Cardiac Myocyte-Specific Role of PKG I Alpha in Aging-Dependent Cardiac Remodelin

描述(由申请人提供)：这份新的R03提案探索了cGMP依赖的蛋白激酶Iα(PKGI？)在体内抑制衰老相关心脏重构的机制。它是基于PI的实验室中进行的大量新的和未公布的数据。初步研究探索了在PKGI亮氨酸拉链相互作用区域(PKGI亮氨酸拉链突变体，或LZM，小鼠)中存在离散突变的小鼠模型的心脏重构反应。在基线状态下，LZM小鼠发生进行性的、与年龄相关的左心室肥厚(LVH)，这是重塑的关键组成部分。在老年患者常见的危险因素LV压力超负荷的背景下，与野生型对照相比，LZM小鼠出现了LVH增加，收缩功能恶化，死亡率显著加快。这些数据揭示了PKGI在抑制体内衰老相关的心脏重塑中的新作用。与WT TAC心脏相比，我们进一步鉴定了LZM小鼠TAC后心脏中抗重塑JNK通路的早期钝化，并鉴定了PKGI与上游MAPKKK和JNK激活物混合谱系激酶3(MLK3)的新的相互作用。这些初步数据支持一种机制，即PKGI通过与MLK3相互作用，激活JNK，以抑制与衰老相关的心脏重构。基于这些和其他初步数据，这项应用建议检验中心假设，即心肌细胞中的PKGI？及其下游靶点作为治疗衰老相关心脏重构的新靶点。我们建议通过三个特定的目标，使用一些小鼠模型来检验这一假设。在SA1中，我们将通过检测我们实验室创造的新的心肌细胞特异性PKGI？基因敲除小鼠(PKGI？CKO)的心脏结构和功能，探讨PKGI？抑制年龄依赖性左室肥厚和体内重构的机制。我们还将从PKGI？CKO小鼠中分离心肌细胞(CMS)，以探索PKGI？抑制重构的细胞机制。在SA2中，我们将探讨PKGI？CKO小鼠对TAC诱导的左心室压力超负荷的反应，并将测试CM特异性PKGIβ缺失在多大程度上消除磷酸二酯酶5抑制剂西地那非的抗重构作用。在SA3中，我们将探索全身MLK3基因敲除小鼠(MLK3-/-)的心脏重构反应，以检验PKGI相互作用蛋白MLK3在体内抑制衰老相关重构的临床相关假设。这些拟议的研究将确定心肌中的PKGI抑制心脏重构的具体机制。这一项目意义重大，因为它直接解决了老年人中普遍存在的问题，并直接测试了新的治疗目标。它是高度的 创新是因为它使用了独特的活体模型，还因为它采用了一种创新的策略，探索特定于CM的心脏重构调节因子，这可能转化为新颖的特定于CM的治疗方法。 公共卫生相关性：在老年人中，充血性心力衰竭是发病率和死亡率的主要原因，与年轻患者相比，老年患者的预后更差。心力衰竭是心脏重塑过程的结果。我们建议的项目研究心脏重塑，试图找到治疗老年心力衰竭的新策略。该项目基于大量的新数据，并建议研究一种可能在限制心脏重构过程中具有直接重要性的途径。该项目的目标是了解这一新途径(称为PKGI途径)在人体特定细胞(称为心肌细胞)中的作用。这项工作的最终希望是，通过展示PKGI在心肌细胞中的工作方式，它可能会建议利用这些细胞中的PKGI活性的新药物治疗，这与老年患者特别相关。

DESCRIPTION (provided by applicant): This new R03 proposal explores the mechanisms through which the cGMP-dependent protein kinase I alpha (PKGI¿) inhibits aging-related cardiac remodeling in vivo. It is based on extensive new and unpublished data performed in the PI's laboratory. Preliminary studies have explored the cardiac remodeling response in a mouse model harboring discrete mutations in the PKGI¿ leucine zipper interacting domain (the PKGI¿ leucine zipper mutant, or LZM, mouse). In the baseline state, the LZM mice develop progressive, age-related left ventricular hypertrophy (LVH), a key component of remodeling. In the setting of LV pressure overload, a common risk factor in elderly patients, the LZM mice develop increased LVH, worsening contractile function, and striking accelerated mortality, compared with wild type littermate controls. These data reveal a novel role of PKGI¿ in inhibiting aging-related cardiac remodeling in vivo. We have further identified early blunting of the anti-remodeling JNK pathway in hearts of LZM mice after TAC, compared with WT TAC hearts, and have identified a novel interaction of PKGI¿ with the upstream MAPKKK and JNK activator mixed lineage kinase 3 (MLK3). These preliminary data support a mechanism by which PKGI¿, via interaction with MLK3, activates JNK to inhibit aging-related cardiac remodeling. Based on these, and other, preliminary data, this application proposes to test the central hypothesis that PKGI¿ and its downstream targets in the cardiac myocyte serve as novel targets to treat aging-related cardiac remodeling. We propose to test this hypothesis through three specific aims, using a number of mouse models. In SA1 we will explore the mechanisms by which PKGI¿ inhibits age-dependent LVH and remodeling in vivo, by examining the cardiac structure and function of a new cardiac myocyte-specific PKGI¿ knockout mouse (PKGI¿cKO) created in our laboratory. We will also isolate cardiac myocytes (CMs) from PKGI¿cKO mice to explore the cellular mechanisms by which PKGI¿ inhibits remodeling. In SA2 we will explore the response of the PKGI¿cKO mice to LV pressure overload induced by TAC, and will test the degree to which CM specific PKGI¿ deletion abolishes the anti-remodeling effect of the phosphodiesterase 5 inhibitor sildenafil. In SA3 we will explore the cardiac remodeling response in a whole body MLK3 knockout mouse (MLK3-/-) to test the clinically relevant hypothesis that the PKGI¿-interacting protein MLK3 inhibits aging-related remodeling in vivo. These proposed studies will define the specific mechanisms through which PKGI¿ in the CM inhibits cardiac remodeling. This project is highly significant because it directly addresses a problem which is highly prevalent in the elderly and because it directly tests new therapeutic targets. It is highly innovative because it uses unique in vivo models, and also because it employs an innovative strategy of exploring CM-specific regulators of cardiac remodeling, which could translate into novel, CM-specific treatments. PUBLIC HEALTH RELEVANCE: In the elderly, the disease of congestive heart failure is a major cause of morbidity and mortality, and carries a worse prognosis in elderly patients compared with younger patients. Heart failure results from a process termed cardiac remodeling. Our proposed project studies cardiac remodeling to try to find new treatment strategies for heart failure in the elderly. This project is based on extensive new data and proposes to examine a pathway which may be of direct importance in limiting the cardiac remodeling process. The goals of the project are to understand the role of this new pathway (called the PKGI¿ pathway) in a specific cell of the body, called the cardiac myocyte. The ultimate promise of this work is that, by showing how PKGI¿ works in the cardiac myocyte, it might suggest new medical treatments which take advantage of PKGI¿ activity in these cells, which is particularly relevant to aging patients.