乙型肝炎病毒X蛋白（HBx蛋白）基于1 H-NMR-代谢揭示核酸代谢

StephenW

Hepatitis B virus X protein (HBx)-induced abnormalities of nucleic acid metabolism revealed by 1H-NMR-based metabonomics

    Dan Yue, Yuwei Zhang, Liuliu Cheng, Jinhu Ma, Yufeng Xi, Liping Yang, Chao Su, Bin Shao, Anliang Huang, Rong Xiang & Ping Cheng

    Scientific Reports 6, Article number: 24430 (2016)
    doi:10.1038/srep24430
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        Cancer metabolismOncogenesis

Received:
    07 October 2015
Accepted:
    30 March 2016
Published online:
    14 April 2016

Abstract

Hepatitis B virus X protein (HBx) plays an important role in HBV-related hepatocarcinogenesis; however, mechanisms underlying HBx-mediated carcinogenesis remain unclear. In this study, an NMR-based metabolomics approach was applied to systematically investigate the effects of HBx on cell metabolism. EdU incorporation assay was conducted to examine the effects of HBx on DNA synthesis, an important feature of nucleic acid metabolism. The results revealed that HBx disrupted metabolism of glucose, lipids, and amino acids, especially nucleic acids. To understand the potential mechanism of HBx-induced abnormalities of nucleic acid metabolism, gene expression profiles of HepG2 cells expressing HBx were investigated. The results showed that 29 genes involved in DNA damage and DNA repair were differentially expressed in HBx-expressing HepG2 cells. HBx-induced DNA damage was further demonstrated by karyotyping, comet assay, Western blotting, immunofluorescence and immunohistochemistry analyses. Many studies have previously reported that DNA damage can induce abnormalities of nucleic acid metabolism. Thus, our results implied that HBx initially induces DNA damage, and then disrupts nucleic acid metabolism, which in turn blocks DNA repair and induces the occurrence of hepatocellular carcinoma (HCC). These findings further contribute to our understanding of the occurrence of HCC.

Introduction

With a high degree of malignancy, rapid progression, and high mortality rate, hepatocellular carcinoma (HCC) is the third most frequent cause of cancer-related death worldwide1. Hepatitis B virus (HBV) infection is a major risk factor for HCC2. In recent years, ~2 billion individuals have been found to be infected with HBV, of which, more than 350 million are chronically infected3,4. Thus, HBV has become one of the most important pathogens threatening human health.

As is well known, the main function of the liver is metabolism, thus plays a central role in metabolism of glucose, lipids, amino acids and nucleic acids. HBV infection leads to liver dysfunction, which in turn induces alterations of metabolism. Several recent studies have shown that HBV-related liver disease resulted in many metabolic abnormalities. One study showed that metabolite components of HCC markedly differed from those of adjacent non-tumor tissues, and low-grade HCC had clear metabolomics differences from high-grade HCC5. Another study reported that HBV-related HCC exhibited glucose and lipid metabolic abnormalities6. The levels of many indicators associated with lipid metabolism are altered in patients with chronic HBV infection7,8. Additionally, HBV infection can induce other abnormalities of metabolism such as amino acids, cholesterol, and choline9,10,11.

Hepatitis B virus X protein (HBx) is a multifunctional regulator, which is involved in viral replication, transcriptional regulation, cell cycle progression, DNA repair, apoptosis, and genetic stability12. Recently, several studies have also shown that HBx can disturb cell metabolism. HBx can interact with peroxisome proliferator-activated receptor-γ (PPAR-γ) and sterol regulatory element binding protein 1, increase their respective mRNA and protein expression, and lead to lipid accumulation in hepatocytes7. HBx can induce activation of lipogenic genes and fatty acid accumulation, which contributes to disease progression by promoting steatosis and liver inflammation in transgenic mice13. However, no systematic investigations about the effects of HBx on cell metabolism have been reported thus far.

Metabonomics is a systematic approach in studying low-molecular-weight compounds in cells, tissues and biofluids, which directly reflects the phenotypic features of an organism and provides important information on disease processes, biochemical functions, and drug toxicity14,15. Here, an NMR-based metabolomics approach was applied to study the mechanisms of HBx-mediated carcinogenesis. Studying the HBx-related mechanisms contributes to understanding of the occurrence of HCC.

StephenW

乙型肝炎病毒X蛋白（HBx蛋白）基于1 H-NMR-代谢揭示核酸代谢的诱导异常

    淡月，余味张，郑溜溜，金湖马，裕丰兮，杨丽萍，诏素，邵斌，黄安良，戎祥与平城

    科学报告6，商品编号：24430（2016）
    DOI：10.1038 / srep24430
    下载文献
        癌症metabolismOncogenesis

收稿日期：
    2015年10月07日
公认：
    2016年3月30日
网络发布时间：
    2016年4月14日

抽象

乙型肝炎病毒X蛋白（HBx蛋白）起着HBV相关肝癌中起重要作用;然而，潜在的HBx蛋白介导的肿瘤发生机制尚不清楚。在这项研究中，基于NMR的代谢途径被施加到系统地研究的HBx对细胞代谢的影响。的EdU掺入测定法进行检查的HBx对DNA合成，核酸代谢的重要特征的影响。结果显示了HBx打乱葡萄糖，脂质和氨基酸的代谢，特别是核酸。理解核酸代谢的HBx蛋白诱导的异常的可能机制，表达HBx的HepG2细胞的基因表达图谱进行了调查。结果表明，参与DNA损伤和DNA修复29个基因中的HBx表达HepG2细胞中差异表达。 HBx蛋白诱导的DNA损伤是由染色体核型分析，彗星试验，免疫印迹法，免疫荧光和免疫组化分析进一步显现。许多研究先前报道，DNA损伤可诱导核酸代谢异常。因此，我们的结果暗示了HBx最初诱导DNA损伤，然后破坏核酸代谢，这反过来块DNA修复和诱导肝细胞癌（HCC）的发生。这些发现进一步有助于我们肝癌的发生的理解。

介绍

随着恶性程度高，病情进展快，死亡率率高的，肝细胞癌（HCC）是癌症相关死亡worldwide1的第三个最常见的原因。乙型肝炎病毒（HBV）感染是HCC2的主要危险因素。近年来，〜已经找到2十亿个人被感染乙肝病毒，其中350多万是长期infected3,4。因此，乙肝已成为威胁人类健康的最重要病原菌之一。

如众所周知的，肝脏的主要功能是代谢，从而起到在葡萄糖，脂质，氨基酸和核酸的代谢中心作用。乙肝病毒感染导致的肝功能障碍，而这又引起代谢的改变。最近的几项研究表明，HBV相关肝病导致许多代谢异常。一项研究表明，肝癌的代谢产物成分明显不同于相邻的非肿瘤组织的不同，低档肝癌已经从高档HCC5明确的代谢差异。另一项研究报告说，乙肝相关的肝癌表现出血糖和血脂代谢abnormalities6。与脂质代谢有关的许多指标水平改变了慢性HBV infection7,8。此外，HBV感染可诱导代谢的其他异常如氨基酸，胆固醇和choline9,10,11这样。

乙型肝炎病毒X蛋白（HBx蛋白）是一种多功能的调节器，其涉及病毒复制，转录调控，细胞周期进展，DNA修复，细胞凋亡，和遗传stability12。最近，一些研究还表明了HBx可以扰乱细胞的新陈代谢。 HBx蛋白可与过氧化物酶体增殖物激活受体γ（PPAR-γ）和固醇调节元件结合蛋白1相互作用，增加它们各自的mRNA和蛋白的表达，并导致在hepatocytes7脂质积聚。 HBx蛋白能诱导生脂基因和脂肪酸的积累，促进脂肪变性和肝脏炎症在转基因mice13有助于疾病进展的激活。然而，关于HBx蛋白对细胞代谢的影响没有系统的调查，迄今已报道。

代谢组是在研究中的细胞，组织和生物流体，它直接反映了生物体的表型特征，并提供关于疾病的进程，生化功能，和药物toxicity14,15重要信息的低分子量化合物的系统方法。在这里，一个基于核磁共振的代谢组学方法被用于研究HBx蛋白介导的致癌机制。研究HBx蛋白相关的机制，有助于肝癌的发生的理解。

StephenW

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