Research Of Influence On The Biological Characteristics Of Breast Cancer Cell Line MDA-MB-231 With Silencing LOX By SiRNA Interference

Objective:To clone LOX gene and inspect LOX gene quantitive expression from low invasion breast cancer cell MCF-7 and high invasion/metastic breastcell MDA-MB-231.Meanwhile to study mechanism how LOX participated in invasional and metastatic complex biological behaviour of breast cancer, by investigating the change of invasion,movement and prolifertion of high invasion/metastic breast cancer MDA-MB-231 with silencing LOX by siRNA interferenceMethods:After LOX gene cloned by RT-PCR was sequenced,inspect quantitive expression of MCF-7 and MDA-MB-231 by the method of Real-time PCR SYBR Green I.LOX siRNA interference sequence designed and syntheticed transiently transfect MDA-MB-231 cell,inspection interference efficiency with RT-PCR,RealtimePCR and cells immunofluorescence, screen the best siRNA in interference efficiency. Seting up Blank group, negative control group and experimental group that eath group have two equal holes,then start experiment as Transwell invasional experiment, Scarification experiment and CCK-8 proliferation experiment, respectively in transfection 24h,48h and 72h, to observe the influence of invasional and proliferation biological behaviour in mda-mb-231 after LOX gene interference.Result:LOX gene cloned by RT-PCR was confirmed concordance to targent gene from sequencing, and breast cell MDA-MB-231 expressed higher LOX gene,which compared with MCF-7.After transient transfection of LOX siRNA, the expression of LOXmRNA descends obviously in transfection 48h. after interfering the LOX gene expression.The results of movement experience in vitro (Transwell) demonstrates that the numbers of MDA-MB-231 cuting through microporous membrane in RNAi group,Mock group and Con group is respectively 50±2,48±4 and 30±1.In scarification experiments the picture and migration lenth in Oh,6h,12h and 24h whichexecutes after transfection 48h,demonstrate that RNAi group comparing with Mock group and Con group, its capacity of migration and locomotory decreases obviously.CCK-8 cell proliferation experiment demonstrate the cell growth inhibition rates of RNAi group in transfection 24h,48h,72h and 96h later respectively are 30.5%,40.8%,50.4% and 36.95%.while the cells of Mock group and Congroup have no inhibition obviously. So RNAi group comparing with blank group and Mock group the capicity of cell proliferation suffers inhibition obviously,which has statistical significance (p<0.05).Conclusion:First, the experiments certified that high invasion/metastic breast cell MDA-MB-231 has a higher expression of LOX gene.Second, after inferencing the expression of LOX gene of breast adenoma cell MDA-MB-231, the invasional and lomotory capacity decreased apparently, and similarly LOX gene participate on cellular proliferation of MDA-MB-231,the capicity of cellular proliferation of MDA-MB-231 attenuated obviously with silencing LOX by siRNA interference.