Gopinath Adithya, Badov Martin, Francis Madison, Shaw Gerry, Collins Anthony, Miller Douglas R, Hansen Carissa A, Mackie Phillip, Tansey Malú Gámez, Dagra Abeer, Madorsky Irina, Ramirez-Zamora Adolfo, Okun Michael S, Streit Wolfgang J, Khoshbouei Habibeh
Department of Neuroscience, University of Florida, Center for Translational Research in Neurodegenerative Disease, Norman Fixel Institute for Neurological Diseases, Gainesville, FL, USA.
Encor Biotechnology Inc., Gainesville, FL, USA.
NPJ Parkinsons Dis. 2021 Jul 20;7(1):62. doi: 10.1038/s41531-021-00201-x.
Most, if not all, peripheral immune cells in humans and animals express tyrosine hydroxylase (TH), the rate limiting enzyme in catecholamine synthesis. Since TH is typically studied in the context of brain catecholamine signaling, little is known about changes in TH production and function in peripheral immune cells. This knowledge gap is due, in part, to the lack of an adequately sensitive assay to measure TH in immune cells expressing lower TH levels compared to other TH expressing cells. Here, we report the development of a highly sensitive and reproducible Bio-ELISA to quantify picogram levels of TH in multiple model systems. We have applied this assay to monocytes isolated from blood of persons with Parkinson's disease (PD) and to age-matched, healthy controls. Our study unexpectedly revealed that PD patients' monocytes express significantly higher levels of TH protein in peripheral monocytes relative to healthy controls. Tumor necrosis factor (TNFα), a pro-inflammatory cytokine, has also been shown to be increased in the brains and peripheral circulation in human PD, as well as in animal models of PD. Therefore, we investigated a possible connection between higher levels of TH protein and the known increase in circulating TNFα in PD. Monocytes isolated from healthy donors were treated with TNFα or with TNFα in the presence of an inhibitor. Tissue plasminogen activator (TPA) was used as a positive control. We observed that TNFα stimulation increased both the number of TH+ monocytes and the quantity of TH per monocyte, without increasing the total numbers of monocytes. These results revealed that TNFα could potentially modify monocytic TH production and serve a regulatory role in peripheral immune function. The development and application of a highly sensitive assay to quantify TH in both human and animal cells will provide a novel tool for further investigating possible PD immune regulatory pathways between brain and periphery.
大多数（如果不是全部的话）人和动物的外周免疫细胞都表达酪氨酸羟化酶（TH），它是儿茶酚胺合成中的限速酶。由于TH通常是在脑儿茶酚胺信号传导的背景下进行研究的，因此对外周免疫细胞中TH产生和功能的变化了解甚少。这种知识差距部分是由于缺乏一种足够灵敏的检测方法来测量与其他表达TH的细胞相比，表达较低TH水平的免疫细胞中的TH。在此，我们报告了一种高度灵敏且可重复的生物ELISA方法的开发，用于在多个模型系统中定量皮克水平的TH。我们已将此检测方法应用于从帕金森病（PD）患者血液中分离出的单核细胞以及年龄匹配的健康对照。我们的研究意外地发现，与健康对照相比，PD患者外周单核细胞中TH蛋白的表达水平显著更高。肿瘤坏死因子（TNFα）是一种促炎细胞因子，在人类PD的大脑和外周循环以及PD动物模型中也已显示其水平升高。因此，我们研究了TH蛋白水平升高与PD中已知的循环TNFα升高之间可能存在的联系。用TNFα或在存在抑制剂的情况下用TNFα处理从健康供体分离出的单核细胞。组织纤溶酶原激活剂（TPA）用作阳性对照。我们观察到TNFα刺激增加了TH +单核细胞的数量以及每个单核细胞中TH的量，而没有增加单核细胞的总数。这些结果表明，TNFα可能潜在地改变单核细胞TH的产生，并在外周免疫功能中发挥调节作用。开发和应用一种高度灵敏的检测方法来定量人和动物细胞中的TH，将为进一步研究大脑和外周之间可能的PD免疫调节途径提供一种新工具。
