MECHANISMS OF IMPAIRED REGENERATION IN CIRRHOTIC LIVER

这项研究计划将探讨基本机制， 导致肝脏生长的严重临床问题。 的 作者在开发几个方面有丰富的经验， 动物模型是深入研究肝脏疾病的理想动物模型， 再生，并建议使用这些模型在一个严格的， 补充的方式，试图研究的机制，关于 控制正常和肝硬化肝脏的肝再生。 的 有待检验假设包括：1）所谓的 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 再生的大鼠在体内。 2)的显著改变 肽结合发生在再生和再生的质膜上， 肝，并负责，部分减少增殖 在肝硬化中观察到的反应; 3）抗增殖因子，如 生长抑素-14（SS-14）和转化生长因子β（TGF-β） 直接作用于第二信使抑制肝细胞增殖 大鼠肝细胞中的途径，这可以通过特异性 药理学探针 特别是双脱氧腺苷（DDA）（抑制剂 腺苷酸环化酶的催化亚基）和其他药理学 试剂可以模拟SS-14的作用， 4）GI肽的信号转导机制 肝增殖的受体与G蛋白有关， 肝硬化时G蛋白结构和功能发生改变; 5)肝硬化中存在早期基因表达模式的改变， 负责衰减增殖反应;具体目标 这一建议将是：1）试图增加肝再生 通过给予外源性生长因子在痴呆大鼠中的作用。 （二） 研究肝再生和肝硬化对结合的影响 EGF、TGF-α、胰岛素、胰高血糖素、SS-14和TGF-β与肝血浆 3）比较不同药理学方法的抗增殖作用 腺苷酸环化酶的探针-其可以模拟外源肽- 系统对静止期和肝切除正常肝细胞的影响， 4）确定G蛋白在信号转导中的作用 导致肝细胞增殖的转导途径，并比较 G蛋白在正常肝组织中的表达与在肝病肝组织中的表达的差异 肝切除术和肝硬化对肝组织中即刻表达的影响 早期反应基因，已知在肝脏中起作用， 再生 用于实现特定目标的模型 包括：1）对假肝切除大鼠进行促肝细胞因子刺激 通过质膜结合分析，第二信使分析， 结构G蛋白的研究，早期基因调控，和分析 肝DNA合成; 2）70%肝切除和生化及分子生物学 如上所述的分析; Wistar大鼠模型; 4） 分离培养的肝细胞制备物; 5）高度纯化的亚细胞 组分，包括分离的肝质膜制剂。

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.