MECHANISMS OF IMPAIRED REGENERATION IN CIRRHOTIC LIVER

这项研究计划将研究基本的机制。 负责肝脏生长的关键临床问题。这个 作者有丰富的经验，开发了几个 一种理想的深入研究肝脏的动物模型 并建议将这些模型用于严格和 互补的方式，试图研究机制，关于 正常和肝硬变肝脏再生的控制。这个 有待检验的假设包括：1)所谓的管理 促肝因子，如胰岛素、胰高血糖素、表皮生长因子 (EGF)和转化生长因子-α(TGF-α)刺激肝脏 肝硬变大鼠体内再生的研究。2)发生重大变化 再生和肝硬变患者的质膜上存在多肽结合 肝脏，这在一定程度上是导致增殖减少的原因 肝硬变的反应；3)抗增殖因子，如 生长抑素-14和转化生长因子-β 直接作用于第二信使抑制肝细胞增殖 大鼠肝细胞内的通路，可通过特异性的 药理探头。特别是二脱氧腺苷(DDA)(一种抑制剂 腺酰环化酶催化亚单位)和其他药理作用 试剂可以模拟SS-14的作用，并改变细胞内 环磷酸腺苷的蓄积；4)胃肠肽的信号机制 肝脏增殖受体与G蛋白有关，而且 肝硬变时G蛋白结构和功能发生改变； 5)在肝硬变中存在早期基因表达模式的改变 对减弱的增殖反应负责；具体目的 这项提议的目的是：1)尝试增加肝脏再生 外源性生长因子在肝硬变大鼠体内的作用。2) 研究肝再生和肝硬变对结合力的影响 表皮生长因子、转化生长因子-α、胰岛素、胰高血糖素、生长抑素-14和转化生长因子-β对肝血浆的影响 3)比较两种药物的抗增殖作用。 腺酰环化酶的探针--可能模仿外源多肽 静息状态下正常肝细胞和肝切除后肝细胞的系统 肝硬变大鼠；4)确定G蛋白在信号中的作用 导致肝细胞增殖的转导途径，并进行比较 正常肝脏中的G蛋白与肝硬变中的G蛋白；5)研究 肝切除和肝硬变对即刻表达的影响 已知在肝脏中起作用的早期反应基因 再生。用于实现特定目标的模型 包括：1)假手术肝切除大鼠促肝因子刺激 通过质膜结合分析，第二信使分析， 结构G蛋白研究，早期基因调控和分析 肝DNA合成；2)70%肝切除及生化和分子 上述分析；30 Wistar肝硬变大鼠模型；4) 分离培养的肝细胞制剂；5)高纯度的亚细胞 组分，包括分离的肝脏质膜制剂。

This research proposal will examine the fundamental mechanisms responsible for the critical clinical problem of hepatic growth. The authors have had extensive experience in the development of several animal models which are ideal for the in depth study of hepatic regeneration, and propose to use these models in a rigorous and complementary fashion to attempt to study the mechanisms, regarding control of hepatic regeneration in both normal and cirrhotic liver. The hypotheses to be tested include: 1) Administration of so-called hepatotropic factors, such as insulin, glucagon, epidermal growth factor (EGF), and transforming growth factor alpha (TGF-alpha) stimulate liver regeneration of cirrhotic rats in vivo. 2) Significant alterations in peptide binding occur in plasma membranes from regenerating and cirrhotic liver, and are responsible, in part for the decreased proliferative response seen in cirrhosis; 3) Antiproliferative factors, such as somatostatin-14 (SS-14) and transforming growth factor beta (TGF-beta) inhibit hepatic proliferation by direct action on second messenger pathways in rat hepatocytes, which can be mimicked by specific pharmacologic probes. Specifically dideoxyadenosine (DDA) (an inhibitor of the catalytic subunit of adenylyl cyclase) and other pharmacologic reagents may mimic the action of SS-14, and alter intracellular accumulation of cyclic AMP; 4) Signaling mechanisms of GI peptide receptors for hepatic proliferation are linked to G proteins, and alterations of G protein structure and function occur during cirrhosis; 5) Altered patterns of early gene expression exist in cirrhosis which are responsible for the attenuated proliferative response; The specific aims of this proposal will be: 1) To attempt to increase hepatic regeneration in cirrhotic rats by the administration of exogenous growth factors. 2) To study the effect of hepatic regeneration and cirrhosis on the binding of EGF, TGF-alpha, insulin, glucagon, SS-14, and TGF-beta to liver plasma membranes; 3) To compare the antiproliferative effects of pharmacologic probes - which may mimic exogenous peptides - of the adenylyl cyclase system on hepatocytes from quiescent and hepatectomized normal and cirrhotic rat liver; 4) To determine the role of G-proteins in signal transduction pathways leading to hepatocyte proliferation, and to compare G proteins in normal liver to those found in cirrhotic liver; 5) To study the effects of hepatectomy and cirrhosis on the expression of immediate early response genes which are known to play a role in hepatic regeneration. The models utilized to accomplish the specific aims include 1) hepatotropic factor stimulation of the sham hepatectomized rat with analysis of plasma membrane binding, second messenger analysis, structural G-protein studies, early gene regulation, and analysis of hepatic DNA synthesis; 2) 70% hepatectomy and biochemical and molecular analysis as described above; 30 the Wistar cirrhotic rat model; 4) isolated cultured hepatocyte preparations; 5) highly purified subcellular fractions, including isolated liver plasma membrane preparations.