Growth Control of Normal and Cirrhotic Hepatocytes

描述（由申请人提供）： 肝细胞是一种独特的实质细胞，在严重损伤时可以分裂和再生肝脏。然而，这种再生能力会导致慢性损伤的肝细胞生长紊乱。肝细胞对生长控制信号的反应不同。大多数肝细胞癌发生在肝硬化的肝脏中，这为肝硬化的肝细胞改变了生长控制行为提供了证据。肝硬化是一个主要的世界性健康问题，由于酒精滥用和B和C型肝炎病毒的传播，肝硬化的所有主要原因，预计其患病率将增加。在早期的提议（K 08）中，我们假设肝细胞凋亡对细胞凋亡不太敏感。事实上，研究证实，肝硬化肝细胞对多种凋亡剂的反应较低，包括肿瘤坏死因子α（TNF α）、转化生长因子β（TGF β）和紫外线-C照射（UV-C）。对TGF β诱导的细胞凋亡的抗性是特别感兴趣的，因为这种细胞因子是已知的肝细胞生长抑制剂，并且它以高浓度存在于炎性肝中。TGFR诱导肝细胞凋亡的机制尚不清楚，但我们在原代正常小鼠肝细胞中的研究表明，这是通过caspase-8依赖性机制发生的，该机制需要产生活性氧、线粒体去极化和细胞色素c释放以及caspase-3活化。这是caspase-8依赖性的第一个证明。在肝硬化肝细胞中抑制该途径的机制尚不清楚，但可能涉及其他TGF-受体激活的途径，如Smad信号通路。我们假设Smad信号通路是肝细胞凋亡所必需的，并且该通路中的缺陷存在于肝细胞凋亡中。Smad信号通路的改变可以改变控制存活和死亡之间的细胞平衡的促凋亡因子或抗凋亡因子。在这个建议中，我们的目的是确定是否Smad信号转导通路是必要的TGF β诱导的肝细胞凋亡。此外，由于肝硬化肝细胞表现出抗凋亡，这可能是由于在表达的促凋亡和抗凋亡因子的变化，我们假设，正常和肝硬化肝细胞在其基因表达谱不同。我们建议确定和表征正常和肝细胞凋亡刺激之间的基因表达谱的差异。对这些假设的检验将使我们的实验室在深度和广度上都得到发展，因为我们将使用新的微阵列技术，该技术是在Genomics Core and Microarray Facility内提供的。我们将继续寻求我们的K 08导师（大卫布伦纳和洛拉里德博士）的建议，并自由地使用胃肠道生物学和疾病的核心设施，如分子成像核心。

DESCRIPTION (provided by applicant): Hepatocytes are unique parenchymal cells that divide and regenerate the liver in response to major injury. This regenerative capacity, however, leads to disordered hepatocyte growth in the chronically injured, cirrhotic liver. Hepatocytes in the cirrhotic liver respond differently to growth control signals. Most hepatocellular carcinomas arise in a cirrhotic liver, providing evidence that cirrhotic hepatocytes have altered growth control behavior. Cirrhosis is a major worldwide health problem that is expected to increase in prevalence because of increasing alcohol abuse and dissemination of the hepatitis B and C viruses, all major causes of cirrhosis. In an earlier proposal (K08), we hypothesized that cirrhotic hepatocytes would be less sensitive to apoptosis. Indeed, studies confirmed that cirrhotic hepatocytes are less responsive to multiple apoptotic agents including tumor necrosis factor a (TNFalpha), transforming growth factor beta (TGFbeta), and ultraviolet-C irradiation (UV-C). The resistance to TGFbeta-induced apoptosis is of special interest because this cytokine is a known inhibitor of hepatocyte growth, and it is present in high concentrations in the cirrhotic liver. The mechanisms by which TGFR induces hepatocyte apoptosis are not well known, but our studies in primary normal mouse hepatocytes suggest that this occurs through a caspase-8 dependent mechanism that requires the generation of reactive oxygen species, depolarization of the mitochondria with cytochrome c release, and caspase-3 activation. This is the first demonstration of caspase-8 dependence. The mechanisms that inhibit this pathway in cirrhotic hepatocytes are unknown, but may involve other TGFR-activated pathways such as the Smad signaling pathway. We hypothesize that the Smad signaling pathway is necessary for hepatocyte apoptosis, and that defects in this pathway are present in cirrhotic hepatocytes. Alterations in the Smad signaling pathway may alter pro- or anti-apoptotic factors that govern the cellular balance between survival and death. In this proposal, we aim to determine if the Smad signal transduction pathway is necessary for TGFbeta-induced hepatocyte apoptosis. In addition, because cirrhotic hepatocytes exhibit resistance to apoptosis that may be due to changes in expression of pro-and anti-apoptotic factors, we hypothesize that normal and cirrhotic hepatocytes differ in their gene expression profiles. We propose to identify and characterize differences in gene expression profiles between normal and cirrhotic hepatocytes in response to apoptotic stimuli. Examination of these hypotheses will allow our laboratory to growth both in depth and in breadth, because we will use new microarray technology provided within the UNC Genomics Core and Microarray Facility. We will continue to seek counsel from our K08 mentors (Drs. David Brenner and Lola Reid) and use liberally the UNC Center for Gastrointestinal Biology and Disease for core facilities such as the Molecular Imaging Core.