BIN1 is a mediator and marker of cardiac reserve in heart failure.

项目摘要 500多万美国人患有心力衰竭，每年新增50多万例 在日益严重的流行病中。然而，心力衰竭进展的病理生理学机制仍然知之甚少。 因此，我们做出治疗决定的能力仍然有限。制定更具体和更具体的 需要与心力衰竭进展直接相关的预后生物标记物来改善临床治疗。我有过 最近发现，一种关键的支架蛋白BIN1在钙处理改变的发病机制中起重要作用 在人类HF中。有了这些知识，我的长期目标是开发一种新的心脏储备的HF生物标志物 通过了解BIN1的生物学和钙在正常和病变心肌细胞中的处理。这个 这种特殊应用的目的是了解BIN1在调节钙瞬变中的作用。 并使用BIN1开发心力衰竭患者的诊断和预后测试。我的中心假设 BIN1在衰竭的人心肌细胞中的表达减少，组织和血清中的BIN1水平 与心肌组织的恢复潜能相关。其理由是成功地完成了 拟议的研究将填补基于BIN1的心衰钙瞬变调控知识的空白 有助于开发心衰患者心脏储备的创新诊断和预后测试。具体目标#1 是确定BIN1在Cav1.2转运中的作用以及在失败的人类中的钙瞬变调节 心肌细胞。定量逆转录聚合酶链式反应、免疫印迹、酶联免疫吸附试验、免疫荧光和T小管分离 用来评估BIN1和Cav1.2在失败和非失败人类中的细胞表达和定位 心肌细胞。在成年小鼠心肌细胞模型中，表面生物素化和活细胞钙成像将 用于研究Cav1.2在BIN1耗竭心肌细胞中的转运和钙瞬变。具体目标2是 心肌组织BIN1表达作为心力衰竭终末期进展的诊断和预后试验 人类心肌病患者。定量免疫荧光将用于心脏活检，以确定 心脏BIN1表达水平与临床收缩参数和预后数据的分析。特定的 目的#3确定血清BIN1是预测心肌病心脏储备功能的新的心力衰竭生物标志物 病人。心脏特异性酶联免疫吸附试验将在大范围心肌病人群中检测血清BIN1 来自加州大学旧金山分校心力衰竭诊所的患者和年龄匹配的正常对照组人群进行分析 临床数据。这项贡献有望将BIN1确定为心力衰竭的介体和生物标志物，以帮助追踪 心力衰竭的进展和预测临床结果。这一贡献将是重大的，因为 这一发现将改变目前用于监测疾病进展的心肌健康评估的范式 并指导心力衰竭的治疗策略。本申请中提出的研究具有创新性。 因为它专注于一种新的心力衰竭诊断和预后测试，组织和血清BIN1水平可以评估 心脏储备通过直接反映单个心肌细胞的生化健康状况。

Project Summary Over five million Americans have heart failure (HF), with more than 500,000 new cases added each year in what is a growing epidemic. Yet the pathophysiology of heart failure progression remains poorly understood and, as a result, our ability to make therapeutic decisions remains limited. Development of more specific and prognostic biomarkers that are directly tied to HF progression is required to improve clinical management. I have recently found that a key scaffolding protein, BIN1, is important to the pathogenesis of altered calcium handling in human HF. With this knowledge, my long term goal is to develop a novel HF biomarker of cardiac reserve through understanding the biology of BIN1 and calcium handling in normal and diseased cardiomyocytes. The objective of this particular application is to understand the role of BIN1 in regulation of the calcium transients in heart failure, and to use BIN1 to develop a diagnostic and prognostic test in HF patients. My central hypothesis is that BIN1 expression is reduced in failing human cardiomyocytes, and that BIN1 levels in tissue and serum correlate with recovery potential of myocardial tissue. The rationale is that successful completion of the proposed research will fill a gap in the knowledge of BIN1 based regulation of calcium transients in HF which helps to develop an innovative diagnostic and prognostic test of cardiac reserve in HF patients. Specific Aim #1 is to identify the role of BIN1 in Cav1.2 trafficking and calcium transient regulation in failing human cardiomyocytes. Quantitative rtPCR, western blot, ELISA, immunofluorescence, and T-tubule fractionation will be used to assess the cellular expression and localization of BIN1 and Cav1.2 in failing and non-failing human cardiomyocytes. In adult mouse cardiomyocyte models, surface biotinylation and live-cell calcium imaging will be used to study Cav1.2 trafficking and calcium transients in BIN1 depleted cardiomyocytes. Specific Aim #2 is to identify myocardial tissue BIN1 expression as a diagnostic and prognostic test of HF progression in end-stage human cardiomyopathy patients. Quantitative immunofluorescence will be used on heart biopsies to determine ventricular BIN1 expression level for analysis with clinical contractile parameters and outcome data. Specific Aim #3 is to identify serum BIN1 as a novel prognostic HF biomarker of cardiac reserve in cardiomyopathy patients. Serum BIN1 will be measured by cardiac specific ELISA in a large population of cardiomyopathy patients from the UCSF heart failure clinic and an age-matched normal control population for analysis with clinical data. The contribution is expected to identify BIN1 as a HF mediator and biomarker to help track the progression of heart failure and prognosticate clinical outcomes. This contribution will be significant because such discovery will shift current paradigm in myocardial health assessment for monitoring disease progression and guiding therapeutic strategies in heart failure. The research proposed in this application is innovative because it focuses on a new diagnostic and prognostic test of HF, tissue and serum BIN1 level, which assess cardiac reserve through directly reflecting the biochemical health of individual cardiomyocytes.