Abstract
Objective To prepare myeloid cell leukemia-1 (Mcl-1 )small interfering RNA (siRNA)loaded niosomes and to observe its inhibitory effect on human hepatoma carcinoma HepG2 cells.Methods ①Preparation and functional i-dentification of Mcl-1 siRNA loaded niosomes:Ethanol injection method was employed to prepare blank niosomes,which were then separately incubated with FAM-siRNA and Mcl-1 siRNA to form FAM-siRNA and Mcl-1 siRNA loaded niosomes through electrostatic interaction for 30 min.Malvern apparatus was used to determine the average particle size and zeta po-tential of blank niosomes and siRNA loaded niosomes.The encapsulation efficiency and in vitro release profile of siRNA were determined by centrifugal ultrafiltration-fluorescence spectrophotometry method.HepG2 cells were divided into groups A,B,C,and D for 24-hour cultivation,then groups C and D were treated with FAM-siRNA loaded niosomes and FAM-siRNA loaded Lipofectamin,respectively.Group B was exposed to equivalent free FAM-siRNA,while group A was subjec-ted to no disposal.After 6-hour incubation,cellular uptake of FAM-siRNA in HepG2 cells was examined.②The inhibitory effect of transfection of Mcl-1 siRNA loaded niosomes on HepG2 cells:Cells in logarithmic phase were divided into groups 1,2,3,4,and 5 and were seeded at a density of 5 ×105 per well for cultivation.At 24 h,cells in the groups 1,2,3, and 4 were treated with free Mcl-1 siRNA,FAM-siRNA loaded niosomes,Mcl-1 siRNA loaded niosomes,and Mcl-1 siR-NA loaded Lipofectamin for 6 h,respectively.After another 72 h,Mcl-1 protein of cells in each group was detected. HepG2 cells were divided into groups a,b,c,d and the control group,and were treated as previously mentioned.The fi-nal concentration of siRNA of each group was at a gradient of 1,10,50,100,and 200 nmol/L.The growth of cells was examined and cell viability was calculated.Results The particle size of blank niosomes was smaller while its zeta poten-tial was larger as compared with FAM-siRNA loaded niosomes (all P<0.01).The encapsulation efficiency of FAM-siRNA was 82.3%±2.1%.The in vitro cumulative FAM-siRNA release rates of FAM-siRNA loaded noisomes at 1,3,6,12, 24,36,48,72,and 96 h were 13.5%±2.8%,26.1%±1.6%,38.0%±2.9%,50.6%±2.3%,56.8%±2.9%, 59.9%±2.3%,63.3%±2.0%,65.0%±2.7%,67.2%±2.9%,respectively.Compared with group B,cellular up-take of FAM-siRNA in groups C and D was better.The Mcl-1 protein expression levels of groups 1,2,3,4 and 5 were 102.0 ±4.9,103.8 ±8.3,25.2 ±3.7,29.4 ±6.4,and 96.1 ±5.8,respectively.Obviously,the level of group 3 was significantly less than that of groups 1,2,and 5 (all P<0.01).Cell viability of groups c and d was significantly higher that that of groups a and b,meanwhile,the cell viability of group d was higher than that of group c (all P<0.01).Con-clusion We successfully prepare Mcl-1 siRNA loaded niosomes and the expression of Mcl-1 protein is low expressed in HepG2 cells,and HepG2 cells are inhibited after transfection of Mcl-1 siRNA loaded niosomes.关键词
髓样细胞白血病1/小干扰RN A/RN A干扰/囊泡/肝癌/细胞存活率Key words
myeloid cell leukemia-1/small interfering RNA/RNA interference/niosomes/liver carcinoma/cell via-bility分类
医药卫生
