Calpain Activates Intracellular TGF-beta1 in Pulmonary Hypertension

描述(申请人提供)：肺动脉高压(PAH)是一种严重的进展性疾病，其主要特征是肺血管重塑，与细胞外基质(包括胶原)的积聚和血管细胞的增殖、中膜和外膜增厚、闭塞性新生内膜和复杂的丛状病变有关，导致毛细血管前动脉闭塞和肺动脉压持续升高。血小板衍生生长因子、表皮生长因子、血管内皮生长因子和转化生长因子β1参与了肺血管重塑过程。我们最近首次报道，在缺氧性肺动脉高压中，Calain的全局敲除可以防止肺血管重构，并且Calain抑制剂MDL28170可以防止野百合碱诱导的已建立的肺动脉高压的进展。钙蛋白酶是一类钙依赖的非溶酶体中性半胱氨酸内切酶家族，在包括肺血管细胞在内的哺乳动物细胞中通过对底物蛋白进行有限的蛋白分解来发挥作用。Calain是PDGF、EGF和VEGF的下游信号转导分子，介导PDGF和EGF诱导的肺动脉平滑肌细胞(PASMCs)的胶原合成和增殖，以及VEGF诱导的肺动脉内皮细胞(PAECs)的血管生成。我们最近发现，Calain裂解并激活了转化生长因子β1，而细胞内的转化生长因子β1。PASMCs中存在信号通路。为了在动物模型中验证Calain在肺血管重构中的作用，我们将利用肺高压突破计划(PHBI)提供的人类肺组织，并验证我们的新假设，即Calain在与IPAH相关的肺血管重构中激活细胞内转化生长因子-1。为了验证这一假说，我们提出了一种多学科的方法，集中于以下特定的目的：#1是确定Calain抑制是否阻止或减弱特发性肺动脉高压(IPAH)患者PASMCs和PAECs的胶原合成和增殖；#2是确定Calain是否激活IPAH患者PASMCs和PAECs的细胞内转化生长因子？1。Calain对肺血管细胞胶原合成和增殖调控的证据可能会导致新的药理策略的发展，该药物将针对与肺动脉高压相关的肺血管重构中的Calain。 公共卫生相关性：我们最近报道，在缺氧性肺动脉高压中，全局Calain基因敲除可防止肺血管重构，Calain抑制剂MDL28170可防止野百合碱诱导的已建立的肺动脉高压的进展。这项建议是为了研究一种新的假说，即在特发性肺动脉高压中，Calain激活细胞内的转化生长因子？1。这项拟议中的研究对肺动脉高压领域的贡献有望导致新的药理策略的开发，这些策略将针对与肺动脉高压相关的肺血管重构中的钙蛋白酶。该项目的顺利完成将为肺动脉高压的防治提供一种新的方法。

DESCRIPTION (provided by applicant): Pulmonary arterial hypertension (PAH) is a severe and progressive disease, a key feature of which is pulmonary vascular remodeling that is associated with accumulation of extracellular matrix, including collagen, and vascular cell proliferation, medial and adventitial thickening, occlusive neointima, and complex plexiform lesions, leading to obliteration of precapillary pulmonary arteries and sustained elevation of pulmonary arterial pressure. PDGF, EGF, VEGF, and TGF?1 participate in the process of pulmonary vascular remodeling. We recently reported for the first time that global knockout of calpain prevents pulmonary vascular remodeling in hypoxia-induced pulmonary hypertension and that the calpain inhibitor MDL28170 prevents the progression of established pulmonary hypertension induced by monocrotaline. Calpain is a family of calcium-dependent non-lysosomal neutral cysteine endopeptidases that act via limited proteolysis of substrate proteins in mammalian cells including pulmonary vascular cells. Calpain is the downstream signal transduction molecule of PDGF, EGF and VEGF, and mediates PDGF- and EGF-induced collagen synthesis and proliferation of pulmonary artery smooth muscle cells (PASMCs) and VEGF-induced angiogenesis of pulmonary artery endothelial cells (PAECs). We have recently found that calpain cleaves and activates TGF?1 and that an intracrine TGF? signal pathway exists in PASMCs. To validate the role of calpain in pulmonary vascular remodeling in animal models, we will take advantage of the human lung tissues provided by the Pulmonary Hypertension Breakthrough Initiative (PHBI) and test our novel hypothesis that calpain activates intracellular TGF?1 in pulmonary vascular remodeling associated with IPAH. To test this hypothesis, we propose a multidisciplinary approach focusing on the following specific aims: #1 is to determine whether calpain inhibition prevents or attenuates collagen synthesis and proliferation of PASMCs and PAECs from patients with idiopathic pulmonary arterial hypertension (IPAH); #2 is to determine whether calpain activates intracellular TGF?1 in PASMCs and PAECs from patients with IPAH. Proof of regulation of collagen synthesis and proliferation by calpain in pulmonary vascular cells may lead to development of new pharmacologic strategies that will target calpain in pulmonary vascular remodeling associated with pulmonary hypertension. PUBLIC HEALTH RELEVANCE: We recently reported that global calpain knockout prevents pulmonary vascular remodeling in hypoxia- induced pulmonary hypertension and the calpain inhibitor MDL28170 prevents the progression of established pulmonary hypertension induced by monocrotaline. This proposal is to study a novel hypothesis that calpain activates intracellular TGF?1 in idiopathic pulmonary arterial hypertension. The contribution of the proposed research to the field of pulmonary hypertension is expected to be leading to development of new pharmacologic strategies that will target calpain in pulmonary vascular remodeling associated with pulmonary hypertension. Successful completion of this project will help provide a new method for the prevention and treatment of pulmonary hypertension.