PARK14/Calcium signaling as a novel biomarker for Parkinson disease

帕金森氏病(PD)是一种无法治愈的神经退行性疾病，进展缓慢(没有临床症状 在帕金森病相关运动功能障碍的首发症状出现前多年。绝大多数人 约85%的帕金森病患者为特发性帕金森病(IdPD)，目前尚无特异性生物标志物可用于早期诊断 这种毁灭性的疾病在年老的人类中。博洛蒂纳博士实验室的最新发现(周等人，《自然》 通信，2016)导致在PARK14/PLA2G6中识别出以前未知的缺陷(S)- 依赖的钙信号(PLA2g6/Ca2+)，可在idPD患者的外周细胞中检测到，并可能 用于开发全新的生物标记物战略。我们还证实，这种缺陷会导致 老年性帕金森病患者多巴胺能神经元的进行性死亡和年龄依赖性帕金森病样运动功能障碍的发展 在人类体内模拟idpd的新小鼠模型。重要的是，我们发现人类idPD与 随着PLA2G6(L)表达/功能的显著丧失和存储操作的钙离子进入(SOCE)的损伤， 我们可以在idPD患者的试验组的原代皮肤成纤维细胞(PSF)中检测到这种情况。 在这里，我们假设血小板和/或PSF中PLA2G6/钙离子信号的特征缺陷可能是使用的 作为一种新的检测老年人idPD的生物标志物。我们正在为一项试点计划寻求支持，该计划将 重点验证我们新的基于PLA2g6/Ca2+的生物标记物的特异性，并将测试使用的可行性 人血小板和/或皮肤成纤维细胞中的这些生物标志物是检测idPD的一种新方法。我们的研究 团队是唯一有资格进行拟议研究的团队：博洛蒂纳博士(PI)拥有独特的知识并建立了 在人类受试者的试验组中验证PLA2g6/钙离子生物标记物的实验平台，以及Saint- 希莱尔(临床联合调查员)目前参与了帕金森进展标志物倡议(PPMI)的研究， 并有一个完善的捐赠者招募和样本收集平台。所有方法都已建立，并且 由PI和Co-Investigator出版。初步数据充分支持我们的假设，并证明了我们的 求婚。我们建议的具体目标是： 目的1：利用现有的原发瘤样本验证特征PLA2g6/Ca~(2+)缺陷对idPD的特异性 NINDS资料库中的皮肤成纤维细胞。PLA2g6(L)、钙离子通道和内质网的表达和功能 将使用分子、生化和成像技术分析钙离子水平，并在成纤维细胞中进行比较 来自idPD、家族性PD(LRRK2或GBA突变)、阿尔茨海默病、亨廷顿病、 并控制非神经科捐赠者。所有样本将从NINDS人体细胞和数据库中获得。 目的2：检测是否能在idPD患者的血小板中检测到PLA2g6/钙信号的标志性缺陷 病人。活的血小板和皮肤成纤维细胞的样本将从老年患者的试验组中获得 诊断为早期idPD，并与年龄匹配的对照组非神经科供者进行比较。 PLA2g6/Ca~(2+)信号将使用分子、生化和成像技术进行分析。

Parkinson’s disease (PD) is an incurable neurodegenerative disorder that progresses silently (without clinical manifestations) for years prior to onset of the first symptoms of PD-associated motor dysfunction. The vast majority of PD cases (about 85%) are idiopathic (idPD), and currently, there are no specific biomarkers for early diagnosis of this devastating disease in aging humans. Recent discoveries in Dr. Bolotina’s lab (Zhou et al, Nature Communications, 2016) resulted in identification of the previously unknown defect(s) in PARK14 / PLA2g6- dependent Ca2+ signaling (PLA2g6/Ca2+), which could be detected in peripheral cells from idPD patients, and may be used for development of a brand new biomarker strategy. We also established that such defects lead to progressive demise of dopaminergic neurons and development of age-dependent PD-like motor dysfunction in a new mouse model that closely mimics idPD in humans. Importantly, we found a strong association of human idPD with significant loss of PLA2g6(L) expression/function and impairment of the store-operated Ca2+ entry (SOCE), which we could detect in primary skin fibroblasts (PSFs) from a pilot group of idPD patients. Here we hypothesize that signature defects in PLA2g6/Ca2+ signaling in platelets and/or PSFs could be used as a novel biomarker for detection of idPD in aging humans. We are seeking support for a pilot program that will focus on validation of the specificity of our new PLA2g6/Ca2+-based biomarker, and will test the feasibility of using these biomarkers in human platelets and/or skin fibroblasts as a novel approach for detection of idPD. Our research team is uniquely qualified for proposed studies: Dr. Bolotina (PI) has unique knowledge and established experimental platform for validation of PLA2g6/Ca2+ biomarkers in the pilot groups of human subjects, and Dr. Saint- Hilaire (clinical Co-Investigator) is currently involved in the Parkinson’s Progression Markers Initiative (PPMI) study, and has a well-established platform for donor recruitment and sample collection. All approaches are established and published by PI and Co-Investigator. Preliminary data fully support our hypothesis and demonstrate feasibility of our proposal. Specific Aims of our proposal are: Aim 1: To validate specificity of the signature PLA2g6/Ca2+ defects for idPD using existing samples of primary skin fibroblasts from NINDS repository. Expression and function of PLA2g6(L), store-operated Ca2+ entry and ER Ca2+ levels will be analyzed using molecular, biochemical, and imaging techniques, and compared in fibroblasts from the patients with idPD, familial PD (mutations in LRRK2 or GBA), Alzheimer’s disease, Huntington’s disease, and control non-neurologic donors. All samples will be obtained from NINDS human cell and data repository. Aim 2: To test if the signature defects in PLA2g6/Ca2+ signaling could be detected in platelets from idPD patients. The samples of live platelets and skin fibroblasts will be obtained from a pilot group of aged patients diagnosed with early stages of idPD, and compared with the age-matched group of control non-neurologic donors. PLA2g6/Ca2+ signaling will be analyzed using molecular, biochemical, and imaging techniques.