三氯生对离体鹅尾臀腺和人乳腺癌细胞脂肪酸合成酶的抑制作用

Inhibition of Triclosan to Fatty Acid Synthase from Goose Uropygial Glands and Human Breast Cancer Cells in Vitro

背景与目的:研究表明,人乳腺癌早期和病程中癌细胞脂肪酸合成酶(fattyacidsynthase,FAS)呈高效表达。抑制FAS可引起癌细胞凋亡。本研究采用酶促反应动力学实验观察三氯生对离体鹅尾臀腺FAS的影响,建立实验方法,研究三氯生对人乳腺癌SKBr3细胞FAS的抑制作用。方法:运用超速离心和SuperdexPG200层析技术分离纯化鹅尾臀腺FAS,用超速离心技术部分纯化人乳腺癌SKBr3细胞FAS。不同浓度三氯生与FAS相互作用不同时间后,加入底物,用分光光度法观察三氯生对FAS的抑制作用。结果:在鹅尾臀腺组,FAS被纯化为SDS-PAGE电泳单条区带(分子量250kDa),三种浓度三氯生(12.5、25.0、100.0μmol/L)与FAS在催化作用前相互作用0、5和10min,对FAS的抑制率分别为26.40%、28.30%和43.93%(12.5μmol/L);46.22%、50.28%和97.05%(25μmol/L);98.11%、97.75%和97.37%(100μmol/L)。在人乳腺癌SKBr3细胞组FAS被部分纯化,25、50、100和200μmol/L三氯生分别与FAS在催化前相互作用5min,对FAS活性的抑制率分别为20.00%、26.67%、60.00%和100.00%。结论:三氯生对离体鹅尾臀腺FAS和人乳腺癌SKBr3细胞FAS均具有抑制作用;作用强度既依赖于抑制剂浓度,又依赖于抑制剂与酶相互作用时间。

BACKGROUND &OBJECTIVE: It has been indicated that fatty acid synthase (FAS) is abnormally overexpressed in human breast cancer compared with normal human tissue. Inhibition of FAS induces apoptosis of human breast cancer cells. The aim of this study was to observe the inhibition of triclosan on FAS from goose uropygial glands for establishing the method and to study the inhibition of triclosan on FAS from human breast cancer SKBr3 cells in vitro. METHODS: The goose uropygial glands FAS was purified by ultra centrifugation and Superdex PG 200 chromatography; the human breast cancer SKBr3 cell FAS was partially purified by ultra centrifugation. The FAS was interacted with different concentrations of Triclosan with different times before catalyzing. Then the substrates of FAS were added to the reaction system. The inhibitory activities of triclosan against the FAS were investigated using spectrophotometric assays. RESULTS: In the goose uropygial gland group, FAS was purified as a single band at 250kDa with SDS PAGE. The inhibitory activities of triclosan(12.5 μmol/L) at 0, 5, and 10 minute on FAS were 26.40%, 28.30%, and 43.93%, respectively. The inhibitory activities of triclosan (25.00 μmol/L) at 0, 5, and 10 minute on FAS were 46.22%, 50.28%, and 97.05%, respectively. The inhibitory activities of triclosan (100.00 μmol/L) at 0, 5, and 10 minute on FAS were 98.11%, 97.75%, and 97.37%, respectively. In human SKBr3 breast cancer cell group, the inhibitory activities of triclosan (25, 50, 100, and 200 μmol/L) at 5 minute on FAS were 20.00%, 26.67%, 60.00%, and 100%, respectively. CONCLUSION: Triclosan inhibits the FAS from goose uropygial glands and human breast cancer SKBr3 cells. The inhibitory activities depended on the concentrations of triclosan and the interaction times between triclosan and FAS before catalyzing.