
8352180
Study Section
Special Emphasis Panel (ZHL1-CSR-N (M1))
Program Officer
Evans, Frank
Project Start
2012-08-23
Project End
2016-06-30
Budget Start
2012-08-23
Budget End
2013-06-30
Support Year
1
Fiscal Year
2012
Total Cost
$385,000
Indirect Cost
$135,000
Name
University of California Los Angeles
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
092530369
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
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