基于网络药理学及实验验证探讨山慈菇-漏芦药对治疗乳腺癌作用机制  

Mechanism of Cremastrae Pseudobulbus-Rhapontici Radix Against Breast Cancer:An Exploration Based on Network Pharmacology and Experimental Verification

作　　者：王冰[1] 王玥[1] 朱仲康 崔勇[1] 于嵩 刘海兴[1] 王旭[1] WANG Bing;WANG Yue;ZHU Zhong-kang;CUI Yong;YU Song;LIU Hai-xing;WANG Xu(Liaoning University of Traditional Chinese Medicine,Shenyang 110847,China;Shenyang Sports University,Shengyang 110105,China)

机构地区：[1]辽宁中医药大学,沈阳110847 [2]沈阳体育学院,沈阳110115

出　　处：《中国实验方剂学杂志》2022年第1期65-71,共7页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：辽宁省自然科学基金计划重点项目(20180540105)。

摘　　要：目的:应用网络药理学的研究方法筛选山慈菇-漏芦药对治疗乳腺癌的作用靶点及相关信号通路,通过体外实验明确其作用机制。方法:通过运用中药系统药理学分析平台(TCMSP)数据库检索获取山慈菇-漏芦的主要化学成分及相关作用靶点;使用GendCards筛选乳腺癌疾病的相关靶标基因;通过Venn软件筛选出药物与疾病的共同作用靶点;构建出药物-疾病的靶点网络,蛋白质-蛋白质相互作用(PPI)网络构建;通过基因本体(GO)富集和京都基因与基因组百科全书(KEGG)通路富集对有效作用靶点进行分析;通过细胞增殖与活性检测(CCK-8)实验、流式细胞术检测细胞凋亡实验、蛋白免疫印迹法(Western blot)实验进行进一步的实验验证。结果:通过筛选得到山慈菇-漏芦有效化学成分7个,有效作用靶点61个,其中作用于乳腺癌的相关靶点55个。通过GO富集分析共富集到条目832个,在生物学过程方面集中体现。KEGG通路富集分析共富集到85个条目,包括肿瘤抑制基因信号通路p53,血管内皮生长因子(VEGF)信号通路,人表皮生长因子受体(EGFR)信号通路、磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/Akt)信号通路等。通过体外实验验证了山慈菇-漏芦醇提取物可抑制人乳腺癌MDAMB-231细胞增殖,并诱导细胞的凋亡,与空白组,二甲基亚砜(DMSO)组(0.1%溶剂组)相比较,给药组MDA-MB-231细胞吸光度显著降低(P<0.01),凋亡率显著增高(P<0.01),Western blot实验结果表示,与空白组和DMSO组相比较,各给药组细胞中磷酸化(p)-PI3K/PI3K,p-Akt/Akt比值均明显降低(P<0.05)。结论:山慈菇-漏芦醇提取物能有效抑制人乳腺癌MDA-MB-231细胞增殖,并可以诱导其凋亡,作用机制可能与抑制PI3K/Akt信号通路活化相关。Objective:To screen out the main targets and related signaling pathways of the herbal pair Cremastrae Pseudobulbus-Rhapontici Radix in treating breast cancer based on network pharmacology and verify their action mechanism in in vitro experiments. Method: The main chemical components and related targets of Cremastrae Pseudobulbus-Rhapontici Radix were retrieved from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),and the target genes related to breast cancer from GeneCards. Following the screening of the common targets of Cremastrae Pseudobulbus-Rhapontici Radix and breast cancer using Venn,the Cremastrae Pseudobulbus-Rhapontici Radix-breast cancer network and proteinprotein interaction(PPI)network were constructed. The effective targets were then subjected to gene ontology(GO)and Kyoto encyclopedia of genes and genomes(KEGG)pathway enrichment analysis. The resulting outcomes were then verified by cell counting kit(CCK)-8 assay,flow cytometry,and Western blot. Result:The screening yielded seven effective components and 61 targets of Cremastrae Pseudobulbus-Rhapontici Radix,among which 55 targets were involved in breast cancer. The GO analysis revealed 832 entries,which were mainly enriched in the biological processes. According to KEGG pathway enrichment analysis,85 signaling pathways were obtained, including tumor suppressor p53, vascular endothelial growth factor(VEGF),epidermal growth factor receptor(EGFR),and phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt). It was verified in in vitro experiments that the alcohol extract of Cremastrae Pseudobulbus-Rhapontici Radix inhibited the proliferation of human breast cancer MDA-MB-231 cells and induced their apoptosis. Compared with the blank control group and the dimethyl sulfoxide(DMSO,0.1% solvent)group,the medication groups exhibited obviously decreased absorbance in MDA-MB-231 cells(P<0.01)and increased apoptosis rate(P<0.01). The results of Western blot demonstrated that compared with the blank control group an

关 键 词：山慈菇 漏芦 网络药理学 乳腺癌 作用机制 药对 

分 类 号：R285[医药卫生—中药学] R289[医药卫生—中医学]