Mathematical Model of Parotid Acinar Differentiation

描述（由申请人提供）：该项目的长期目标是确定诱导细胞分化为成熟唾液浆液腺泡细胞的信号网络的发展，从而允许基因治疗方法再生或替换患者的唾液组织。数以百万计的患者因格林综合征或放射治疗而遭受唾液腺功能丧失。了解唾液腺细胞的分化是使患病或受损的腮腺唾液腺组织得以恢复的必要步骤。以前的工作已经从组织学上描述了腺泡细胞的终末分化，并通过表征分化标志物的表达，但没有使用基因组学水平的方法或数学模型来定义调控途径。当前应用的主要目标是建立正式的数学和统计模型，以确定导致腮腺腺泡细胞最终分化的网络。动态数学模型将用于产生将被测试的假设，并且模型将通过合并新数据而反复改进。这个应用程序是响应RFA“唾液腺生物学的A系统方法”。我们对这些研究的总体假设是，一个数学模型可以识别控制腮腺腺泡细胞分化的关键调控途径。具体目标1将使用激光捕获显微解剖（LCM）从胚胎和新生大鼠腮腺腺泡细胞中获取RNA，用于微阵列分析基因表达模式在分化期间。将创建一个耦合的常微分方程（ODE）模型来描述指导微分过程的假设相互作用。将通过RT-PCR、免疫组化和Western blots的结合对假设进行检验，并改进ODE模型。由于microRNAs是重要的发育调节因子，特异性Aim 2将定义microRNAs在腺泡细胞中的表达，以及分化过程中的变化模式。目前还没有描述腮腺组织中的microrna的出版物。所得结果将用于修正微分的数学模型。具体目标3将创建一个统计算法，通过定义偏差和变异的来源，以及通过评估模型的整体预测能力，以及在其各个子模块中，来验证和修改ODE模型。这将允许置信区间与ODE模型中的不同路径相关联。Specific Aim 4将使用ODE模型对腮腺腺泡细胞中调节基因表达的特定途径做出假设。这些假设将通过转染和转导实验进行验证，结果将进一步完善和验证数学模型。这种系统生物学方法应该确定驱动腮腺腺泡细胞分化的分子途径。项目描述：本研究的总体目标是明确控制细胞向分泌性唾液腺泡细胞分化的分子机制。这解决了数百万美国人的需求，这些人由于Sj"gren's综合征、放射治疗或基本药物导致的口干而遭受唾液腺功能障碍。本研究为开发治疗或缓解口干症的基因转移疗法和生物制剂等新技术奠定了必要的基础。

DESCRIPTION (provided by applicant): The long term goal of the proposed project is to define the development of signaling networks that induce differentiation of cells into mature salivary serous acinar cells to allow gene therapy approaches to regenerating or replacing salivary tissue in patients. Millions of patients suffer loss of salivary gland function due to Sj"gren's syndrome or radiation therapy. Understanding the differentiation of salivary cells is a necessary step to enable the restoration of diseased or destroyed parotid salivary tissue. Previous work has described terminal differentiation of acinar cells histologically, and by characterizing the expression of markers of differentiation, but has not used genomics-level approaches, or mathematical models, to define regulatory pathways. The primary goal of the current application is to develop formal mathematical and statistical models that will identify networks which cause terminal differentiation of parotid acinar cells. The dynamic mathematical models will serve to generate hypotheses which will be tested, and the model will be repeatedly refined by the incorporation of new data. This application is in response to the RFA "A Systems Approach to Salivary Gland Biology." Our overall hypothesis for these studies is that a mathematical model can identify key regulatory pathways that control parotid acinar cell differentiation. Specific Aim 1 will use Laser Capture Microdissection (LCM) to obtain RNA from embryonic and newborn rat parotid acinar cells for microarray analysis of the patterns of gene expression across the period of differentiation. A coupled Ordinary Differential Equation (ODE) model will be created to describe the hypothetical interactions that direct the process of differentiation. The hypotheses will be tested, and the ODE model refined, by a combination of RT-PCR, IHC, and Western blots. Since microRNAs are important regulators of development, Specific Aim 2 will define the expression of microRNAs in acinar cells, and the pattern of changes during differentiation. There are currently no publications describing microRNAs in the parotid tissue. The results will be used to revise the mathematical model of differentiation. Specific Aim 3 will create a statistical algorithm to validate and revise the ODE model by defining the sources of bias and variation as well as by assessing the model's predictive power overall, and in its various sub-modules. This will allow confidence intervals to be associated with different pathways within the ODE model. Specific Aim 4 will use the ODE model to make hypotheses about specific pathways regulating gene expression in the parotid acinar cells. These hypotheses will be tested by transfection and transduction experiments, and the results shall be used to refine and validate the mathematical model. This systems biology approach should identify molecular pathways that drive cytodifferentiation of parotid acinar cells. Project Narrative: The overall goal of this research is to define the molecular mechanisms which control differentiation of cells into secretory salivary acinar cells. This addresses the needs of millions of Americans who suffer from salivary gland dysfunction due to Sj"gren's Syndrome, radiation therapy, or xerostomia due to essential medications. This research is a necessary foundation for developing new technologies such as gene transfer therapy and biologics for treating or alleviating the oral symptoms of xerostomia.