Effect Of HIF-1a Gene Sliencing On The Growth Of Human Lung Adenocarcinoma Cell A549in Nude Mice And Survivin Expression

0bjective:To observe the effect of HIF-1α gene silencing on the growth of human lungadenocarcinoma cell A549in nude mice and survivin expression,in order to further tostudy the regulation role of survivinMethods:Human lung cancer cell1ine A549cells were divided into3groups:control group,control group transfected with scrambled sequence plasmid and HIF-1α-MiRNA grouptransfected with HIF-1α-MiRNA eukaryotic expression plasmid.Cultured cells of thethree groups were inoculated in nude mice to establish lung cancer-bearing nude miceeight in every group, Tumor’s dimensions of three groups were measured Per three days andtumor growth curve were maked.After58days,nude mice were killed and tumors wereweighed,the anti-tumor effect was reckoned, HIF-1α MiRNA assay was performed toevaluate the tumor-suppressive effect of HIF-1α silencing on lung cancer-bearing nudemice. RT-PCR was adopted to detect the mRNA expression of HIF-α and survivin.Immunohistochemistry and Western blot assay was done to detect the protein expressionof HIF-1α and Survivin. TUNEL stainin was used to detect apoptosis in tumor tissue.Results:1. Tumor formation rate in nude mice is100%. Subcutaneous Nodules could be touchedafter inoculation4-5days in control group, control group transfected with scrambledsequence plasmid and6-7days in HIF-1α-MiRNA group, the standard(about50mm3)was achieved after10days, The tumor growth in HIF-1α-MiRNA group wassignificantly slower than that in control groups and control group transfected withscrambled sequence plasmid on the25th day after transplantation(P all <0.05),thedifference is much significant after34th days.0n the58th day after transplantation.all nudemice were killed,the tumor weight in the HIF-1α-MiRNA group was (1.14±0.08) g,significantly lower than in the control group transfected with scrambled sequenceplasmid (1.71±0.18)g in the mock group and control group without handed(1.75±0.26)g (P all <0.05). 2. the levels of HIF-1α mRNA in the HIF-1α-MiRNA group detected with RT-PCRwere0.10±0.01, significantly lower than in the control group transfected with scrambledsequence plasmid (0.72±0.04) and control group without handed (0.74±0.03)(Pall<0.05).3. the levels of HIF-1α protein in the HIF-1α-MiRNA group were (20.13±3.15)%and0.53±0.09, significantly lower than in the control group transfected with scrambledsequence plasmid [(66.55±3.55)%and (1.64±0.19)] and control group withouthanded[(69.05±5.32)%and (1.76±0.09)](P all<0.05).4. the levels of survivin protein in the HIF-1α-MiRNA group were (22.18±3.12)%and0.50±0.06, significantly lower than in the control group transfected with scrambledsequence plasmid [(68.53±4.10)%and (1.02±0.09)]and control group withouthanded[(70.35±5.37)%and (1.07±0.08)](P all<0.05).5. the number of apoptotic cells in the HIF-1α-MiRNA group was (22.52±2.81)%,significantly higher than in the control group transfected with scrambled (P all<0.01).Conclusion:1. HIF-1α MiRNA can significant inhibits the growth of A549cell xenograft in nudemice,increases tumor cell apoptosis;2. survivin mRNA and protein expressions were reduced in the HIF-1α-MiRNA group,HIF-1α MiRNA may through downregulate its downstream gene survivin expression,therefore,to promote lung cancer cell apoptosis and exert a tumor–suppressing effect invivo．...