CAJ | 학술논문

目的探讨姜黄素对鼻咽癌细胞回复引导半胱氨酸丰富蛋白含kazal基元( RECK)基因甲基化以及基质金属蛋白酶9(MMP-9)表达的影响。方法体外培养鼻咽癌细胞系CNE-1,用1、10、30μmol/L姜黄素处理后,采用Western blot和实时定量PCR分别检测RECK、MMP-9蛋白和mRNA表达；高效液相色谱-电喷雾质谱检测RECK甲基化；MTT法检测CNE-1细胞的增殖。同时采用明胶酶谱实验观察姜黄素处理前后MMP-9酶活性变化。结果 CNE-1细胞未刺激时RECK表达水平较低,而经1~30μmol/L姜黄素处理后,能显著增强RECK蛋白和mRNA的表达。高效液相色谱-电喷雾质谱结果显示,30μmol/L姜黄素处理后,RECK启动子、全基因组以及细胞核内甲基化水平分别降低为(69.04依10.62)%、(61.13依7.08)%、2.80依1.32。同时,姜黄素能显著降低CNE-1细胞中MMP-9蛋白和mRNA表达以及MMP-9的酶活性。结论姜黄素可能通过能上调RECK基因表达,降低细胞内甲基化水平,从而抑制MMP-9的表达与活性而发挥对CNE-1细胞生长抑制作用。
목적탐토강황소대비인암세포회복인도반광안산봉부단백함kazal기원( RECK)기인갑기화이급기질금속단백매9(MMP-9)표체적영향。방법체외배양비인암세포계CNE-1,용1、10、30μmol/L강황소처리후,채용Western blot화실시정량PCR분별검측RECK、MMP-9단백화mRNA표체；고효액상색보-전분무질보검측RECK갑기화；MTT법검측CNE-1세포적증식。동시채용명효매보실험관찰강황소처리전후MMP-9매활성변화。결과 CNE-1세포미자격시RECK표체수평교저,이경1~30μmol/L강황소처리후,능현저증강RECK단백화mRNA적표체。고효액상색보-전분무질보결과현시,30μmol/L강황소처리후,RECK계동자、전기인조이급세포핵내갑기화수평분별강저위(69.04의10.62)%、(61.13의7.08)%、2.80의1.32。동시,강황소능현저강저CNE-1세포중MMP-9단백화mRNA표체이급MMP-9적매활성。결론강황소가능통과능상조RECK기인표체,강저세포내갑기화수평,종이억제MMP-9적표체여활성이발휘대CNE-1세포생장억제작용。
Objective To investigate the effect of curcumin on reversion-inducing-cysteine-rich protein with kazal motifs (RECK) gene methylation,and expression of matrix metalloproteinase-9 (MMP-9) in nasopharyngeal carcinoma cells. Methods Nasopharyngeal carcinoma cell line CNE-1 was cultured in vitro,and stimulated by 1,10 and 30 μmol/L curcumin for 48h,expression of RECK and MMP-9 were detected by Western blot and real-time PCR,respectively. RECK methylation was determined by HPLC chromatographic and mass spectrometric methods. Cell proliferation was assessed by MTT assay. Expression and the enzymic activity of MMP-9 were detected by Western blot and Gelatin zymography assay,re-spectively. Results The expression level of RECK was very low in unstimulated cells,and 1~30μmol/L could decrease the protein and mRNA level. HPLC chromatographic and mass spectrometric analysis demonstrated that 30 μmol/L could decrease level of the promoter methylation, global DNA methylation and the methylation activity of the nuclear extract to (69.04% ±10.62)%、(61.13±7.08)%、2.80±1.32,respectively,as compared to untreated cells. In addition,curcumin could also decrease the protein and mRNA level of MMP-9, and enzymic activity. Conclusion Cucrcumin increase RECK gene expression and down-regulate its methylation,and then attenuate MMP-9 to inhibit CNE-1 cells growth.