Sweet receptor (T1R2/3) signaling in the upper airway and regulation of immunity

 描述（由申请人提供）：我们最近发现，人鼻窦孤立化学感受细胞（SCC）表达T2 R苦味受体，当激活时，刺激Ca 2+驱动的抗菌肽（AMP）从周围上皮细胞分泌，在先天免疫中起重要作用。我们假设，在体内，SCC使用T2 R通过激活AMP分泌来响应苦味细菌产物。此外，SCC T2 R介导的Ca 2+反应被T1 R3甜味受体抑制，以响应人工甜味剂、甜味D-氨基酸（由许多细菌产生）和气道表面液体（ASL）中生理相关的葡萄糖浓度。这种抑制可能通过cAMP信号转导的T1 R激活而发生。免疫荧光表明T2 R和T1 R3受体在相同的SCC中表达，可能与不同的细胞内信号传导途径偶联。我们假设T1 R介导的T2 R抑制存在于相对健康的时期部分抑制AMP释放，因为AMP的完全分泌和耗尽可能使上皮过度脆弱。在急性感染期间，细菌可能消耗可用的ASL葡萄糖，降低葡萄糖浓度并使T2 R的T1 R抑制失活，从而允许AMP释放。T1 R抑制AMP分泌的能力也受到气道表面液体（ASL）pH值的调节，这可能代表了对抗细菌产生D-氨基酸的宿主适应性机制。 虽然我们已经确定了许多人类SCC T2 R信号转导，但T1 R通路尚不清楚。我们的中心假设是，T1 R2和/或T1 R3甜味受体亚基和T2 R，在相同的鼻窦SCC中表达，耦合到不同的细胞内信号传导途径。为了测试这一点，我们将采取鼻窦组织外植体和人和小鼠鼻窦细胞的分化原代培养物，并且（1）使用免疫荧光来检查T1 R在人SCC中的分布，（2）鉴定SCC中的T1 R信号转导通路，包括衍生小鼠SCC的纯培养物或富集培养物以生化表征SCC T1 R信号转导，（3）研究ASL对SCC T1 R信号的pH调节机制。我们还将对人类Tas 1 R2和Tas 1 R3多态性进行基因分型，以确定它们对SCC中T1 R信号传导的影响。 这些目标的目的是确定鼻腔鼻窦T1 R受体的定位和信号转导，这可能是非常重要的，以开发新的治疗呼吸道感染。ASL葡萄糖在某些情况下异常升高，如糖尿病和慢性鼻窦炎，这两种疾病通常都涉及慢性呼吸道感染。过量的ASL葡萄糖可能过度抑制T2 R介导的先天免疫，使T1 R拮抗剂（例如，lactisole）成为恢复某些患者先天免疫应答的治疗选择。此外，如果Tas 1 R2/3多态性在鼻窦T1 R功能中发挥重要作用，那么未来的研究可能会揭示Tas 1 R遗传学和/或味觉测试可能有助于预测上呼吸道感染的易感性和/或T1 R靶向治疗的有效性。

 DESCRIPTION (provided by applicant): We recently discovered that human sinonasal solitary chemosensory cells (SCCs) express T2R bitter taste receptors that, when activated, stimulate Ca2+ -driven secretion of antimicrobial peptides (AMPs) from surrounding epithelial cells, playing an important role in innate immunity. We hypothesize that, in vivo, SCCs use T2Rs to respond to bitter bacterial products by activating AMP secretion. Furthermore, SCC T2R-mediated Ca2+ responses are inhibited the T1R3 sweet receptor in response to artificial sweeteners, sweet D-amino acids (produced by many bacteria), and physiologically-relevant glucose concentrations in the airway surface liquid (ASL). This inhibition may occur through T1R-activation of cAMP signaling. Immunofluroescence suggests that T2R and T1R3 receptors are expressed within the same SCCs, likely coupled to different intracellular signaling pathways. We hypothesize that T1R-mediated inhibition of T2Rs exists to partially inhibit AMP release during times of relative health, as complete secretion and depletion of AMPs may leave the epithelium excessively vulnerable. During acute infection, bacteria may consume available ASL glucose, decreasing glucose concentration and deactivating T1R inhibition of T2Rs, allowing AMP release. The ability of T1Rs to inhibit AMP secretion is also modulated by airway surface liquid (ASL) pH, which may represent a host adaptive mechanism to counter D-amino acid production by bacteria. While we have defined much of human SCC T2R signaling, this T1R pathway is unknown. Our central hypothesis is that T1R2 and/or T1R3 sweet receptor subunits and T2Rs, expressed in the same sinonasal SCCs, are coupled to different intracellular signaling pathways. To test this, we will take sinonasal tissue explants and differentiated primary cultures of human and mouse sinonasal cells and (1) use immunofluorescence to examine T1R distribution in human SCCs, (2) identify T1R signal transduction pathway(s) in SCCs, including deriving pure or enriched cultures of mouse SCCs to biochemically characterize SCC T1R signaling, and (3) examine the mechanism of ASL pH regulation of SCC T1R signaling. We will also genotype human Tas1R2 and Tas1R3 polymorphisms to determine their influence on T1R signaling in SCCs. The goal of these aims is to identify the localization and signal transduction of sinonasal T1R receptors, which may be very important to developing new therapies for respiratory infections. ASL glucose is abnormally elevated in certain conditions such as diabetes mellitus and chronic rhinosinusitis, both of which often involve chronic respiratory infections. Excessive ASL glucose may overly inhibit T2R-mediated innate immunity, making T1R antagonists (eg, lactisole) therapeutic options to restore innate immune responses in certain patients. Moreover, if Tas1R2/3 polymorphisms play an important role in sinonasal T1R function, then future studies could reveal that Tas1R genetics and/or taste testing may be useful for making predictions about susceptibility to upper respiratory infections and/or the effectiveness of T1R-targeted therapeutics.