Liver SVCT Systems: Vitamin C Uptake, Roles & Regulation

描述(由申请人提供)： 这项建议旨在推动首席调查员(PI)朝着学术独立的目标前进。为了实现这一目标，他将在研究肝脏中维生素C的调节时学习先进的分子生物学和小鼠遗传技术以及肝脏病理学/组织学。抗坏血酸(维生素C)是人体必需的微量营养素和有效的抗氧化剂。钠依赖维生素C转运体的两种亚型(SVCT1和SVCT2)在许多人和小鼠组织中表达，包括肝脏。关于SVCT系统对肝脏整体维生素C摄取过程的调节或相对贡献，目前尚不清楚。了解这些机制具有临床意义，因为许多肝脏相关疾病都受益于优化维生素C体内的动态平衡。我们的研究将使用体外和体内两种方法来全面检查人类肝脏维生素C的摄取过程和调节。我们将确定维生素C摄取的特征/动力学，表征hSVCT1和hSVCT2启动子，使用siRNA方法确定hSVCT1和hSVCT2对总摄取的相对贡献，并进行关于维生素C摄取过程适应性调节的影响的研究，所有这些都是在人类肝细胞中进行的。此外，我们将确定与人类一样无法合成维生素C的体内小鼠模型肝脏维生素C摄取的特征/动力学，利用cre/lox系统独立生成每个基因的肝脏特异性敲除，建立mSVCT1和mSVCT2对总摄取的相对贡献，并继续我们对该体内模型中维生素C摄取的适应性调节的研究。我们对维持和调节正常维生素C体内稳态的机制的研究将潜在地使临床医生能够在缺乏的情况下为患者制定有效的策略。广泛的培训和教育机会将使PI能够开始作为独立学术研究人员的职业生涯

DESCRIPTION (provided by applicant): This proposal has been designed to advance the Principle Investigator (PI) towards his goal of academic independence. To attain that objective he will learn advanced molecular biology and mouse genetic techniques as well as liver pathology/histology in his studies of vitamin C regulation in the liver. Ascorbic acid (vitamin C) is a required essential micronutrient and effective antioxidant in humans. Two isoforms of the sodium-dependent vitamin C transporters (SVCT1 and SVCT2) are expressed in many human and mouse tissues, including the liver. Nothing is known regarding the regulation or relative contribution of the SVCT systems toward the overall vitamin C uptake process in the liver. Understanding these mechanisms is clinically relevant since many liver related diseases benefit from optimizing vitamin C body homeostasis. Our studies will use both an in vitro and in vivo approach to perform a comprehensive examination of the human liver vitamin C uptake process and regulation. We will determine the characteristics/kinetics of vitamin C uptake, characterize the hSVCT1 and hSVCT2 promoters, establish the relative contribution of hSVCT1 and hSVCT2 toward total uptake using an siRNA approach and perform studies regarding the effects of adaptive regulation of the vitamin C uptake process, all in human liver cells. In addition we will determine the characteristics/kinetics of vitamin C uptake in the liver of an in vivo mouse model that like humans is unable to synthesize vitamin C, establish the relative contribution of mSVCT1 and mSVCT2 toward total uptake using the cre/lox system to generate liver specific knockouts of each gene independently, and continue our studies into adaptive regulation of vitamin C uptake in this in vivo model. Our studies into the mechanisms involved in maintaining and regulating normal vitamin C body homeostasis will potentially allow clinicians to develop effective strategies for patients during conditions of deficiency. The extensive training and educational opportunities will allow the PI to launch into a career as an independent academic researcher