Gene expression profile of liver tissue in low-dose, repeated diethylnitrosamine (DEN)-treated rat treated with epigallocatechin gallate (EGCG)

Background: Hepatocellular carcinoma (HCC) is increasing in incidence and treatment is often unsuccessful. Thus, prevention in high-risk patients with established cirrhosis has been proposed as an alternative strategy. It has been suggested that the increased inflammation and fibrogenesis observed during cirrhosis predisposes the liver to future malignant transformation to HCC through a “field effect”. A growing body of evidence indicates that the green tea polyphenol, (-)-epigallocatechin gallate (EGCG), can reduce inflammation and fibrogenesis during chronic liver injury. Here, we test the hypothesis that EGCG administered in the setting of cirrhosis can prevent future HCC development. Methods: A rat model of diethylnitrosamine (DEN)-induced cirrhosis was used to examine the efficacy of EGCG for inhibition of HCC formation in these cirrhotic livers. DEN (50 mg/kg) was administered weekly throughout the study while 0.02% EGCG was given in drinking water beginning at the onset of cirrhosis. At the end of the study, rats were sacrificed, livers were sectioned and stained to analyze disease progression and tumor nodules were counted. Liver function tests were performed to determine liver injury and overall liver function. Finally, genome-wide gene expression profiling on the surrounding, non-tumoral liver tissue was used to monitor the “field effect” in response to EGCG. Results: EGCG significantly (p < 0.01) prevented the development of HCC tumor nodules from on average 18.8 in vehicle controls to on average 9.1 in EGCG-treated animals. EGCG also reduced liver injury and improved liver function as assessed by serum chemistry tests. Finally, a gene expression signature predictive of poor survival and HCC development in human cirrhosis patients was reversed in response to EGCG. Conclusions: Our data are consistent with the growing body of evidence suggesting that EGCG has protective effects in liver disease. Further, our results suggest that EGCG is a potentially effective HCC prevention strategy that can be monitored using gene expression signatures.背景：肝细胞癌（Hepatocellular carcinoma, HCC）的发病率持续上升，且临床治疗往往效果欠佳。因此，针对已确诊肝硬化的高风险人群开展预防被提出作为替代治疗策略。已有研究表明，肝硬化进程中加剧的炎症与纤维化可通过“场效应（field effect）”使肝脏易于发生后续恶性转化，进展为肝细胞癌。越来越多的研究证据显示，绿茶多酚类物质(-)-表没食子儿茶素没食子酸酯（(-)-epigallocatechin gallate, EGCG）可减轻慢性肝损伤过程中的炎症与纤维化。本研究旨在验证下述假说：在肝硬化状态下给予EGCG，可预防后续肝细胞癌的发生。 方法：本研究采用二乙基亚硝胺（diethylnitrosamine, DEN）诱导的肝硬化大鼠模型，探究EGCG对肝硬化肝脏中肝细胞癌形成的抑制效果。实验全程每周给予50 mg/kg的DEN，并于肝硬化造模成功后开始在饮用水中添加0.02%浓度的EGCG。实验结束后处死大鼠，对肝脏进行切片染色以分析疾病进展，并计数肿瘤结节数量。同时开展肝功能检测以评估肝损伤程度与整体肝功能状态。最后，对癌周非肿瘤肝组织进行全基因组基因表达谱（genome-wide gene expression profiling）分析，以监测EGCG干预后的“场效应”变化。 结果：EGCG可显著（p < 0.01）抑制肝细胞癌肿瘤结节的形成：溶剂对照组平均结节数为18.8，而EGCG干预组平均仅为9.1。血清生化检测结果显示，EGCG还可减轻肝损伤并改善肝功能。此外，在人类肝硬化患者中可预测不良预后与肝细胞癌发生的基因表达特征，经EGCG干预后得到了逆转。 结论：本研究数据与日益增多的研究证据一致，表明EGCG对肝脏疾病具有保护作用。此外，本研究结果提示EGCG是一种潜在的高效肝细胞癌预防策略，且可通过基因表达特征对其干预效果进行监测。