Investigation of Troponin I-interacting kinase, a cardiomyocyte specific kinase, in the immune response to cardiac injury

 描述（由申请人提供）：病毒性心肌炎（VM）是心力衰竭的重要原因。VM是一种导致扩张型心肌病的隐匿性疾病，通常需要心脏移植。此外，目前仅存在用于对抗VM的支持疗法。对该病的发病机理知之甚少。越来越明显的是，过度的宿主免疫反应在很大程度上决定了VM患者心肌病的进展。基于生物信息学的分析揭示了肌钙蛋白I相互作用激酶（TNNI3K）在VM期间可能调节免疫应答。TNNI3K是心肌细胞（CM）特异性激酶，其调节心脏对缺血和压力超负荷的反应。总体目标是确定TNNI3K作为对抗VM的治疗靶标的有效性。我们的初步研究结果首次表明，TNNI3K加剧了VM期间的免疫反应。虽然免疫细胞浸润是消除病毒病原体所必需的，但急性炎性细胞浸润的严重程度与心功能障碍的发展之间存在明显的有害关系。由于我们的初步发现揭示了（1）在VM的啮齿动物模型中CM特异性TNNI3K的过表达导致心肌免疫细胞浸润增加，以及（2）TNNI3K的CM特异性缺失相反地限制了免疫细胞浸润，我们假设TNNI3K的抑制在急性VM的情况下将是保护性的。此外，由于TNNI3K是CM特异性激酶，因此它是合适的治疗靶标。1）表达野生型或激酶失活的TNNI3K的转基因小鼠，2）条件性CM特异性TNNI3K敲除小鼠（cKO），和3）药理学TNNI3K抑制剂，我们感觉很好地装备来剖析TNNI3K在对心脏损伤的免疫应答中的作用。使用这些工具，我们打算完成以下两个具体目标：1）表征TNNI3K对VM免疫应答的影响。我们将使用流式细胞术和免疫组织化学来表征免疫细胞群和血流动力学，以评估VM小鼠的功能结局。2)研究TNNI3K在调节心脏损伤的免疫应答中的机制。使用分离的成年CM与VM小鼠，我们将使用分子生物学研究底物相互作用，以确定其对免疫反应的影响。总的来说，这些发现将提供关于CM对病毒的免疫应答的见解，提供新的TNNI3K直接相互作用，并回答TNNI3K是否是对抗VM的合适治疗靶标。

 DESCRIPTION (provided by applicant): Viral myocarditis (VM) is a significant contributor to heart failure. VM is an insidious disease that results in dilated cardiomyopathy and often requires cardiac transplantation. Furthermore, only supportive therapies currently exist for combating VM. Very little is known about the pathogenesis of the disease. It is becoming apparent that an excessive host immune response largely defines the progression of cardiomyopathy in patients with VM. Bioinformatics based analyses revealed Troponin I-interacting kinase (TNNI3K) to potentially regulate the immune response during VM. TNNI3K is a cardiomyocyte (CM) specific kinase that regulates the cardiac response to ischemia and pressure overload. The overall goal is to determine the validity of TNNI3K as a therapeutic target for combating VM. Our preliminary findings are the first to show that TNNI3K exacerbates the immune response during VM. Although immune cell infiltration is required for elimination of viral pathogen, there is a clear detrimental relationship between severity of acute inflammatory cell infiltrate and the development of cardiac dysfunction. As our preliminary findings reveal (1) Overexpression of CM-specific TNNI3K in a rodent model of VM results in an increased myocardial immune cell infiltration and (2) CM-specific deletion of TNNI3K conversely limits immune cell infiltration, we hypothesize that inhibition of TNNI3K will be protective in a setting f acute VM. Furthermore, as TNNI3K is a CM-specific kinase, it makes for a suitable therapeutic target. With 1) transgenic mice expressing wild-type, or kinase in-active TNNI3K, 2) conditional CM-specific TNNI3K knockout mice (cKO), and 3) a pharmacological TNNI3K inhibitor, we feel well equipped to dissect the role of TNNI3K in the immune response to cardiac injury. Using these tools, we intend to complete the following two specific aims: 1) Characterize the effect of TNNI3K on the immune response to VM. We will use flow cytometry and immunohistochemistry to characterize immune cell populations and hemodynamics to assess functional outcomes in mice with VM. 2) Examine the mechanism of TNNI3K in regulation of the immune response to cardiac injury. Using isolated adult CM from mice with VM, we will use molecular biology to investigate substrate interactions to determine their effect on the immune response. Overall, these findings will provide insight on the CM immune response to virus, provide novel TNNI3K direct interactions, and answer if TNNI3K is a suitable therapeutic target for combating VM.