Odorant-Receptor Interactions in Olfactory Neurons

描述(申请人提供)：这个项目的长期目标是研究调节嗅觉上皮(OE)中祖细胞增殖和神经发生的细胞间信号机制。短期内，使用靶向消融诱导神经元凋亡，随后是巨噬细胞活化/渗透和祖细胞增殖的同步波，该项目重点关注巨噬细胞作为调节细胞间信号转导的生物活性分子来源的作用。转基因小鼠在先导研究中的创新应用表明，在没有激活的巨噬细胞的情况下，靶细胞切除后祖细胞的增殖显著减少。这些数据表明趋化因子巨噬细胞炎性蛋白(MIP)-1α及其受体CCR1参与巨噬细胞的募集，并提示A类巨噬细胞清道夫受体(MSR-A)参与了凋亡神经元的清除，从而将凋亡、吞噬、细胞增殖和神经发生联系在一起。拟议的体内实验主要关注靶细胞消融后2小时至72小时发生的细胞和分子事件，当时巨噬细胞的渗透和随后的信号传递开始，通过解决三个特定的目标。第一个是研究靶点消融后MIP-1α和CCR1mRNA表达的变化过程，并将它们的蛋白定位于OE。二是评价MIP-1α-/-小鼠体内MIP-1α蛋白缺失对巨噬细胞浸润和祖细胞增殖的影响，以及外源性MIP-1α蛋白能否调节MIP-1α基因敲除小鼠的巨噬细胞浸润和祖细胞增殖。第三个是通过比较野生型小鼠和MRS-A-/小鼠的清除率来检测MSR-A作为巨噬细胞吞噬清除凋亡神经元的中介的表达。基因图谱方法将用于基因敲除小鼠的实验，以检查额外的趋化因子受体和/或信号级联的参与。这些研究将阐明OE中导致祖细胞增殖和神经发生的信号通路的关键组成部分，并建立两个新的动物模型来研究OE细胞在体内的动力学。了解OE中调节神经发生的信号将有助于深入了解因衰老、创伤和神经退行性疾病而受损的嗅觉功能恢复机制。

DESCRIPTION (provided by applicant): The long-term objective of this project is to investigate intercellular signaling mechanisms that regulate progenitor cell proliferation and neurogenesis in the olfactory epithelium (OE). Short-term, using target ablation to induce neuronal apoptosis followed by a synchronous wave of macrophage activation/infiltration and progenitor cell proliferation, this project focuses on the role of macrophages as sources of bioactive molecules that regulate intercellular signaling. Innovative use of transgenic mice in pilot studies demonstrated that in the absence of activated macrophages, progenitor cell proliferation is substantially reduced following target ablation. The data implicate the chemokine macrophage inflammatory protein (MIP)-1alpha and its receptor CCR1 in macrophage recruitment and suggest that the class A macrophage scavenger receptor (MSR-A) is involved in the clearance of apoptotic neurons, thus linking apoptosis, phagocytosis, cell proliferation, and neurogenesis. The proposed in vivo experiments focus on cellular and molecular events that occur at 2 his to 72 hrs after target ablation, when macrophage infiltration and subsequent signaling is initiated, by addressing three specific aims. The first is to characterize the tune course of mRNA expression for MIP-1alpha and CCR1 following target ablation and to localize their proteins in the OE. The second is to assess the effects of the absence of MIP-1alpha protein in MIP-1alpha-/- mice on macrophage infiltration and progenitor cell proliferation and to determine if the effects can be modulated by exogenous MIP-1alpha protein in knockout mice. The third is to examine the expression of MSR-A as a mediator of phagocytic clearance of apoptotic neurons by macrophages by comparing clearance in wild-type mice and MRS-A-/ mice. Gene profiling methodology will be used in the experiments on knockout mice to examine the involvement of additional chemokine receptors and/or signaling cascades. These studies will elucidate key components of the signaling pathways leading to progenitor cell proliferation and neurogenesis in the OE and establish the use of two new animal models in which to investigate OE cell dynamics in vivo. Understanding the signaling regulating neurogenesis in the OE will yield insights into mechanisms for restoring olfactory function compromised by aging, trauma, and neurodegenerative diseases.