基于网络药理学和体外实验分析莪术醇抗胶质瘤的作用机制

Mechanism of curcumol against glioma based on network pharmacology and in vitro experiments

目的:通过网络药理学和体外实验研究莪术醇抗胶质瘤的潜在靶点和作用机制,为其临床应用及科研提供理论依据。方法:利用TCMSP数据库和Swiss数据库筛选莪术醇潜在作用靶点;通过GeneCards数据库和OMIM数据库获取胶质瘤相关靶点,获得“莪术醇-胶质瘤”的交集靶点并绘制韦恩图,使用STRING数据库构建PPI网络并筛选出核心靶点,使用Metascape数据库进行GO与KEGG富集分析,进一步挖掘莪术醇抗胶质瘤的药理作用机制。最后,采用MTT法检测莪术醇对胶质瘤U87细胞的增殖影响及利用Western blot技术检测PI3K,p-AKT,AKT蛋白表达水平,进一步证实网络药理学分析的结果。结果:从莪术醇预测102个潜在作用靶点,获取胶质瘤疾病靶点4821个,莪术醇与胶质瘤一共有67个交集作用靶点,主要核心靶点为MAPK1、EGFR、MAPK8、MAPK14、JAK3。GO富集分析得到1003个生物过程,其中BF为854个、MF为72个、CC为77个,KEGG分析得到136条信号通路,其中PI3K/AKT通路等可能在抗胶质瘤中起关键作用。分子对接结果发现莪术醇与EGFR、JKA3、MAPK1、MAPK8、MAPK结合较好,且均具有能够自发结合成较为稳定的构像。实验证明随着浓度的增加,莪术醇对胶质瘤U87细胞增殖抑制活性明显增强,呈时间浓度依赖性。Western blot结果显示莪术醇能降低胶质瘤U87细胞的p-AKT蛋白的表达从而抑制PI3K/AKT通路(P<0.01)。结论:莪术醇治疗恶性胶质瘤具有多靶点、多通路的特点,其机制可能与调控p-AKT的蛋白表达有关。

Objective:To study the potential targets and mechanisms of curcumol against glioma using network pharmacology and in vitro experiments,and to provide theoretical basis for its clinical application and scientific research.Methods:TCMSP database and Swiss database were used to screen the potential targets,geneCards database and OMIM database were used to obtain glioma-related targets.The intersection targets of curcumol-glioma were obtained and the Wayne diagram was drawn.The PPI network was constructed using STRING database and the core targets were screened.Metascape database was used for GO and KEGG enrichment analysis to further explore the pharmacological mechanism of curcumol against glioma.Finally,MTT method was used to detect the proliferation effect of curcumol on glioma U87 cells,and western blot was used to detect the expression levels of PI3K,p-AKT and AKT proteins,to further confirm the results of network pharmacology analysis.Results:A total of 4821 glioma targets were obtained from 102 potential targets predicted by curcumol.There were 67 intersection targets between curcumol and glioma,and the main core targets were MAPK1,EGFR,MAPK8,MAPK14 and JAK3.GO enrichment analysis showed 1003 biological processes,including 854 BFs,72 MFs,and 77 CCs.KEGG analysis showed 136 signaling pathways,among which PI3K/AKT pathway may play a key role in anti-glioma.Molecular docking showed that zedoaryl could spontaneously combine with EGFR,JKA3,MAPK1,MAPK8 and MAPK to form a relatively stable conformation.The results showed that the inhibitory activity of curcumol on the proliferation of glioma U87 cells was significantly enhanced in a time-and concentration-dependent manner with the increase of concentration.WB results showed that curcumol could reduce the expression of p-AKT protein in glioma U87 cells to inhibit the PI3K/AKT pathway(P<0.01).Conclusion:Curcumol could inhibit malignant glioma with multiple targets and multiple pathways,and its mechanism may be related to the regulation of p-AKT protein expression.