Mechanism of Alcohol/Alcoholism-induced Liver Neoplasia

描述（由申请人提供）：肝细胞癌（HCC）是实体器官恶性肿瘤中常见的死亡原因。酒精是肝细胞癌的一个致病因素。迄今为止，没有动物模型证明酒精/酒精中毒在没有其他HCC危险因素的情况下诱导HCC。进行了初步研究，以确定是否自愿消耗大量酒精的大鼠长时间可以作为HCC的动物模型。为酒精偏好选择性饲养的成年雄性大鼠（P大鼠）自由获取食物和水（“非饮用者”）或食物、水和10%（v/v）酒精溶液（“饮用者”）18个月. P大鼠饮酒者的平均酒精摄入量为6.7 g/kg BW/天。在18个月结束时，P饮酒者肝脏的大体检查显示100%的肿瘤发生率，肝脏组织学显示多个异型增生灶，腺瘤性高度异型增生结节和HCC。相反，在不饮酒者中没有发现肉眼可检测到的肿瘤病变。为了确定酒精偏好和肝肿瘤发生之间的关联不是偶然的，拟议的赠款研究将描述两个单独的大鼠系的肝肿瘤发生的特征，这些大鼠系选择性地饲养酒精偏好（P：Wistar背景; HAD：NIH背景）。肝肿瘤发生的程度和时间将通过超声、病理学和组织病理学（H&E，GSTp+）检查来确定。还将测定增殖（BrDU摄取）、凋亡（TUNEL/ApopTag）和血管生成（CD 31）的指数。为了评估酒精/酒精中毒诱导的肝肿瘤形成的机制是否涉及这些动物中代谢肝酶表达的改变，肝酒精代谢酶的基础水平将测定P与Wistar对照大鼠和HAD与NIH对照大鼠的CYP 2 E1、ADH和ALDH;还将在P和HAD大鼠中测量这些酶的酒精诱导水平，并将其与肿瘤发生，血液酒精水平，血清营养参数、酒精摄入量和研究期间的食物摄入量。以前，我们已经在多种模型中证明了MEK-ERK信号在肝脏肿瘤发生中的作用，但这在酒精/酒精中毒诱导的HCC中尚未探索。为了评估酒精/酒精中毒诱导的肝肿瘤形成的机制是否依赖于改变的ERK/MAPK途径信号传导，将测定P与Wistar对照大鼠和HAD与NIH对照大鼠中总ERK和磷酸化ERK的基础水平;还将测量P和HAD大鼠中酒精诱导的水平并将其与肿瘤发生、血液酒精水平、血清营养参数酒精摄入量和食物摄入量。最后，ERK/MAPK通路信号传导的作用将通过用口服活性MEK抑制剂与安慰剂对照治疗P大鼠来进一步测试，以确定对酒精诱导的肝肿瘤形成的影响。这种酒精/酒精中毒诱导的HCC动物模型可能被证明是一个宝贵的工具，用于研究支持这种特定的HCC病因学的机制，并进一步了解酒精诱导的癌症的一般致病机制。 公共卫生相关性：这项拟议的研究将涉及描述两种不同的动物模型，以表明在没有其他危险因素的情况下，长期饮酒会导致肝癌的发展。可能的机制也将被调查。这种动物模型将是研究酒精暴露导致肝癌的机制的宝贵工具，也将进一步加深我们对酒精导致癌症的机制的理解。

DESCRIPTION (provided by applicant): Hepatocellular carcinoma (HCC) is a common cause of death among solid organ malignancies. Alcohol is an etiologic factor for HCC. No animal model to date has demonstrated that alcohol/alcoholism induces HCC in the absence of other HCC risk factors. Preliminary studies were conducted to determine whether rats which voluntarily consume large quantities of alcohol for prolonged periods can serve as an animal model of HCC. Adult male rats selectively bred for alcohol preference (P rats) were given free access to food and water ("non- drinkers"), or food, water and a 10% (v/v) alcohol solution ("drinkers") for 18 months. Average alcohol intake was 6.7 g/kg BW/day in P rat drinkers. At the end of 18 months, gross examination of livers of P drinkers revealed 100% tumor incidence, and liver histology showed multiple dysplastic foci, adenomatous high grade dysplastic nodules, and HCC. In contrast, no grossly detectable neoplastic lesions were found in P non- drinkers. To establish that the association between alcohol preference and hepatic tumorigenesis is not fortuitous, the proposed grant studies will characterize hepatic tumorigenesis in two separate lines of rats selectively bred for alcohol preference (P: Wistar background; HAD: NIH background). Extent and timing of hepatic tumorigenesis will be determined by ultrasound, pathologic and histopathologic (H&E, GSTp+) examination. Indices of proliferation (BrDU uptake), apoptosis (TUNNEL/ApopTag), and angiogenesis (CD31) will also be determined. To evaluate whether mechanisms of alcohol/alcoholism-induced liver neoplasia involve altered metabolic liver enzyme expression in these animals, basal levels of hepatic alcohol metabolizing enzymes (CYP2E1, ADH and ALDH) in P vs Wistar control and HAD vs NIH control rats will be determined; alcohol-induced levels of these enzymes will also be measured in P and HAD rats and correlated with tumorigenesis, blood alcohol level, serum nutritional parameters, alcohol intake and food intake over the timecourse of the study. Previously, we have demonstrated a role for MEK-ERK signaling in liver tumorigenesis in multiple models, but this has not been explored in alcohol/alcoholism-induced HCC. To evaluate whether mechanisms of alcohol/alcoholism-induced liver neoplasia depend upon altered ERK/MAPK pathway signaling, basal levels of total and phosphorylated ERK in P vs Wistar control and HAD vs. NIH control rats will be determined; alcohol-induced levels in P and HAD rats will also be measured and correlated with tumorigenesis, blood alcohol level, serum nutritional parameters, alcohol intake and food intake over the time course of the study. Finally, the role of ERK/MAPK pathway signaling will be further tested by treating P rats with an orally active MEK inhibitor vs placebo control to determine the effect on alcohol-induced liver neoplasia. This animal model of alcohol/alcoholism-induced HCC may prove to be an invaluable tool for studying the mechanisms which support this specific etiology of HCC and further our understanding of the general pathogenic mechanisms of alcohol-induced cancers. Public Health relevance: The proposed research will involve characterizing two distinct animal models to show that prolonged alcohol intake leads to the development of liver cancer in the absence of other risk factors. Possible mechanisms involved will also be investigated. Such an animal model will be an invaluable tool for studying the mechanism by which alcohol exposure causes liver cancer and also further our understanding of the mechanisms by which alcohol causes cancer in general.