Chen Ting, Zhang Siwen, Zhou Dongmei, Lu Peipei, Mo Xianglai, Tamrakar Rashi, Yang Xi
Department of Endocrinology, First Affiliated Hospital, Guangxi Medical University, Nanning, China.
Department of Gastrointestinal Surgery, First Affiliated Hospital, Guangxi Medical University, Nanning, China.
Front Oncol. 2022 Aug 11;12:911808. doi: 10.3389/fonc.2022.911808. eCollection 2022.
Non-alcoholic fatty liver disease (NAFLD) is a risk factor for hepatocellular carcinoma (HCC). However, its carcinogenic mechanism is still unclear, looking for both diseases' transcriptome levels, the same changes as we are looking for NAFLD may provide a potential mechanism of action of HCC. Thus, our study aimed to discover the coexisting pathogenic genes of NAFLD and HCC.
We performed a variance analysis with public data for both diseases. At the same time, weighted gene correlation network analysis (WGCNA) was used to find highly correlated gene modules in both diseases. The darkturquoise gene module was found to be highly correlated with both diseases. Based on the diagnosis related module genes and the differential genes of the two diseases, we constructed diagnostic and prognostic models by logistic regression, univariate Cox regression, and LASSO regression. Public datasets verified the results. Meanwhile, we built a competing endogenous RNA (ceRNA) network based on the model genes and explored the related pathways and immune correlation involved in the two diseases by using Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and gene set enrichment analyses. Immunohistochemistry was used to verify the different expression of ABCC5 and TUBG1 among the normal liver, NAFLD, and HCC tissues. Sodium palmitate/sodium oleate was used to establish high-fat cell models, and Real Time Quantitative Polymerase Chain Reaction (RT-qPCR) was used to verify the messenger RNA (mRNA) expression of ABCC5 in lipidization cells.
A total of 26 upregulated genes and 87 downregulated genes were found using limma package identification analysis. According to WGCNA, the darkturquoise gene module was highly correlated with the prognosis of both diseases. The coexisting genes acquired by the two groups were only three central genes, that is, ABCC5, DHODH and TUBG1. The results indicated that the diagnostic and prognostic models constructed by ABCC5 and TUBG1 genes had high accuracy in both diseases. The results of immunohistochemistry showed that ABCC5 and TUBG1 were significantly overexpressed in NAFLD and HCC tissues compared with normal liver tissues. The Oil Red O staining and triglyceride identified the successful construction of HepG2 and LO2 high-fat models using PA/OA. The results of RT-qPCR showed that the lipidization of LO2 and HepG2 increased the mRNA expression of ABCC5.
The gene model constructed by ABCC5 and TUBG1 has high sensibility and veracity in the diagnosis of NAFLD as well as the diagnosis and prognosis of HCC. ABCC5 and TUBG1 may play an important role in the development of NAFLD to HCC. In addition, lipidization could upregulate the mRNA expression of ABCC5 in HCC.
非酒精性脂肪性肝病（NAFLD）是肝细胞癌（HCC）的一个危险因素。然而，其致癌机制仍不清楚，在两种疾病的转录组水平上寻找相同的变化，可能为HCC提供潜在的作用机制。因此，我们的研究旨在发现NAFLD和HCC共存的致病基因。
我们对两种疾病的公共数据进行了方差分析。同时，使用加权基因共表达网络分析（WGCNA）来寻找两种疾病中高度相关的基因模块。发现暗绿松石色基因模块与两种疾病都高度相关。基于诊断相关模块基因和两种疾病的差异基因，我们通过逻辑回归、单变量Cox回归和LASSO回归构建了诊断和预后模型。公共数据集验证了结果。同时，我们基于模型基因构建了竞争性内源性RNA（ceRNA）网络，并通过基因本体论、京都基因与基因组百科全书和基因集富集分析探索了两种疾病中涉及的相关途径和免疫相关性。免疫组织化学用于验证正常肝脏、NAFLD和HCC组织中ABCC5和TUBG1的差异表达。使用棕榈酸钠/油酸钠建立高脂细胞模型，并使用实时定量聚合酶链反应（RT-qPCR）验证脂化细胞中ABCC5的信使核糖核酸（mRNA）表达。
使用limma包鉴定分析共发现26个上调基因和87个下调基因。根据WGCNA，暗绿松石色基因模块与两种疾病的预后高度相关。两组获得的共存基因只有三个核心基因，即ABCC5、二氢乳清酸脱氢酶（DHODH）和TUBG1。结果表明，由ABCC5和TUBG1基因构建的诊断和预后模型在两种疾病中都具有很高的准确性。免疫组织化学结果显示，与正常肝脏组织相比，ABCC5和TUBG1在NAFLD和HCC组织中显著过表达。油红O染色和甘油三酯鉴定了使用棕榈酸/油酸成功构建HepG2和LO2高脂模型。RT-qPCR结果显示，LO2和HepG2的脂化增加了ABCC5的mRNA表达。
由ABCC5和TUBG1构建的基因模型在NAFLD诊断以及HCC诊断和预后方面具有高敏感性和准确性。ABCC5和TUBG1可能在NAFLD向HCC的发展中起重要作用。此外，脂化可上调HCC中ABCC5的mRNA表达。
