The Fetal Hepatocyte Phenotype and Cell-Based Therapy for Liver Disease

描述（申请人提供）：肝脏的生长和功能分化对妊娠晚期胎儿代谢、围产期过渡和新生儿代谢适应至关重要。胎儿肝脏发育的生物学也对胎儿体细胞生长的调节、肝癌的发生和肝脏疾病的细胞治疗有影响。我们的实验室长期以来一直关注啮齿动物妊娠晚期肝脏的发育。在此过程中，我们确定了一种由调节胎儿肝细胞生长、增殖和基因表达的信号通路定义的胎儿肝细胞表型。最近，我们证明了妊娠晚期胎鼠肝细胞具有关键肝细胞标志物亮氨酸氨基肽酶（LAP）的能力，可以重新填充受伤的成年肝脏。这种能力是成年大鼠肝细胞所不具备的。目前的建议是基于一种中心假设，即组蛋白变异和组蛋白翻译后修饰（PTMs）通过对染色质结构的影响，解释了妊娠晚期胎鼠肝细胞的信号表型以及移植到成人肝脏微环境后这种表型的持久性。该项目的长期目标是确定解释胎儿肝细胞新特征的遗传和表观遗传机制，从而将对胎儿肝脏发育的理解应用于肝脏疾病细胞治疗的发展。这一目标将通过三个具体目标来实现。特异性目的1是通过检测胎儿晚期和成年期间肝脏中的染色质结构、组蛋白变异和组蛋白PTMs来鉴定胎儿肝细胞的表观遗传特征。特异性目的2是表征胎儿到成年过渡期间肝脏中组蛋白变异、组蛋白PTMs、染色质结构和关键生长调节基因调控之间的功能关系。这一目标将验证特定组蛋白变异和ptm与关键生长调节基因的发育变化相关的假设。在具体目标3中，我们将评估目标1和目标2中确定的表观遗传特征在确定和维持妊娠晚期胎鼠肝细胞重新填充受伤成年大鼠肝脏的能力中的作用。这一目的将验证下述假设：胎儿肝脏表观遗传特征是具有肝细胞和胆管标记的胎儿肝细胞亚群的特征，并且这种表观遗传特征在移植前和移植后都存在。这些目标的完成将提供对决定发育中的肝脏细胞行为的分子机制的深入了解，同时推进基于细胞的肝脏疾病治疗的发展。该项目在许多方面都具有创新性。我们将获得有关肝脏发育过程中基因表达的表观遗传调控的新信息。在此过程中，我们将描述胎儿肝脏中先前未被识别的多能细胞群

DESCRIPTION (provided by applicant): Growth and functional differentiation of the liver are critical to late gestation fetal metabolism, the perinatal transition and metabolic adaptation by the newborn. The biology of fetal liver development also has implications for regulation of fetal somatic growth, hepatic carcinogenesis, and cell-based therapy for liver disease. Our laboratory has long focused on late gestation liver development in the rodent. In doing so, we have identified a fetal hepatocyte phenotype that is defined by the signaling pathways that regulate fetal hepatocyte growth, proliferation and gene expression. More recently, we demonstrated the ability of late gestation fetal rat liver cells possessing a key hepatocyte marker, leucine amino peptidase (LAP), to repopulate an injured adult liver. This is a capacity not shared by adult rat hepatocytes. The present proposal is based on the central hypothesis that histone variants and histone posttranslational modifications (PTMs), acting through effects on chromatin structure, account for the signaling phenotype of late gestation fetal rat hepatocytes and the persistence of this phenotype following transplantation into the adult liver microenvironment. The long term goal of the project is to identify genetic and epigenetic mechanisms that account for the novel characteristics of fetal liver cells, thereby applying an understanding of fetal liver development o the development of cell-based therapy for liver disease. This goal will be pursued through three Specific Aims. Specific Aim 1 is to identify a fetal hepatocyte epigenetic signature by examining chromatin structure, histone variants and histone PTMs in liver during late fetal versus adult life Specific Aim 2 is to characterize the functional relationship between histone variants, histone PTMs, chromatin structure and regulation of key growth-regulating genes in liver during the fetal to adult transition. This aim will test the hypothesis that specific histone variants and PTMs are associated with developmental changes in critical growth-regulating genes. In Specific Aim 3, we will assess the role of the epigenetic signature identified in Aims 1 and 2 in defining and maintaining the ability of late gestation fetal rat liver cells to repopulate an injured adult rat iver. This aim will test the hypothesis that a fetal hepatic epigenetic signature is characteristic of a subpopulation of fetal hepatocytes that possess both hepatocyte and bile ductular markers, and that this epigenetic signature is present both before transplantation and after engraftment. Completion of these aims will provide insight into molecular mechanisms that determine cell behavior in the developing liver while advancing the development of cell-based therapy for liver disease. The project is innovative in a number of aspects. We will generate new information regarding the epigenetic regulation of gene expression during liver development. In doing so, we will characterize a previously unidentified, multipotent cell population in fetal liver that can repopulate an injured adult liver. Finally, we will use innovative methods throughout these studies, thus advancing the ability to study epigenetic regulation in the context of mammalian developmental physiology.