Δ6-去饱和酶和C20-延伸酶抑制乳腺癌细胞MCF-7增殖的研究  

Δ6-desaturase and C20-elongase Can Inhibit Breast Cancer Cells MCF-7 Proliferation

作　　者：马晓磊 崔安芳 张向阳 孟圆 MA Xiao-Lei;CUI An-Fang;ZHANG Xiang-Yang;MENG Yuan(Jining Medical University,Basic Medical College,Institute of Biochemistry and Molecular Biology,Jining 272067,China)

机构地区：[1]济宁医学院基础医学院生物化学与分子生物学教研室,山东济宁272067

出　　处：《中国海洋大学学报（自然科学版）》2018年第A01期44-49,共6页Periodical of Ocean University of China

基　　金：山东省自然科学基金项目(ZR2013CL009)资助~~

摘　　要：为探讨外源基因Δ6-去饱和酶与C20-延伸酶基因对肿瘤细胞增殖的抑制作用,本研究选取了微拟球藻(Nannochloropsis oculata)作为研究对象,对其Δ6-去饱和酶与C20-延伸酶基因进行了逆转录PCR扩增,并将获得的上述2个基因的编码区与真核表达质粒pEGFP-C3构建重组表达质粒,转染乳腺癌细胞MCF-7,以pEGFP-C3为对照,培养48h后,在荧光显微镜下观察转染细胞中的GFP荧光信号以判断重组质粒表达情况,荧光实时定量PCR分析转染细胞中外源基因的转录水平,MTT比色法检测外源基因对MCF-7细胞的增殖抑制程度,高效气相色谱检测转染细胞中多不饱和脂肪酸(Polyunsaturated fatty acids,PUFAs)的含量变化。结果表明2个外源基因均可在MCF-7细胞中表达,并能有效地抑制MCF-7细胞的增殖,改变MCF-7细胞中2种PUFAs的相对含量,且n-3PUFAs与n-6PUFAs的比值升高。推测过表达的Δ6-去饱和酶与C20-延伸酶可能通过改变乳腺癌细胞MCF-7细胞膜中PUFAs的相对含量,达到抑制乳腺癌细胞MCF-7增殖的效果。Polyunsaturated fatty acids(PUFAs)played an important part in many human body life activity.Previous studies have shown that the ratio of n-3/n-6 PUFAs play a key role in maintaining cell stability and normal growth and the ideal n-3/n-6 PUFAs ratio should be between 1∶4 and 1∶1.n-3 PUFAs can inhibit the growth of tumors and cancer cells,while n-6 PUFAs have the opposite effect.Clinical trials have shown that continuous addition of n-3 PUFAs to the diet of colorectal and breast cancer patients is a good inhibitor of tumor development.To explore the mechanism of exogenous gene in tumor cell proliferation inhibition,Δ6-desaturase(Nanoc-D6)and C20-elongase(Nanoc-E)were cloned from Nannochloropsis oculata with the method of the reverse transcription PCR.Plasmid pEGFP-C3 was used to construct respectively 2 recombinant expression plasmid,pEGFP-C3-Nan-D6 and pEGFP-C3-Nan-E.Then recombinant expression plasmid transfected breast cancer cells MCF-7 with X-fect Polymer.pEGFP-C3 transfected MCF-7 cells was as control with the same treatment.Transfection Cells were cultured for 48 hours and used to analysis 4 projects.Fluorescence microscope observed eGFP fluorescent signal to determind recombinant expression plasmid transfection success.Real-time PCR was used to analysis 2 exogenous genes transcription levels.MTT was used to detect the degree of transfection cells proliferation inhibition.The determination of variation of PUFAs content in transfection cells with the method of Gas chromatography.Meantime,cells transfected pEGFP-C3 were used as control.The results proved that 2 exogenous genes can express separately in MCF-7 cells,and effectively restrain the transfection MCF-7 cells proliferation.The content of PUFAs in transfection MCF-7 cells was changed by 2 exogenous genes,especially n-3/n-6 PUFAs increased in transfection MCF-7 cells.The above evidence proved that Nanoc-D6 and Nanoc-E catalyst can occur and product can significantly change the volume inside MCF-7 cells.So it is suggested that 2 exogenous genes chan

关 键 词：Δ6-去饱和酶 C20-延伸酶 MCF-7 增殖 PUFAS 

分 类 号：R34[医药卫生—基础医学]