背景:Tanacetumparthenium(L.)Schultz-Bip,通常被称为狂热者,传统上被用来治疗发烧,偏头痛,类风湿性关节炎,和癌症。小白菊内酯(PTL),从小白菊芽中分离出的主要生物活性成分,是一种具有抗炎和抗肿瘤特性的倍半萜内酯。以前的研究表明,PTL在各种癌症中发挥抗癌活性,包括肝癌,胆管癌,急性髓系白血病,乳房,前列腺,还有结直肠癌.然而,PTL抗癌作用的代谢机制尚不清楚。
目的:探讨PTL在人胆管癌细胞中的抗癌活性及其潜在机制。
方法:在这项调查中,通过基于液相色谱/质谱(LC/MS)的代谢组学方法研究了PTL对人胆管癌细胞的作用和机制.首先,使用细胞计数试剂盒-8(CCK-8)评估细胞增殖和凋亡,流式细胞术分析,和西方印迹。然后,已构建了基于LC/MS的代谢谱分析以及正交偏最小二乘判别分析(OPLS-DA),以区分TFK1细胞中阴性对照组和PTL处理组之间的代谢变化。接下来,应用酶联免疫吸附试验(ELISA)研究与显着警告的代谢物相关的代谢酶的变化。最后,与关键代谢酶相关的代谢网络,代谢物,代谢途径是使用MetaboAnalyst5.0和京都基因和基因组百科全书(KEGG)途径数据库建立的。
结果:PTL处理可以浓度依赖性地诱导TFK1的增殖抑制和凋亡。确定了43个与PTL抗肿瘤作用相关的潜在生物标志物,主要与谷氨酰胺和谷氨酸代谢有关,丙氨酸,天冬氨酸和谷氨酸代谢,苯丙氨酸,酪氨酸和色氨酸的生物合成,苯丙氨酸代谢,精氨酸生物合成,精氨酸和脯氨酸代谢,谷胱甘肽代谢,烟酸和烟酰胺代谢,嘧啶代谢,脂肪酸代谢,磷脂分解代谢,和鞘脂代谢。上游和下游代谢物的路径分析,我们发现了三种关键的代谢酶,包括谷氨酰胺酶(GLS),γ-谷氨酰转肽酶(GGT),和肉碱棕榈酰转移酶1(CPT1),主要参与谷氨酰胺和谷氨酸代谢,谷胱甘肽代谢,和脂肪酸代谢。与显著提醒的代谢物相关的代谢酶的变化与代谢物的水平一致。以及与关键代谢酶相关的代谢网络,代谢物,并建立了代谢途径。PTL可能通过干扰代谢途径发挥其对胆管癌的抗肿瘤作用。此外,我们选择了两种被认为是胆管癌治疗一线化疗标准的阳性对照药物,以验证我们的PTL代谢组学研究的可靠性和准确性.
结论:这项研究增强了我们对PTL治疗胆管癌细胞的代谢谱和机制的理解,为进一步研究其他药物的抗癌机制提供了参考。
BACKGROUND: Tanacetum parthenium (L.) Schultz-Bip, commonly known as feverfew, has been traditionally used to treat fever, migraines, rheumatoid arthritis, and cancer. Parthenolide (PTL), the main bioactive ingredient isolated from the shoots of feverfew, is a sesquiterpene lactone with anti-inflammatory and antitumor properties. Previous studies showed that PTL exerts anticancer activity in various cancers, including hepatoma, cholangiocarcinoma, acute myeloid leukemia, breast, prostate, and colorectal cancer. However, the metabolic mechanism underlying the anticancer effect of PTL remains poorly understood.
OBJECTIVE: To explore the anticancer activity and underlying mechanism of PTL in human cholangiocarcinoma cells.
METHODS: In this investigation, the effects and mechanisms of PTL on human cholangiocarcinoma cells were investigated via a liquid chromatography/mass spectrometry (LC/MS)-based metabolomics approach. First, cell proliferation and apoptosis were evaluated using cell counting kit-8 (CCK-8), flow cytometry analysis, and western blotting. Then, LC/MS-based metabolic profiling along with orthogonal partial least-squares discriminant analysis (OPLS-DA) has been constructed to distinguish the metabolic changes between the negative control group and the PTL-treated group in TFK1 cells. Next, enzyme-linked immunosorbent assay (ELISA) was applied to investigate the changes of metabolic enzymes associated with significantly alerted metabolites. Finally, the metabolic network related to key metabolic enzymes, metabolites, and metabolic pathways was established using MetaboAnalyst 5.0 and Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway Database.
RESULTS: PTL treatment could induce the proliferation inhibition and apoptosis of TFK1 in a concentration-dependent manner. Forty-three potential biomarkers associated with the antitumor effect of PTL were identified, which primarily related to glutamine and glutamate metabolism, alanine, aspartate and glutamate metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, phenylalanine metabolism, arginine biosynthesis, arginine and proline metabolism, glutathione metabolism, nicotinate and nicotinamide metabolism, pyrimidine metabolism, fatty acid metabolism, phospholipid catabolism, and sphingolipid metabolism. Pathway analysis of upstream and downstream metabolites, we found three key metabolic enzymes, including glutaminase (GLS), γ-glutamyl transpeptidase (GGT), and carnitine palmitoyltransferase 1 (CPT1), which mainly involved in glutamine and glutamate metabolism, glutathione metabolism, and fatty acid metabolism. The changes of metabolic enzymes associated with significantly alerted metabolites were consistent with the levels of metabolites, and the metabolic network related to key metabolic enzymes, metabolites, and metabolic pathways was established. PTL may exert its antitumor effect against cholangiocarcinoma by disturbing metabolic pathways. Furthermore, we selected two positive control agents that are considered as first-line chemotherapy standards in cholangiocarcinoma therapy to verify the reliability and accuracy of our metabolomic study on PTL.
CONCLUSIONS: This research enhanced our comprehension of the metabolic profiling and mechanism of PTL treatment on cholangiocarcinoma cells, which provided some references for further research into the anti-cancer mechanisms of other drugs.