Effects Of Antisense Oligonucleotide Targeting Hsa-miR-21on The Growth And Sensitivity To Chemoradiotherapy Of Human Colorectal Cancer HT-29Cell

Objective:To observe lentivirus-mediated miR-21antisense nucleotide chain loweredcolorectal cancer HT-29cells express how to impact the biological behavior ofcolorectal cancer cells and radiotherapy and chemotherapy sensitivity.Discuss miR-21to become a new target for the possibility of colorectal cancer targeted therapy.Methods:MicroRNA-21genomic sequence was amplified by PCR and cloned intoLentiviral vector construct with miR-21antisense oligonucleotide chain and by viraltransfection inhibition of miR-21function and expression, screening targeted themiR-21ASO fragment into colorectal cancer HT-29cells stably transfected cell linesas experimental group, and the other normal HT-29cells transfected with no loadlentivirus group,transfection U6snRNA group as the control group,with a probemethod PCR detection of the groups HT-29cells,miR-21expression differences,direct cell counting method cell proliferation was detected by flow cytometrycellcycle and apoptosis, cell colony formation assay cell clones Transwell invasion assaycell invasion, MTT assay different gradient of drug concentration in the tumorinhibition rate, as well as by the total10Gy X growth inhibition rate of tumor cellsafter irradiation.Results:1.Transfected with miR-21ASO treated group compared with HT-29cells groupmiR-21levels decreased significantly,a significant difference(P<0.01),there was nosignificant difference between no-load Lentiviral transfection group and U6positivecontrol group and the normal group of miR-21.2.Counting the number of cells by direct cell counting method for7days,the cellgrowth curve,cell proliferation rate of the experimental group was significantly lowerthan other control cells(P<0.01),no significant difference between the groups and thecontrol group. 3.The flow cytometry results showed that transfection cells the miR-21ASOafter early apoptosis rate compared with the control group increased significantly,theG0/G1share percentage of the total number of cells increased, while the decline inthe percentage of cells in S phase.4.Cell colony formation assay display HT-29/ASO group of colony formationwas significantly lower than the HT-29group,HT-29/NC group and HT-29/U6ofHT-29group,HT-29/NC group and the formation rate of of the cell clones in thebetween the the HT-29/U6groups was no significant difference,5.Cell invasion assay in the experimental group transfected with miR-21ASOcell invasion was significantly lower than the control group,lowered the expression ofmiR-21ASO group through the least number of cells in the Matrigel membrane,andthe other group, the difference was statistically significant(P<0.01),NC and U6lentivirus group with non-transfected HT-29,no significant difference(P>0.05).6.MTT assay cut miR-21HT-29cells to5-FU drug sensitivity experiments showthat different drug concentrations of5-FU the ASO group of cell growth inhibitionwas significantly higher than the control cell growth inhibition rate, the differencesignificant(P<0.05),the ASO group of sensitizing multiple of2.35times.7.MTT assay cut miR-21HT-29cells to radio therapy sensitivity experiments,results showed that10Gy total X-ray irradiation after HT-29/ASO tumor inhibitionrate was significantly lower than HT-29,HT-29/NC HT-29/U6group, while nosignificant difference between the control groups.Conclution:Lentivirus-mediated miR-21ASO intransfected HT-29cells,cell proliferation,invasion,cloning diminished capacity,increased sensitivity to radiotherapy andchemotherapy,apoptosis enhancement.miR-21may be the growth of colorectal cancerinvasion and metastasis and chemotherapy sensitivity plays a role ininhibition ofcancer development, there may be a new target for colorectal cancer treatment.