Analysis of interferon related gene regulation as a proapoptotic function against hepatocellular carcinoma

干扰素（IFN）联合5-氟尿嘧啶（5-FU）治疗在晚期肝细胞癌（HCC）患者中显示出显着的效果。IFN - 2广泛用于慢性肝脏疾病。但IFN α 8是几种癌细胞系中诱导凋亡最有效的亚型。在本研究中，我们研究了IFN α 2或α 8联合5-FU诱导肝癌细胞系凋亡的分子机制。在没有或存在5- fu的情况下，用IFN α诱导5种肝癌细胞株（Hep3B、Huh7、HLE、PLC/PRL/5和HepG2）凋亡。Hep3B对IFN + 5-FU处理最敏感。caspase- 3, -9，尤其是caspase-8的活性与IFN + 5-FU相比，高于IFN或5-FU。当在两种IFN α处理中添加5-FU时，JAK-STAT通路转录因子ISRE被更协同地激活。抑制caspase-3，-8,9，c-Jun n-末端激酶（JNK），磷脂酰亚苷3-激酶（PI3K）和p38丝裂原活化蛋白激酶（p38 MAPK）表明caspase-8抑制是降低IFN和/或5-FU凋亡作用的最有效的治疗方法。在JAK1和isgf3g沉默的Hep3B细胞中，IFN诱导凋亡和caspase-8激活水平，即使与5-FU联合使用，也被取消。虽然IFN α 8比α 2更有效地诱导细胞凋亡，并且在芯片分析中表现出不同的簇，但JAK-STAT和caspase的激活状态相似。我们认为caspase-8是控制ifn和5-FU诱导的肝癌细胞凋亡的最重要因子。

Interferon (IFN) combined with 5-Fluorouracil (5-FU) treatment is recently reported to show marked effects in patients with advanced hepatocellular carcinoma (HCC). IFN alpha 2 is widely provided for chronic liver diseases. But IFN alpha 8 is the most effective subtype to induce apoptosis in several cancer cell lines. In this study, we investigated the molecular mechanisms of apoptosis induction in hepatoma cell lines with IFN alpha 2 or alpha 8 in combination with 5-FU. Five hepatoma cell lines (Hep3B, Huh7, HLE, PLC/PRL/5, and HepG2) were tested for apoptosis inducibility by IFN alpha in the absence or presence of 5-FU. Hep3B was the most apoptosis sensitive to IFN plus 5-FU treatment. Caspases-3, -9, and especially caspase-8 activities were higher with IFN alpha plus 5-FU than IFN or 5-FU alone. The JAK-STAT pathway transcriptional factor ISRE was activated more synergistically when 5-FU was added to both IFN alpha treatments. Inhibition of caspase-3,-8,9,c-Jun N-terminal kinase (JNK), phosphatidylinositide 3-kinase (PI3K), and p38 mitogen-activated protein kinase (p38 MAPK) revealed that caspase-8 inhibition was the most effective treatment to decrease apoptotic effects of IFN and/or 5-FU. In JAK1, and ISGF3g-silenced Hep3B cells, the apoptosis induction and caspase-8 activation levels by IFN, even in combination with 5-FU, were abrogated. Although IFN alpha 8 induced apoptosis more effectively than alpha 2 and showed different cluster in microarray analysis, the JAK-STAT and caspase activation status were similar.We conclude that caspase-8 is the most important factor that controls IFN-and 5-FU- induced apoptosis in hepatoma cell lines.