Mathematical Model of Parotid Acinar Differentiation

描述（由申请人提供）：拟议项目的长期目标是确定信号网络的发展，诱导细胞分化为成熟的唾液浆液性腺泡细胞，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干燥综合症或放射治疗而丧失唾液腺功能。了解唾液细胞的分化是恢复患病或破坏的腮腺唾液组织的必要步骤。以前的工作已经从组织学角度描述了腺泡细胞的终末分化，并通过表征分化标记物的表达，但尚未使用基因组学水平的方法或数学模型来定义调节途径。当前申请的主要目标是开发正式的数学和统计模型动态数学模型将用于生成将被测试的假设，并且该模型将通过纳入新数据来重复完善，以响应 RFA“唾液腺生物学的系统方法”。我们对这些研究的总体假设是，数学模型可以识别控制腮腺腺泡细胞分化的关键调节途径。从胚胎和新生大鼠腮腺腺泡细胞中获取 RNA，用于对整个分化时期的基因表达模式进行微阵列分析，将创建一个耦合常微分方程 (ODE) 模型来描述指导分化过程的假设相互作用，并通过 RT-PCR、IHC 和蛋白质印迹的组合来完善 ODE 模型，因为 microRNA 是发育的重要调节因子。目前还没有出版物描述腮腺组织中的 microRNA。特定目标 3 将创建一个统计算法，通过定义偏差和变异的来源以及评估模型的整体预测能力及其各个子模块来验证和修改 ODE 模型。目标 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，这些假设将用于完善和验证数学模型，该系统生物学方法应确定驱动腮腺腺泡细胞细胞分化的分子途径。这满足了数百万因干燥综合症、放射治疗或因基本药物治疗而导致唾液腺功能障碍的美国人的需求。这项研究是开发基因转移疗法和生物制剂等治疗或缓解口干症口腔症状的新技术的必要基础。

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.