Role of Glutathione S-Transferase P in Heart Failure

描述（由申请人提供）：尽管氧化应激是心力衰竭（HF）的标志，但针对第一代活性氧（ROS）（如O2和H2 O2）的抗氧化剂的临床试验尚未产生令人信服的益处。然而，ROS也产生来自脂质过氧化的次级中间体，包括放大氧化损伤的过氧化物和醛。谷胱甘肽S-转移酶（GST）通过催化醛与谷胱甘肽（GSH）的结合来代谢醛。选择GST同种型也有重要的非催化功能，如与c-Jun N-末端激酶（JNK）的物理相互作用，这是由氧化应激调节。虽然GST在氧化应激反应中起着至关重要的作用，但GST如何影响HF尚不清楚。我们的目标是确定GSTP，最丰富的心脏GST亚型，在HF中的功能作用。我们的初步研究表明GSTP在HF中下调，并且GSTP缺乏会导致心脏重塑，增加蛋白醛加合物，抑制循环内皮祖细胞（EPCs），并损害新血管形成。因此，我们的中心假设是GSTP是梗死后HF中一种重要的心脏保护蛋白，可改善重塑并促进心脏修复。为了验证这个假设，我们将执行三个特定目标。在目的1中，我们将通过检测野生型（WT）、GSTP-/-和心脏特异性hGSTP 1 *A和hGSTP 1 *C转基因（Tg）小鼠的梗死后LV重构来确定GSTP以及人GSTP变体hGSTP 1 *A和hGSTP 1 *C在HF中的作用。我们将评估细胞凋亡，纤维化，炎症以及谷胱甘肽水平，蛋白加合醛，JNK激活在心脏。在目标2中，我们将确定GSTP对衰竭心脏中脂质过氧化产物解毒的代谢贡献。在WT、GSTP-/-和hGSTP Tg小鼠的离体、灌注假手术和衰竭心脏中，使用同位素标记和质谱法，我们将表征不饱和醛的代谢和解毒。在组织匀浆中，我们还将测定GST相关的过氧化物酶活性以及醛类和脂质过氧化物的水平。在目标3中，我们将描述心脏和骨髓（BM）相关的影响，其中GTSP调节新生血管在衰竭的心脏。我们将首先确定WT和GSTP -/-小鼠中的EPC和BM祖细胞功能，包括伴随和不伴随JNK抑制。接下来，我们将定义GSTP消融和hGSTP过表达如何影响Aim 1的假手术和衰竭心脏中的新血管形成和血管生成基因表达。最后，我们将评估在嵌合小鼠中的梗死后重塑、炎症、EPC动员和新血管形成：具有GSTP-/- BM的WT小鼠和具有WT BM的GSTP-/-小鼠。这些研究将确定心肌局部与BM局部GSTP在衰竭心脏重塑、新血管形成和炎症过程中的作用。总的来说，这项工作将建立一个新的范式GSTP作为一个重要的抗氧化剂，抗炎，促血管生成蛋白在HF。该模型对于HF患者的氧化损伤具有重要的诊断和治疗意义。 公共卫生相关性：这些研究将确定谷胱甘肽S-转移酶P（GSTP）作为心力衰竭中的关键抗氧化剂和组织修复蛋白，并确定氧化损伤的新决定因素。因此，这些结果可以帮助设计新的，非经典的抗氧化剂和再生疗法治疗心力衰竭。我们还将评估人GSTP变体在衰竭心脏中的心脏保护效力; GSTP变体之间的重要差异可以为心脏中抗氧化反应的变异性建立遗传基础，并为心力衰竭的进展提供新的生物标志物。

DESCRIPTION (provided by applicant): Although oxidative stress is a hallmark of heart failure (HF), clinical trials with antioxidants targeting first generation reactive oxygen species (ROS) such as O2/ and H2O2 have not yielded compelling benefits. However, ROS also generate secondary intermediates derived from lipid peroxidation, including peroxides and aldehydes that amplify oxidative injury. Glutathione S-transferases (GSTs) metabolize aldehydes by catalyzing their conjugation with glutathione (GSH). Select GST isoforms also have important non-catalytic functions such as physical interactions with c-Jun N-terminal kinase (JNK) that are modulated by oxidative stress. Although GSTs play a vital role in oxidative stress responses, how GSTs impact HF is unknown. Our goal is to define the functional role of GSTP, most abundant cardiac GST isoform, in HF. Our preliminary studies indicate that GSTP is downregulated in HF, and that GSTP deficiency worsens cardiac remodeling, augments protein- aldehyde adducts, depresses circulating endothelial progenitor cells (EPCs), and impairs neovascularization. Our central hypothesis, therefore, is that GSTP is a critical cardioprotective protein in post-infarction HF that ameliorates remodeling and promotes cardiac repair. To test this hypothesis, we will perform three Specific Aims. In Aim 1, we will define the role of GSTP, and the human GSTP variants hGSTP1*A and hGSTP1*C, in HF by examining post-infarction LV remodeling in wild-type (WT), GSTP-/-, and cardiac-specific hGSTP1*A and hGSTP1*C transgenic (Tg) mice. We will evaluate apoptosis, fibrosis, and inflammation together with glutathione levels, protein-adducted aldehydes, and JNK activation in the heart. In Aim 2, we will determine the metabolic contribution of GSTP to the detoxification of lipid peroxidation products in the failing heart. In isolated, perfused sham-operated and failing hearts from WT, GSTP-/- and hGSTP Tg mice, using isotope labeling and mass spectrometry, we will characterize the metabolism and detoxification of unsaturated aldehydes. In tissue homogenates, we will also determine GSTP-related peroxidase activity and levels of aldehydes and lipid peroxides. In Aim 3, we will delineate the cardiac and bone marrow (BM)-related effects by which GTSP modulates neovascularization in the failing heart. We will first determine EPC and BM progenitor cell function in WT and GSTP -/- mice, both with and without concomitant JNK inhibition. Next, we will define how GSTP ablation and hGSTP overexpression affect neovascularization and angiogenic gene expression in the sham and failing hearts from Aim 1. Lastly, we will evaluate post-infarction remodeling, inflammation, EPC mobilization, and neovascularization in chimeric mice: WT mice with GSTP-/- BM and GSTP-/- mice with WT BM. These studies will establish the role of myocardium-localized versus BM-localized GSTP in the process of remodeling, neovascularization, and inflammation in the failing heart. Collectively, this work will establish a novel paradigm of GSTP as an essential antioxidant, anti-inflammatory, and pro-angiogenic protein in HF. This model can have important diagnostic and therapeutic implications for HF patients with regard to oxidant injury. PUBLIC HEALTH RELEVANCE: These studies will establish glutathione S-transferase P (GSTP) as a critical antioxidant and tissue reparative protein in heart failure and identify new determinants of oxidative injury. Hence, the results can help design new, non-classical antioxidant and regenerative therapies in heart failure. We will also evaluate the cardioprotective potency of human GSTP variants in the failing heart; important differences between GSTP variants can establish a genetic basis for variability of antioxidant responses in the heart and provide a novel biomarker for the progression of heart failure.