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Abstract：
BACKGROUND: A-to-I RNA editing is an abundant post-transcriptional modification event in hepatocellular carcinoma (HCC). Evidence suggests that adenosine deaminases acting on RNA 1 (ADAR1) correlates to oxidative stress that is a crucial factor of HCC pathogenesis. The present study investigated the effect of ADAR1 on survival and oxidative stress of HCC, and underlying mechanisms.
METHODS: ADAR1 expression was measured in fifty HCC and normal tissues via real-time quantitative PCR, and immunohistochemistry. For stable knockdown or overexpression of ADAR1, adeno-associated virus vectors carrying sh-ADAR1 or ADAR1 overexpression were transfected into HepG2 and SMMC-7721 cells. Transfected cells were exposed to oxidative stress agonist tBHP or sorafenib Bay 43-9006. Cell proliferation, apoptosis, and oxidative stress were measured, and tumor xenograft experiment was implemented.
RESULTS: ADAR1 was up-regulated in HCC and correlated to unfavorable clinical outcomes. ADAR1 deficiency attenuated proliferation of HCC cells and tumor growth and enhanced apoptosis. Moreover, its loss facilitated intracellular ROS accumulation, and elevated Keap1 and lowered Nrf2 expression. Intracellular GSH content and SOD activity were decreased and MDA content was increased in the absence of ADAR1. The opposite results were observed when ADAR1 was overexpressed. The effects of tBHP and Bay 43-9006 on survival, apoptosis, intracellular ROS accumulation, and Keap1/Nrf2 pathway were further exacerbated by simultaneous inhibition of ADAR1.
CONCLUSIONS: The current study unveils that ADAR1 is required for survival and oxidative stress of HCC cells, and targeting ADAR1 may sensitize HCC cells to oxidative stress via modulating Keap1/Nrf2 pathway.
METHODS: ADAR1 expression was measured in fifty HCC and normal tissues via real-time quantitative PCR, and immunohistochemistry. For stable knockdown or overexpression of ADAR1, adeno-associated virus vectors carrying sh-ADAR1 or ADAR1 overexpression were transfected into HepG2 and SMMC-7721 cells. Transfected cells were exposed to oxidative stress agonist tBHP or sorafenib Bay 43-9006. Cell proliferation, apoptosis, and oxidative stress were measured, and tumor xenograft experiment was implemented.
RESULTS: ADAR1 was up-regulated in HCC and correlated to unfavorable clinical outcomes. ADAR1 deficiency attenuated proliferation of HCC cells and tumor growth and enhanced apoptosis. Moreover, its loss facilitated intracellular ROS accumulation, and elevated Keap1 and lowered Nrf2 expression. Intracellular GSH content and SOD activity were decreased and MDA content was increased in the absence of ADAR1. The opposite results were observed when ADAR1 was overexpressed. The effects of tBHP and Bay 43-9006 on survival, apoptosis, intracellular ROS accumulation, and Keap1/Nrf2 pathway were further exacerbated by simultaneous inhibition of ADAR1.
CONCLUSIONS: The current study unveils that ADAR1 is required for survival and oxidative stress of HCC cells, and targeting ADAR1 may sensitize HCC cells to oxidative stress via modulating Keap1/Nrf2 pathway.
摘要：
背景：A到IRNA编辑是肝细胞癌（HCC）中丰富的转录后修饰事件。有证据表明，腺苷脱氨酶作用于RNA1（ADAR1）与氧化应激相关，氧化应激是HCC发病的关键因素。本研究调查了ADAR1对肝癌患者生存和氧化应激的影响。和潜在的机制。
方法：通过实时定量PCR检测50例HCC和正常组织中ADAR1的表达，和免疫组织化学。为了稳定敲低或过表达ADAR1,将携带sh-ADAR1或ADAR1过表达的腺相关病毒载体转染到HepG2和SMMC-7721细胞中。将转染的细胞暴露于氧化应激激动剂tBHP或索拉非尼Bay43-9006。细胞增殖，凋亡，并测量了氧化应激，并进行了肿瘤异种移植实验。
结果：ADAR1在HCC中上调，并与不良临床结局相关。ADAR1缺乏减弱HCC细胞的增殖和肿瘤的生长并增强凋亡。此外，它的损失促进了细胞内ROS的积累，和升高Keap1和降低Nrf2表达。在没有ADAR1的情况下，细胞内GSH含量和SOD活性降低，MDA含量增加。当ADAR1过表达时,观察到相反的结果。tBHP和Bay43-9006对存活率的影响，凋亡，细胞内ROS积累，同时抑制ADAR1进一步加剧Keap1/Nrf2通路。
结论：目前的研究表明，ADAR1是肝癌细胞存活和氧化应激所必需的，靶向ADAR1可能通过调节Keap1/Nrf2通路使肝癌细胞对氧化应激敏感。
方法：通过实时定量PCR检测50例HCC和正常组织中ADAR1的表达，和免疫组织化学。为了稳定敲低或过表达ADAR1,将携带sh-ADAR1或ADAR1过表达的腺相关病毒载体转染到HepG2和SMMC-7721细胞中。将转染的细胞暴露于氧化应激激动剂tBHP或索拉非尼Bay43-9006。细胞增殖，凋亡，并测量了氧化应激，并进行了肿瘤异种移植实验。
结果：ADAR1在HCC中上调，并与不良临床结局相关。ADAR1缺乏减弱HCC细胞的增殖和肿瘤的生长并增强凋亡。此外，它的损失促进了细胞内ROS的积累，和升高Keap1和降低Nrf2表达。在没有ADAR1的情况下，细胞内GSH含量和SOD活性降低，MDA含量增加。当ADAR1过表达时,观察到相反的结果。tBHP和Bay43-9006对存活率的影响，凋亡，细胞内ROS积累，同时抑制ADAR1进一步加剧Keap1/Nrf2通路。
结论：目前的研究表明，ADAR1是肝癌细胞存活和氧化应激所必需的，靶向ADAR1可能通过调节Keap1/Nrf2通路使肝癌细胞对氧化应激敏感。
