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

描述(由申请人提供)：超过500万美国人患有心力衰竭(HF)，每年新增超过50万例，这是一种日益增长的流行病。然而，心力衰竭进展的病理生理学仍然知之甚少，因此，我们做出治疗决定的能力仍然有限。为了改善临床治疗，需要开发与心力衰竭进展直接相关的更具特异性和预后的生物标记物。我最近发现，一种关键的支架蛋白BIN1在人类心力衰竭钙处理改变的发病机制中起着重要作用。有了这些知识，我的长期目标是通过了解BIN1的生物学和正常和疾病心肌细胞中钙的处理来开发一种新的心脏储备的HF生物标志物。这种特殊应用的目的是了解BIN1在调节钙瞬变过程中的作用。 并使用BIN1开发心力衰竭患者的诊断和预后测试。我的中心假设是，BIN1在衰竭的人类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力相关。其基本原理是，这项拟议研究的成功完成将填补基于BIN1的心衰钙瞬变调节知识的空白，这有助于开发一种心衰患者心脏储备的创新诊断和预测测试。具体目标1是确定BIN1在Cav1.2转运和衰竭的人心肌细胞钙瞬变调节中的作用。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管分级将被用来评估BIN1和Cav1.2在衰竭和未衰竭的人心肌细胞中的表达和定位。在成年小鼠心肌细胞模型中，将使用表面生物素化和活细胞钙成像来研究Cav1.2在BIN1耗竭的心肌细胞中的转运和钙瞬变。具体目的#2是确定心肌组织BIN1的表达作为终末期人类心肌病患者心力衰竭进展的诊断和预后测试。定量免疫荧光将用于心脏活检以确定心室BIN1的表达水平，以便与临床收缩参数和结果数据进行分析。具体目的#3是确定血清BIN1是预测心肌病患者心脏储备功能的一种新的心衰生物标志物。加州大学旧金山分校心力衰竭诊所的大量心肌病患者和年龄匹配的正常对照人群的血清BIN1将通过心脏特异性ELISA法进行测定，以便与临床数据进行分析。这项贡献有望将BIN1确定为心力衰竭的介体和生物标记物，以帮助跟踪心力衰竭的进展并预测临床结果。这一贡献将是重大的，因为这一发现将改变目前心肌健康评估中用于监测心力衰竭疾病进展和指导治疗策略的范式。这项应用中提出的研究具有创新性，因为它专注于一种新的诊断和预测心力衰竭、组织和血清BIN1水平的测试，该测试通过直接反映单个心肌细胞的生化健康来评估心脏储备。

DESCRIPTION (provided by applicant): 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.