Functional Regulation of the Lacrimal Duct System

泪道系统的功能调节

• 批准号: 7372957
• 负责人: CHUANQING DING
• 金额: $ 40.75万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-01-01 至 2011-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7372957
• 关键词: Acinar Cell; Acinus organ component; Address; American; Animal Model; Attention; Autoimmune Process; Biology; Caring; Cell Volumes; Cell physiology; Cells; Characteristics; Complex; Condition; Coupled; Data; Development; Diestrus; Dry Eye Syndromes; Duct (organ) structure; Ductal; Ductal Epithelial Cell; Electrolytes; Epithelial; Epithelial Cells; Exhibits; Eye; Female; Flow-It; Fluorescence Microscopy; Functional disorder; GTP-Binding Proteins; Goals; Human; Image; Immunohistochemistry; Impairment; In Situ Hybridization; Ions; Knowledge; Lacrimal gland structure; Lasers; Lead; Liquid substance; Lymphocyte; Mediating; Methods; Microdissection; Modeling; Monitor; Muscarinic Acetylcholine Receptor; Muscarinic Acetylcholine Receptor M3; Ophthalmology; Oryctolagus cuniculus; Patients; Phenotype; Physiological; Pilocarpine; Polymerase Chain Reaction; Pregnancy; Production; Receptor Signaling; Regulation; Research; Research Personnel; Risk; Role; Scanning; Syndrome; System; Techniques; Testing; Time; Vision research; Visit; Water; absorption; eye dryness; innovation; insight; lacrimal; laser capture microdissection; multi-photon; ocular surface; programs; water channel

DESCRIPTION (provided by applicant): Dry eye afflicts millions of Americans, and it is the leading cause of visits to eye care clinicians. Dry eye often results from insufficient tear formation from the lacrimal gland. Lacrimal fluid is initially formed in acini and then modified during passage through the duct system. However, little is known about the ducts and their function. It has been assumed that the decreased lacrimal production in dry eye is caused primarily by decreased acinar secretion. However, our preliminary data suggest that a competing hypothesis also must be considered, i.e., lacrimal hyposecretion may result from inappropriate or excessive reabsorption of fluid in the ducts. In this proposal, the investigators will use normal rabbits and two rabbit models, pregnancy and induced autoimmune dacryoadenitis [IAD], both of which exhibit decreased basal lacrimal secretion and pathological features of lacrimal insufficiency, to address three aims: #1 Determine whether the volume and composition of lacrimal fluid is modified in isolated duct segments and deduce the functional capabilities of duct epithelial cells in normal control rabbits. #2 Determine whether transepithelial ion and fluid transport functions of isolated duct segments are altered in pregnancy and IAD. #3 Identify cellular mechanisms underlying decreased lacrimal fluid production in pregnancy and IAD. These studies will employ a broad repertoire of innovative techniques. Microdissection and microperfusion methods will be used to determine ionic and volumetric changes in fluids perfused through specific duct segments under basal and pilocarpine-stimulated conditions, and to assess the potential influence of periductal lymphocytes on these functions. Ex vivo real time imaging using multi-photon excitation laser scanning fluorescence microscopy will be used to assess changes in cytosolic Ca2+ associated with M3 muscarinic acetylcholine receptor (M3AChR) signaling; changes in cell volume associated with activation of K+ and Cl- efflux mechanisms and regulatory activation of Na+ and Cl- influx mechanisms; and changes in cytosolic electrolyte contents associated with activation or inhibition of ion transporters hypothesized to mediate transepithelial absorption or secretion. Laser capture microdissection, real time RT- PCR, immunohistochemistry and in situ hybridization will be used to determine whether expression of aquaporins, M3AChR, G proteins coupled to M3AChR (i.e., Gq and G11), and various ion transporters associated with epithelial electrolyte and fluid absorption and secretion is altered in duct cells in the two models. Therefore, these studies will define the functional roles of specific duct segments within the lacrimal gland in production of lacrimal fluid. This research is highly relevant to the Program Goals of the National Plan for Eye and Vision Research, and knowledge gained from these studies has considerable potential to enhance the development of more effective treatment strategies.Dry eye is the most prevalent problem in ophthalmology which afflicts millions of Americans, and is the leading cause of visits to eye care clinicians. The proposed studies take advantage of two excellent animal models to study the biology of the lacrimal gland duct system, which has heretofore received little attention. Gaining an understanding of the lacrimal duct system is critical to enhancing our ability to develop more effective treatment strategies ultimately benefiting millions of dry eye patients.

描述(申请人提供)：干眼症困扰着数以百万计的美国人，是眼科医生就诊的主要原因。干眼症通常是由于泪腺形成的泪液不足所致。泪液最初在腺泡中形成，然后在通过导管系统的过程中被修改。然而，人们对这些管道及其功能知之甚少。一般认为干眼症患者泪液分泌减少主要是由于腺泡分泌物减少所致。然而，我们的初步数据表明，还必须考虑一个相互竞争的假设，即泪道排出不足可能是由于导管内液体的不适当或过度再吸收所致。在这项研究中，研究人员将使用正常兔和两种兔模型-妊娠和诱导性自身免疫性泪腺炎[IAD]，这两种模型都表现出基础泪液分泌减少和泪腺功能不全的病理特征，以解决三个目标：#1确定分离的泪道节段泪液的体积和组成是否发生改变，并推断正常对照兔泪道上皮细胞的功能能力。#2确定妊娠和IAD时离体管段的跨上皮离子和液体运输功能是否改变。#3确定妊娠和IAD患者泪液分泌减少的细胞机制。这些研究将采用广泛的创新技术。将使用显微解剖和微灌流方法来确定在基础和匹罗卡品刺激条件下通过特定管段灌流的液体的离子和体积变化，并评估导管周围淋巴细胞对这些功能的潜在影响。使用多光子激发激光扫描荧光显微镜的体外实时成像将用于评估与M3 M胆碱型乙酰胆碱受体(M3AChR)信号相关的细胞内钙的变化；与激活K+和Cl-外流机制以及调节激活Na+和Cl-内流机制相关的细胞体积的变化；以及与激活或抑制可能介导跨上皮吸收或分泌的离子转运体相关的细胞液电解质含量的变化。采用激光捕获显微切割、实时定量RT-PCR、免疫组织化学和原位杂交等方法，检测两种动物模型中导管细胞水通道蛋白、M3AChR、与M3AChR偶联的G蛋白(即Gq和G11)以及与上皮电解质和液体吸收分泌相关的各种离子转运体的表达是否发生改变。因此，这些研究将确定泪腺内特定管段在泪液产生中的功能作用。这项研究与国家眼睛和视力研究计划的计划目标高度相关，从这些研究中获得的知识具有相当大的潜力来促进更有效的治疗策略的开发。干眼是眼科中最普遍的问题，困扰着数百万美国人，也是眼科护理临床医生就诊的主要原因。建议的研究利用两个优秀的动物模型来研究泪腺导管系统的生物学，到目前为止，这一系统几乎没有受到关注。对泪道系统的了解对于提高我们制定更有效的治疗策略的能力至关重要，最终使数百万干眼患者受益。