ANGIOTENSIN II-SMAD SIGNALING IN CARDIAC FIBROSIS

描述（由申请人提供）：Ang II在慢性心脏病中起关键作用，然而，其导致心脏纤维化的信号通路仍不清楚。众所周知，Ang II通过刺激TGF-b介导心脏纤维化，然而，我们的初步研究发现Ang II能够通过两种机制直接激活TGF-b信号通路：1)通过激活ERK/p38 MAP激酶的急性途径（5-30分钟）。这是与TGF-b无关的，因为Ang II能够在缺乏TGF-b受体的细胞中激活Smad2和3；这种反应被ERK或p38抑制剂阻断；2)通过自分泌TGF-b导致纤维化的晚期机制（24小时）。此外，我们还发现smad缺失的小鼠可以防止心脏纤维化，而Smad2有条件缺失的小鼠则可以增强Ang II对心脏纤维化的反应。因此，我们假设Smad信号是Ang II反应中心脏纤维化发展的关键。我们计划通过追求三个具体目标来检验这一假设。在Specific Aim 1中，我们提出确定Ang II介导心脏纤维化的新信号通路。我们将证明Ang II通过AT1-R信号并通过ERK/p38 mapk依赖机制激活急性Smad信号（5-30分钟）。我们还建议通过经典的tgf -b依赖机制（24小时）激活晚期Smad信号来确定Ang II的长期影响。在Specific Aim 2中，我们将分析Smad2或SmadS在缺乏Smad2或SmadS的小鼠胚胎成纤维细胞和不表达Smad2或Smad2有条件敲除的心脏成纤维细胞中angll介导的纤维化中的具体作用。在Specific Aim 3中，我们将通过皮下注射大剂量的Ang II进一步研究Smad2或SmadS在无SmadS KO小鼠或Smad2导致条件性KO小鼠心脏纤维化中的功能作用。我们期望获得的结果将支持中心假设，为angll介导的心脏纤维化的发病机制提供新的见解，并为开发新的治疗策略提供信息。

DESCRIPTION (provided by applicant): Ang II plays a pivotal role in chronic cardiac disease, however, its signaling pathways leading to cardiac fibrosis remain largely unclear. It is known that Ang II acts by stimulating TGF-b to mediate cardiac fibrosis, however, our preliminary studies found that Ang II is able to directly activate the TGF-b signaling pathway by two mechanisms: 1) an acute pathway (5-30 minutes) via activation of the ERK/p38 MAP kinases. This is TGF-b-independent since Ang II is able to activate Smad2 &3 in cells lacking TGF-b receptors; this response is blocked by ERK or p38 inhibitors; 2) a late mechanism (24 hours) that acts through autocrine TGF-b and leads to fibrosis. Furthermore, we also found that mice null for SmadS are protected against cardiac fibrosis, while mice that are conditionally deleted for Smad2 enhance fibrosis in response to Ang II. Thus, we hypothesize that Smad signaling is a key to the development of cardiac fibrosis in response to Ang II. We plan to test this hypothesis by pursuing three specific aims. In Specific Aim 1, we propose to identify new signaling pathways whereby Ang II mediates cardiac fibrosis. We will demonstrate that Ang II signals through the AT1-R and activates an acute Smad signaling via the ERK/p38 MAPK-dependent mechanism (5-30 mins). We also propose to identify that a long-term effect of Ang II by activating a late Smad signaling via the classic TGF-b-dependent mechanism (24hrs). In Specific Aim 2, we will dissect the specific role of Smad2 or SmadS in Ang ll-mediated fibrosis in mouse embryonic fibroblasts lacking Smad2 or SmadS and in cardiac fibroblasts that do not express SmadS or have conditional knockout for Smad2. In Specific Aim 3, we will further investigate the functional role of Smad2 or SmadS in cardiac fibrosis in mice null for SmadS KO mice or have conditional KO for Smad2 by subcutaneous infusion of large doses of Ang II. We expect that the outcomes obtained will support the central hypothesis, providing new insights into the pathogenesis of Ang ll-mediated cardiac fibrosis and information for the development of new therapeutic strategies.