Basal-Lateral/Endomembrane Traffic in Lacrimal Acini

描述（由申请人提供）：干燥综合征（SjS）和其他免疫相关疾病在200万至400万美国人中导致泪道功能障碍、视力受损和眼表炎症。原发性泪源性缺乏症（PLD），也可能有免疫相关成分，影响至少600多万人。这些概念通过以下假设得到统一：泪腺的环境反映了一种涉及调节性淋巴细胞、自身免疫效应淋巴细胞和分泌性上皮细胞的免疫稳态；淋巴细胞和炎症细胞介质调节上皮细胞功能；免疫稳态的进化是对上皮细胞功能改变的反应。然而，关于改变上皮功能引发疾病的触发因素，女性比男性更频繁发生的原因，以及导致泪道功能障碍的分子机制，我们知之甚少。研究人员发现，用毒蕈碱受体（MAChR）激动剂（carbachol）慢性刺激泪腺泡细胞，可引起上皮分泌静止，并激活异常的膜运输程序，阻止溶酶体的运动，潜在地增加构成性自身抗原的暴露，并启动先前隐性表位的暴露。与MAChR、Gq和G11结合的G蛋白是多种介质的受体共享的，它们的信号传导受到性激素的影响。因此，研究人员提出：(a)生理扰动可以启动不适当的Gq/G11信号，激活异常的交通程序；(b) SjS和PLD的局部环境刺激不适当的Gq/G11信号，导致潜在的泪液功能障碍的功能静止。具体目标激活异常膜交通程序的信号是什么？这个

DESCRIPTION (provided by applicant): Sjogren's syndrome (SjS) and other immune-related diseases cause lacrimal dysfunction, impaired vision, and ocular surface inflammation in 2 to 4 million Americans. Primary lacrimal deficiency (PLD), which also may have an immune-related component, affects at least 6 million more. These concepts are unified by the hypothesis that: the milieu in the lacrimal glands reflects an immunohomeostasis involving regulatory lymphocytes, autoimmune effector lymphocytes, and secretory epithelial cells; lymphocyte and inflammatory cell mediators modulate epithelial cell functions; and the immunohomeostasis evolves in response to altered epithelial cell functions. However, little is known about the triggers that alter epithelial function to initiate the diseases, the reasons they occur more frequently in women than in men, and the molecular mechanisms that cause lacrimal dysfunction. The investigators have found that chronic stimulation of lacrimal acinar cells with the muscarinic receptor (MAChR) agonist, carbachol, causes epithelial secretory quiescence and also activates an aberrant endomembrane traffic program that blocks movement to lysosomes, potentially increasing exposure of constitutive autoantigens and initiating exposure of previously cryptic epitopes. The G proteins that classically couple to MAChR, Gq and G11, are shared by receptors for a wide variety of mediators, and their signaling is influenced by the sex hormones. Therefore, the investigators propose that: (a) physiological perturbations can initiate inappropriate Gq/G11 signaling that activates the aberrant traffic program, and (b) the local environments in SjS and PLD stimulate inappropriate Gq/G11 signaling that causes functional quiescence underlying lacrimal dysfunction. Specific Aim 1. What are the signals that activate the aberrant membrane traffic program? The central hypothesis is that MAChR remain activated and continue to activate Gq/G11. Specific Aim 2. Can chronic stimulation of other receptors also cause functional quiescence and activate the aberrant traffic program? The central hypothesis is that these changes can be elicited by agonists for receptors that utilize Gq/G11, including histamine, 5-hydroxytryptamine, PGE2, and estradiol. Specific Aim 3. What traffic effectors are responsible for the aberrant program? The central hypothesis is that chronic stimulation decreases dynein motor function but increases p150(Glued) association with kinesin II, thereby increasing kinesin-mediated traffic. Specific Aim 4. How extensively does the aberrant program alter traffic of lysosomal proteins? The investigators will use a GFP-cathepsin S fusion protein and confocal microscopy to test the hypothesis that lysosomal proteins accumulate in the endosomes.