Wang Hui, Lv Yuan, Wang Cao, Leng Dongjing, Yan Yan, Blessing Fasae Moyondafoluwa, Madiha Zahra Syeda, Jiang Yanan, Wang Zhiguo, Yang Baofeng, Bai Yunlong
Department of Pharmacology (State-Province Key Laboratories of Biomedicine-Pharmaceutics of China, Key Laboratory of Cardiovascular Medicine Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin, China.
Chronic Disease Research Institute, Translational Medicine Research and Cooperation Center of Northern China, Heilongjiang Academy of Medical Sciences, Harbin, China.
Front Genet. 2019 Aug 13;10:687. doi: 10.3389/fgene.2019.00687. eCollection 2019.
Hepatocellular carcinoma (HCC) is an extremely fatal malignancy. Intestinal microRNAs, which can be detected in fecal samples in humans may be involved in the pathological process of HCC. Therefore, screening for functional intestinal microRNAs in fecal samples and investigating their potential roles in the molecular progression of HCC are necessary. Quantitative real-time PCR (qRT-PCR) has been widely used in microRNA expression studies. However, few genes have been reported as reference genes for intestinal microRNAs in fecal samples. In order to obtain a more accurately analyzed intestinal microRNAs expression, we first searched for reliable reference genes for intestinal microRNAs expression normalization during qRT-PCR, using three software packages (GeNorm, NormFinder, and Bestkeeper). Next we screened and predicted the target genes of the differentially intestinal microRNAs of control and HCC mice through quantitative RT-PCR or miRtarBase. Finally, we also analyzed the mRNA targets for enrichment of Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways using the DAVID Bioinformatic Resources database. This study has successfully screened relatively suitable reference genes and we have discovered that the differential intestinal microRNAs play significant roles in the development of HCC. The top reference genes identified in this study could provide a theoretical foundation for the reasonable selection of a suitable reference gene. Furthermore, the detection of intestinal microRNAs expression may serve as a promising therapeutic target for the diagnosis and treatment of HCC.
肝细胞癌（HCC）是一种极其致命的恶性肿瘤。可在人类粪便样本中检测到的肠道微小RNA可能参与了HCC的病理过程。因此，有必要在粪便样本中筛选功能性肠道微小RNA，并研究它们在HCC分子进展中的潜在作用。定量实时聚合酶链反应（qRT-PCR）已广泛应用于微小RNA表达研究。然而，很少有基因被报道可作为粪便样本中肠道微小RNA的参考基因。为了更准确地分析肠道微小RNA的表达，我们首先使用三个软件包（GeNorm、NormFinder和Bestkeeper）搜索用于qRT-PCR期间肠道微小RNA表达标准化的可靠参考基因。接下来，我们通过定量RT-PCR或miRtarBase筛选并预测对照小鼠和HCC小鼠差异肠道微小RNA的靶基因。最后，我们还使用DAVID生物信息资源数据库分析了基因本体论（GO）术语和京都基因与基因组百科全书（KEGG）通路富集的mRNA靶标。本研究成功筛选出了相对合适的参考基因，并且我们发现差异肠道微小RNA在HCC的发展中发挥着重要作用。本研究中确定的顶级参考基因为合理选择合适的参考基因提供了理论基础。此外，肠道微小RNA表达的检测可能成为HCC诊断和治疗的一个有前景的治疗靶点。
