Research Of Effect Of PEX Gene-modified Glioma Stem Cells On Bioactivity

Objective：To separate and train glioma stem cells (glioma stem cells, GSCs) from C6glioma cell lines，purify appraisal to get reliable GSCs cell lines; To study the effects ofPEX gene on the GSCs of biological activities and micro environmental.Methods：（1） With the stick wall culture method training C6glioma cells, no serumsuspended separation and purification amplification monoclonal training GSCs,cellular immune fluorescent chemical dyeing GSCs identification.（2） Liposomes mediated pDsRed2-C1-PEX transfect rats C6glioma stem cells andfluorescence microscope Observate fluorescence, the expression of MMP-2andVEGFmRNA were detected by using RT-PCR.（3） Effects of PEX on GSCs growth were observed by water soluble tetrazoliumcell proliferation and cytotoxicity assay kit. Flow cytometric analysis detectionthe cell cycle changes of GSCs.Results：（1） Successfully separate and cultivate GSCs, monoclonal training GSCs sphericalgrowth, is not easy to blow, refraction sex is stronger, the cellular immunefluorescence staining identified tumor ball express CD133, its differentiationthe progeny of cells express NSE or GFAP, differentiated cells with neuronsand glia for similar cancer cells.（2） Successfully transfect PEX into GSCs, high expression of GSCs in vitro andthe proliferation of GSCs had significant inhibitory effect on the detectioninstrument, Flow cytometric analysis detection: The group which carrying PEX cell cycle G0/G1phase cell percentage rise is81.10%, S period cells obviouslydecreased was12.04%, differences have percentage of statistical significance(P <0.05).（3） Transgenic group the expression of the blank group for2.0times, for emptycarrier group of1.8times. Transgenic group and the rest two groups ofcomparisons difference was statistically significant (P <0.05), MMP-2andVEGFmRNA expression, the opposite is true, all is Transgenic groupexpression, and reduce the blank group, the empty carrier group comparingstatistical significance (P <0.05).Conclusion：（1） In C6glioma cell lines，we can separate and train glioma stem cells (gliomastem cells, GSCs)，PEX gene in C6glioma, and its expression of the stemcells in vitro can directly suppress the GSCs proliferation.（2） PEX genes on the GSCs significantly cut the expression of MMP-2andVEGF, can speculate PEX gene may be by suppressing the secretion ofthe GSCs MMPs way is inhibit or vasogenic GSCs way of proliferation andto influence their growth. Thus will gene therapy and GSCs this targetcombined, precisely targeted GSCs as an glioma treatments with effectivemeans.