Effect of ultrasound-mediated triplex-forming oligonucleotide lipid ultrasonic microbubble on cellular absorption rate of TFO and expression of tissue factor / 解放军医学杂志

ABSTRACT

Objective To investigate the effect of lipid ultrasonic microbubble on the cellular absorption rate of antiparallel phosphorothioate triplex-forming oligonucleotide(TFO) and the expression of the tissue factor(TF) in endothelial cells.Methods The GT21-apsTFO,specific to the gene sequence of the shearing stress response element,was synthesized and the lipid ultrasonic microbubble carrying TFO was constructed.The ECV304 cells were divided into 4 groupscontrol group(SSRE),received shear stress only for 6h;TFO+ ultrasonic irradiation group(TFO+U),treated with 60?l TFO in final concentration of 0.2?mol/L and ultrasonic irradiation for 30s simultaneously;TFO-lipid microbubble group(TFO-M),treated with 60?l lipid microbubble loaded with TFO in final concentration of 0.2?mol/L;and TFO-lipid microbubble + ultrasonic irradiation group(TFO-M+U),treated with 60?l lipid microbubble loaded with TFO(final concentration was 0.2?mol/L) and ultrasonic irradiation for 30s simultaneously.Cells in the last 3 groups were treated with shear stress for 6h with pressure of 1.2Pa 24h after transfection.The fluorescent microscope was used to observe the cellular absorption rate of TFO.The immunofluorescence method and RT-PCR were employed to determine the protein and mRNA expressions of TF.Results The transfection efficiency of the TFO was higher in TFO-M+U group(38.83%?6.52%) than in TFO-M group(9.50%?2.88%) and TFO+U group(12.66%?3.01%,P