Novel heterotypic cell cultures for liver toxicology studies

描述（由申请人提供）：我们提案的目的是研究酒精损伤期间实质和非实质肝细胞之间的旁分泌纤维化信号。尽管非实质细胞（星状细胞）仍然是肝脏发育的主要参与者，但实质细胞（肝细胞）在这一过程中的作用正在引起越来越多的关注。更好地了解实质和非实质肝细胞在酒精损伤中所起的作用，将转化为更好、更有效和更有针对性的治疗方法。然而，不同肝细胞类型之间复杂的信号交换很难用体内模型或标准的体外细胞培养方法来描述。在本提案中，我们将利用一种新颖的肝细胞培养方法进行研究！该假说认为，肝脏产生的转化生长因子(TGF)-¿在启动星状细胞的成纤维程序中起重要作用。这一假设将通过一种新的细胞培养系统进行验证，该系统将细胞黏附物和/或信号分子印迹到培养底物中，从而将肝细胞和星状细胞的离散组定位在确定的位置并彼此靠近（见图1 (a，B)）。如图1c所示，这种新型细胞培养皿将用于在酒精损伤期间选择性地刺激抗纤维化生长因子（GFs）（例如HGF和BMP7）共培养中的肝细胞，以研究GF信号传递给肝细胞如何影响邻近星状细胞的激活。所提出的平台的新颖性在于我们能够定义在同一培养皿中培养的两种肝细胞类型的相互作用，从而调节一种细胞类型的表型并研究另一种细胞类型的反应。这种方法特别适合于提高我们对肝纤维化背后复杂的异型信号的理解。这里描述的新型细胞培养系统被设想为肝毒理学研究和肝保护分子的高通量筛选的使能技术。

DESCRIPTION (provided by applicant): The goal of our proposal is to investigate paracrine fibrogenic signaling between parenchymal and non parenchymal liver cells during alcohol injury. Whereas the nonparenchymalcompartment (stellate cells) remains the major player in the development of liver, the role of the parenchymal cells (hepatocytes) in this process is drawing increasing attention. Improved understanding of the roles played by parenchymal and nonparenchymalliver cells during alcohol insult will translate into better, more effective and targeted therapeutics. However, the complex signaling exchange between different liver cell types is difficult to delineate with in vivo models or standard in vitro cell culture approaches. In this proposal, we will utilize a novel liver cell culture approach to investigate !he hypothesis that hepatic production of transforming growth factor (TGF)-¿ plays an important role in initiating the fibrogneic program of stellate cells. This hypothesis will be explored using a novel cell culture system where cell adhesive and/or signaling molecules are imprinted into a culture substrate so as to position discrete groups of hepatocytes and stellate cells in defined locations and in close proximity to each other (see Figure 1 (A,B)). As shown in Figure 1 C, this novel cell culture dish will be used to selectively stimulate hepatocytes within the co-cultures with anti-fibrotic growth factors (GFs) (e.g. HGF and BMP7) during alcohol injury in order to investigate how GF signals delivered to hepatocytes impact activation of neighboring stellate cells. The novelty of the proposed platform lies in our ability to define interactions of two liver cell types cultured in the same dish so as to modulate the phenotype of one cell type and to study the response of the other cell type. This approach is particularly well-suited for improving our understanding of the complex heterotypic signaling underlying liver fibrosis. The novel cell culture system described here is envisioned as an enabling technology for liver toxicology studies and for high-throughput screening of liver-protective molecules. PUBLIC HEALTH RELEVANCE: Liver is the metabolic center of the human body and is largely responsible for carrying out detoxification functions in the body. Consumption of alcohol has an adverse effect on the liver and can lead to liver dysfunction. We are developing novel liver cell culture in order to better understand effects of alcohol on the liver and to enable development of liver-protective therapies.