扶正消癥方通过调控 circRNA-miRNA-mRNA 网络改善脂糖代谢发挥抗肝癌作用。

Fuzheng Xiaozheng prescription exerts anti-hepatocellular carcinoma effects by improving lipid and glucose metabolisms via regulating circRNA-miRNA-mRNA networks.

机构信息

School of Basic Medical Sciences, Chengdu University of Traditional Chinese Medicine, Chengdu, 611137, China.

Sichuan Provincial Key Laboratory for Development and Utilization of Characteristic Horticultural Biological Resources, College of Chemistry and Life Sciences, Chengdu Normal University, Chengdu, 611130, China.

出版信息

Phytomedicine. 2022 Aug;103:154226. doi: 10.1016/j.phymed.2022.154226. Epub 2022 Jun 1.

DOI:10.1016/j.phymed.2022.154226

PMID:35689900

Abstract

BACKGROUND

Hepatocellular carcinoma (HCC) is a major threat to human health due to its high lethality. Our previous studies suggested that Fuzheng Xiaozheng prescription (FZXZP), an effective Chinese medicine, demonstrated significant suppressive effects on HCC. However, its underlying mechanism remains largely unclear.

PURPOSE

This study aimed to investigate the anti-HCC mechanisms of FZXZP from transcriptomic sequencing based on a holistic perspective.

METHODS

Rat HCC model was induced by diethylnitrosamine, and then the model was administered with two doses of FZXZP, high and low. Sodium demethylcantharidate was used as a positive control. Subsequently, microarrays of circRNA, miRNA and mRNA were performed on the blank, model, high and low dose groups, respectively, and the competitive binding mechanisms among them were further analyzed by bioinformatics. Then, the circRNA-miRNA-mRNA networks were constructed to mine the targeted-RNAs of FZXZP in HCC, as well as to explore their potential regulatory mechanisms. Finally, functions and pathways of the FZXZP targeted genes in rat HCC were annotated with GO and KEGG, and qRT-PCR was performed to validate the accuracy of the above analyses in this study.

RESULTS

The results showed that FZXZP significantly inhibited the development and progression of HCC in rats, improved the pathological conditions and suppressed the proliferation of HCC cells. Subsequently, after a series of screening, the competing endogenous RNA networks (circRNA-miRNA-mRNA), consisting of 2 circRNAs, 7 miRNAs and 104 mRNAs, were finally established. KEGG and GO analyses of the networks revealed that lipid metabolism related pathways, such as fatty acid metabolism, bile secretion and PPAR pathway, were significantly enriched. In the further hubgene network analysis, in addition to lipid metabolism, aberrant glucose metabolism was found to be ameliorated by G6pc and Pklr in hubgenes. Finally, the qRT-PCR analyses confirmed that the expression tendencies of the above targeted genes were correct and believable in transcriptomic sequencings, and qRT-PCR results of the genes closely related to proliferation, invasion and apoptosis of HCC also indicated the inhibitory effects of FZXZP on HCC obviously.

CONCLUSION

FZXZP demonstrated significant anti-HCC effects through improving lipid and glucose metabolism, restoring the metabolic homeostasis of the liver via circRNA-miRNA-mRNA networks.

摘要

背景

肝细胞癌（HCC）因其高致死率而成为严重威胁人类健康的主要疾病。我们之前的研究表明，扶正消症方（FZXZP）作为一种有效的中药，对 HCC 具有显著的抑制作用。然而，其潜在机制仍不清楚。

目的

本研究从整体角度出发，通过转录组测序探讨 FZXZP 的抗 HCC 机制。

方法

采用二乙基亚硝胺诱导大鼠 HCC 模型，然后给予 FZXZP 高、低两种剂量治疗，阳性对照药为去甲斑蝥酸钠。分别对空白组、模型组、高剂量组和低剂量组进行 microarray 检测，对 circRNA、miRNA 和 mRNA 进行检测，然后通过生物信息学分析其竞争结合机制。进一步构建 circRNA-miRNA-mRNA 网络，挖掘 FZXZP 在 HCC 中的靶向 RNA，并探讨其潜在的调控机制。最后，对 FZXZP 靶向基因在大鼠 HCC 中的功能和通路进行注释，GO 和 KEGG 分析，采用 qRT-PCR 对上述分析在本研究中的准确性进行验证。

结果

结果表明，FZXZP 能显著抑制大鼠 HCC 的发生发展，改善病理状况，抑制 HCC 细胞增殖。随后，经过一系列筛选，最终建立了由 2 个 circRNA、7 个 miRNA 和 104 个 mRNA 组成的竞争性内源性 RNA 网络（circRNA-miRNA-mRNA）。对网络进行 KEGG 和 GO 分析发现，脂肪酸代谢、胆汁分泌和 PPAR 通路等脂质代谢相关通路显著富集。在进一步的 hubgene 网络分析中，除脂质代谢外，还发现 hubgenes 中的 G6pc 和 Pklr 改善了异常的葡萄糖代谢。最后，qRT-PCR 分析证实了上述靶向基因在转录组测序中的表达趋势是正确和可信的，与 HCC 增殖、侵袭和凋亡密切相关的基因 qRT-PCR 结果也表明 FZXZP 对 HCC 有明显的抑制作用。