Pan Yating, Liao Xinyi, Yang Lili, Zhang Chunlei, Wang Jue, Zheng Peiyong, Yu Guanzhen, Song Haiyan
Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China.
Department of Oncology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China.
Front Pharmacol. 2022 Jun 30;13:900128. doi: 10.3389/fphar.2022.900128. eCollection 2022.
The extract of (Roxb.) Moon [Apocynaceae] (MTE) has shown a significant anti-cancer effect on hepatocellular carcinoma (HCC), but its mechanism remains unclear. In this study, we used transcriptomics methods to investigate the underlying mechanism of MTE against HCC. Both MHCC97H and HepG2 cell lines were treated with MTE. The cell viability and migration were measured using the cell counting kit-8 assay and transwell assay. RNA-sequencing was used to identify differentially expressed genes (DEGs) between HepG2 cells treated with and without MTE. The expression levels of selected DEGs-vascular endothelial growth factor-A (VEGFA), platelet-derived growth factor receptor-β (PDGFRB), and von Willebrand factor (VWF)-were verified by RT-PCR and Western blot. The effect of conditioned medium from HCC cells with MTE treatment (CM-MTE) on blood vessels was observed by tube formation assay of HUVECs and chick chorioallantoic membrane (CAM) assay. A mouse model of HCC patient-derived tumor xenograft (PDX) was established and treated with MTE. The effect of MTE on the growth and angiogenesis of HCC-PDX was analyzed. The results demonstrated that MTE inhibited the viability and migration of HCC cells. RNA-seq showed that MTE treatment downregulated multiple genes associated with metabolism and angiogenesis. The expression levels of VEGFA, VWF, PDGFB, and PDGFRB in HCC cells were significantly suppressed by MTE. Meanwhile, MTE effectively inhibited the tube-forming capability of HUVECs and the angiogenesis of chick CAM. experiments revealed that the extract reduced tumor volume, inhibited the proliferation of HCC cells, and expanded the necrotic area of the tumor. Immunohistochemical results showed that the expression levels of CD31, PDGFB, VEGF, VWF, and PDGFRB in the HCC-PDX tumor tissues were all downregulated by MTE in a dose-dependent manner. Taken together, MTE could inhibit angiogenesis by repressing the expression of VEGF, VWF, PDGF, and PDGFRB in HCC cells, a mechanism that may enable MTE to counter HCC development.
萝芙木（Rauvolfia serpentina (Roxb.) Moon）[夹竹桃科]提取物（MTE）已显示出对肝细胞癌（HCC）具有显著的抗癌作用，但其机制尚不清楚。在本研究中，我们使用转录组学方法来探究MTE抗HCC的潜在机制。用MTE处理MHCC97H和HepG2两种细胞系。使用细胞计数试剂盒-8法和Transwell法检测细胞活力和迁移能力。利用RNA测序来鉴定经MTE处理和未处理的HepG2细胞之间的差异表达基因（DEG）。通过RT-PCR和蛋白质免疫印迹法验证所选DEG——血管内皮生长因子-A（VEGFA）、血小板衍生生长因子受体-β（PDGFRB）和血管性血友病因子（VWF）的表达水平。通过人脐静脉内皮细胞（HUVEC）的管腔形成试验和鸡胚绒毛尿囊膜（CAM）试验观察经MTE处理的肝癌细胞条件培养基（CM-MTE）对血管的影响。建立肝癌患者来源的肿瘤异种移植（PDX）小鼠模型并用MTE进行处理。分析MTE对HCC-PDX生长和血管生成的影响。结果表明，MTE抑制肝癌细胞的活力和迁移。RNA测序显示，MTE处理下调了多个与代谢和血管生成相关的基因。MTE显著抑制肝癌细胞中VEGFA、VWF、血小板衍生生长因子B（PDGFB）和PDGFRB的表达水平。同时，MTE有效抑制HUVEC的管腔形成能力和鸡胚CAM的血管生成。体内实验表明，该提取物减小了肿瘤体积，抑制了肝癌细胞的增殖，并扩大了肿瘤的坏死区域。免疫组化结果显示，MTE以剂量依赖的方式下调HCC-PDX肿瘤组织中CD31、PDGFB、VEGF、VWF和PDGFRB的表达水平。综上所述，MTE可通过抑制肝癌细胞中VEGF、VWF、血小板衍生生长因子（PDGF）和PDGFRB的表达来抑制血管生成，这一机制可能使MTE能够对抗HCC的发展。
