Molecular mechanisms regulating leukocyte trafficking

项目摘要/摘要 自主神经系统在压力或“战斗和逃跑”过程中调节先天免疫功能 内源性儿茶酚胺的释放，激活G蛋白偶联受体的反应 肾上腺素能受体家族(GPCRs)在免疫系统中表达。α1-ARs在细胞周期调控中的作用 然而，对先天免疫功能的调节知之甚少，其分子机制 神经激素和先天免疫系统之间潜在的相互作用仍有待确定。这个 趋化因子受体家族(CRS)是调节白细胞募集所必需的，发挥着重要作用 在炎症的方方面面和许多疾病过程中发挥作用。最近，我们发现α1-ARs 与趋化因子(C-X-C基序)受体(CXCR)4及非典型趋化因子形成异低聚复合体 受体(ACKR)3及异构体α1B-AR复合体中α1B-AR对CXCR4的调节作用 人血管平滑肌细胞的迁移。目前尚不清楚CRS之间的异构体络合物是否 白细胞中存在α-1受体。我们在中试实验中观察到，杂低聚化合物之间 α1A/B/D-AR和CXCR4在THP-1细胞表面和新鲜分离的人外周血中均可检测到 血液单个核细胞。此外，我们还进行了初步的分子间生物发光共振。 能量转移(BRET)实验检测到α1b-AR能够与 CR系列的多个成员。这导致了我们的工作假说，杂寡聚体的形成 α-1-ARs和CRs之间的复合体是神经激素通过 系统与先天免疫系统相互作用，调节CR介导的白细胞在健康和 疾病。为了验证这一假设，我们将关注单核细胞作为先天免疫中的主要细胞群。 CRS和α1B-AR之间的系统和异构体，以提供概念验证。我们提出以下建议 具体目的：1.研究CR：α1B-AR异构体复合体在人体内的分布和功能 单核细胞。我们将确定哪些CRS能够与α1B-AR形成异构体，定义 受体异构化，并评估受体异构化对CR信号和功能的影响。2. α1-ARs和CR：α1B-AR异构体复合体在单核细胞调节中的作用 体内炎症过程中的募集。在这里，我们将利用MacBlue小鼠建立腹膜炎模型，以评估 α1-ARs和CR：α1B-AR异构体在炎症中单核细胞迁移调节中的作用这个 拟议的探索性/发育研究将为建立受体提供科学依据 异构化是神经激素系统控制CR介导的白细胞的一种机制 贩卖人口。这些数据可能导致对白细胞调节的理解的范式转变 并有望确定新的分子靶点来调节许多 疾病过程，这有可能促进为患者开发新的治疗策略。

Project Summary/Abstract The autonomous nervous system regulates innate immune functions during the stress or “fight and flight” response through the release of endogenous catecholamines, which activate the G protein-coupled receptor (GPCR) family of adrenergic receptors (ARs) expressed in the immune system. The roles of α1-ARs in the regulation of innate immune functions, however, are poorly understood and the molecular mechanisms underlying cross-talk between the neurohormonal and innate immune system remain to be determined. The family of chemokine receptors (CRs) is essential for the regulation of leukocyte recruitment and plays important roles in all aspects of inflammation and in numerous disease processes. Recently, we discovered that α1-ARs form hetero-oligomeric complexes with chemokine (C-X-C motif) receptor (CXCR) 4 and with atypical chemokine receptor (ACKR) 3, and that α1B-AR within heteromeric CXCR4:α1B-AR complexes regulates CXCR4-mediated migration of human vascular smooth muscle cells. It is unknown whether heteromeric complexes between CRs and α1-ARs exist in leukocytes. We observed in pilot experiments that hetero-oligomeric complexes between α1A/B/D-AR and CXCR4 are detectable on the cell surface of THP-1 cells and in freshly isolated human peripheral blood mononuclear cells. Furthermore, we performed preliminary intermolecular bioluminescence resonance energy transfer (BRET) experiments and detected that α1b-AR is able to form heteromeric complexes with multiple members of the CR family. This leads to our working hypothesis that the formation of hetero-oligomeric complexes between α1-ARs and CRs is a common molecular mechanism through which the neurohormonal system interacts with the innate immune system to regulate CR-mediated leukocyte trafficking in health and disease. To test this hypothesis, we will focus on monocytes as a major cell population in the innate immune system and on heteromers between CRs and α1B-AR to provide proof-of-concept. We propose the following specific aims: 1. To study the distribution and function of CR:α1B-AR heteromeric complexes in human monocytes. We will determine which CRs are able to form heteromers with α1B-AR, define the patterns of receptor heteromerization and assess the effects of receptor heteromerization on CR signaling and function. 2. To assess the roles of α1-ARs and of CR:α1B-AR heteromeric complexes in the regulation of monocyte recruitment during inflammation in vivo. Here we will utilize MacBlue mice in a peritonitis model to evaluate the roles of α1-ARs and CR:α1B-AR heteromers in the regulation of monocyte migration in inflammation. The proposed exploratory/developmental studies will provide the scientific basis to establish receptor heteromerization as a mechanism through which the neurohormonal system controls CR-mediated leukocyte trafficking. These data could lead to a paradigm shift in the understanding of the regulation of leukocyte recruitment and are expected to identify new molecular targets to modulate innate immune functions in numerous disease processes, which has the potential to facilitate the development of new treatment strategies for patients.