Cardiac Myocycte-Role of PKG I Alpha in Aging-Dependent Cardiac Remodeling

描述（由申请人提供）：这项新的R03提案探讨了cgmp依赖性蛋白激酶I α （PKGI¿）在体内抑制衰老相关心脏重构的机制。它是基于PI实验室进行的大量新的和未发表的数据。初步研究已经探索了在PKGI¿亮氨酸拉链相互作用域（PKGI¿亮氨酸拉链突变体，或LZM，小鼠）中具有离散突变的小鼠模型的心脏重塑反应。在基线状态下，LZM小鼠出现进行性、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左室压力过载的情况下，与野生型同窝小鼠相比，LZM小鼠左室压力增加，收缩功能恶化，死亡率显著加快。这些数据揭示了PKGI在体内抑制衰老相关心脏重构中的新作用。与WT TAC心脏相比，我们进一步发现了TAC后LZM小鼠心脏中抗重构JNK通路的早期钝化，并发现PKGI¿与上游MAPKKK和JNK激活剂混合谱系激酶3 （MLK3）的新型相互作用。这些初步数据支持PKGI¿通过与MLK3相互作用激活JNK抑制衰老相关心脏重塑的机制。基于这些和其他初步数据，本应用程序提出验证PKGI¿及其在心肌细胞中的下游靶点作为治疗衰老相关心脏重塑的新靶点的中心假设。我们建议通过三个特定的目标来检验这一假设，使用一些小鼠模型。在SA1中，我们将通过检查在我们实验室创建的新的心肌细胞特异性PKGI敲除小鼠（PKGI¿cKO）的心脏结构和功能，探索PKGI¿抑制年龄依赖性LVH和体内重塑的机制。我们还将从PKGI¿cKO小鼠中分离心肌细胞（CMs），以探索PKGI¿抑制重塑的细胞机制。在SA2中，我们将探讨PKGI¿cKO小鼠对TAC诱导的左室压力过载的反应，并将测试CM特异性PKGI¿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塑作用。在SA3中，我们将探索MLK3敲除小鼠（MLK3-/-）的全身心脏重构反应，以验证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CM中PKGI¿抑制心脏重塑的具体机制。这个项目非常重要，因为它直接解决了一个在老年人中非常普遍的问题，因为它直接测试了新的治疗靶点。它是高度

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 It is based on extensive new and unpublished data performed in the PI's laboratory.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.