参桃软肝方通过 E2F2-p53 通路促进肝癌细胞凋亡。

Shentao Ruangan formula promotes apoptosis via the E2F2-p53 pathway in hepatocellular carcinoma.

机构信息

School of Traditional Chinese Medicine, Southern Medical University, Guangzhou 510515, China; The First School of Clinical Medicine, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong 510405, China.

Artemisinin Research Center, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong 510405, China.

出版信息

Phytomedicine. 2023 Jan;109:154565. doi: 10.1016/j.phymed.2022.154565. Epub 2022 Nov 22.

DOI:10.1016/j.phymed.2022.154565

PMID:36610125

Abstract

BACKGROUND

Hepatocellular carcinoma (HCC) is a malignant tumor with high morbidity and mortality rates. E2F2 is an independent predictor of poor prognosis in HCC; however, The mechanism by which E2F2 promotes the progression of HCC remains unclear. The Shentao Ruangan (STR) formula exhibits antitumor efficacy against HCC; however, the underlying antitumor mechanisms remain unknown.

PURPOSE

To explore the regulatory effect of E2F2 on the p53 signaling pathway and reveal the role and mechanism of STR in promoting cell apoptosis via the E2F2-p53 signaling pathway in HCC.

METHODS

E2F2 overexpression or silencing by lentivirus in HepG2 cells were used to explore their influence on apoptosis and the p53 pathway. An H22 tumor-bearing mice model was used to determine the therapeutic efficacy of STR and its effects on the E2F2-p53 pathway. STR-mediated serum (STR-MS) was prepared, and its chemical constituents were identified using mass spectrometry. The effects of STR-MS on viability and apoptosis of HepG2 cells and the E2F2-p53 pathway were investigated and validated using rescue experiments.

RESULTS

E2F2 overexpression significantly inhibited apoptosis and the p53 pathway in HepG2 cells, whereas E2F2-silenced HepG2 cells showed the reverse. This increased apoptosis was rescued by the addition of a p53 inhibitor (PFT-α) to E2F2-silenced HepG2 cells. In vivo, high doses of STR could remarkably inhibit the growth of xenografts, promote the apoptosis of hepatoma cells, downregulate E2F2, and activate the p53-dependent mitochondrial apoptotic pathway with good safety. In vitro, STR-MS exhibited similar effectiveness, and the best effect was achieved at 30% STR-MS concentration for 48 h. When 30% STR-MS was added to E2F2-overexpressing cells, the increased apoptosis and expression of key proteins in the p53-dependent mitochondrial apoptosis pathway were significantly rescued.

CONCLUSION

Our findings demonstrate, for the first time, that E2F2 inhibits hepatoma cell apoptosis in a p53-dependent manner and that STR may promote apoptosis by regulating the E2F2-p53 pathway in HCC.

摘要

背景

肝细胞癌（HCC）是一种发病率和死亡率都很高的恶性肿瘤。E2F2 是 HCC 预后不良的独立预测因子；然而，E2F2 促进 HCC 进展的机制尚不清楚。参桃软肝方（STR）对 HCC 具有抗肿瘤作用；然而，其潜在的抗肿瘤机制尚不清楚。

目的

探讨 E2F2 对 p53 信号通路的调控作用，并揭示 STR 通过 E2F2-p53 信号通路促进 HCC 细胞凋亡的作用和机制。

方法

采用慢病毒转染 HepG2 细胞过表达或沉默 E2F2，探讨其对细胞凋亡及 p53 通路的影响。构建 H22 荷瘤小鼠模型，观察 STR 的治疗效果及其对 E2F2-p53 通路的影响。制备 STR 介导的血清（STR-MS），并采用质谱鉴定其化学成分。采用 rescue 实验研究 STR-MS 对 HepG2 细胞活力和凋亡及 E2F2-p53 通路的影响，并进行验证。

结果

E2F2 过表达显著抑制 HepG2 细胞凋亡及 p53 通路，沉默 E2F2 的 HepG2 细胞则出现相反作用，E2F2 沉默的 HepG2 细胞加入 p53 抑制剂（PFT-α）后可部分挽救这种促凋亡作用。体内实验结果显示，高剂量 STR 可显著抑制异种移植瘤生长，促进肝癌细胞凋亡，下调 E2F2，激活 p53 依赖性线粒体凋亡通路，且安全性良好。体外实验结果显示，STR-MS 也具有类似作用，其中 30%STR-MS 作用 48 h 效果最佳。当向 E2F2 过表达的细胞中加入 30%STR-MS 时，E2F2 过表达导致的细胞凋亡增加和 p53 依赖性线粒体凋亡通路中关键蛋白的表达上调均得到明显挽救。