不同超声强度及微泡对基因和组织作用的实验研究

目的探讨不同强度超声破坏微泡对绿色荧光蛋白质粒(green fluorcscent protein，GFP)和小鼠骨骼肌组织的作用。方法分别用0．5w／cm^2、1．0w／cm^2、2．5w／cm^2的超声作用于基因及基因和微泡的混合物2min，琼脂糖凝胶电泳观察质粒基因的电泳图谱变化。并将昆明小白鼠16只分为4组，尾静脉输入白蛋白微泡，同时分别用(0．5w／cm^2、1．0W／cm^2、2．0W／cm^2、2．5w／cm^2的超声作用于小鼠骨骼肌2min．取局部组织HE染色观察超声破坏微泡后组织显微结构的变化。结果不同能量的超声和微泡作用后GFP质粒的电泳图谱结果无变化。0．5w／cm^2超声破坏微泡后无血管充血及红细胞渗出；1．0W／cm^2超声破坏微泡后可引起约30％血管充血，极少量红细胞渗出；2．0W／cm^2超声破坏微泡后可引起约60％血管充血，红细胞渗出较明显；2．5W／cm^2的超声破坏微泡后可使部分骨骼肌变性。结论用1．0W／cm^2和2．0W／cm^2超声作用2min后引起血管充血，红细胞渗出；2．5W／cm^2超声作用2min破坏微泡后可损伤组织。1．0～2．0W／cm^2超声强度破坏微泡后对GFP质粒无明显的损害。

Experimental study of effects of ultrasound and microbubble on gene and tissue

Objective To investigate the effects of ultrasound mediated microbubble destruction on green fluorescent protein(GFP) and on mouse skeletal muscles. Methods Ultrasound with different intensity of 0.5 W/cm 2, 1.0 W/cm 2, 2.5 W/cm 2 was applied on GFP plasmid or mixture of plasmid and microbubble for 2 minutes. Then agarose gel electrophoresis was used to analysis the effects of ultrasound and microbubble on plasmid. Ultrasound with the intensity of 0.5 W/cm 2, 1.0 W/cm 2, 2.0 W/cm 2, 2.5 W/cm 2 was applied on the skeletal muscles of mice in 4 groups for 2 minutes after intravenous injection of microbubble. Half hour after the experiment, the skeletal muscles were harvested for HE staining to observe the microstructure of the tissue. Results There was no difference in the results of agarose gel electrophoresis between pure plasmid and the plasmid irradiated by different energy of ultrasound. In vivo experiment, there was no vascular engorgement and red blood cells(RBC) exudation in 0.5 W/cm 2 group; In 1.0 W/cm 2 group, hyperemia in blood vessels and little RBC exudation could be seen; In 2.0 W/cm 2 group, significant hyperemia in blood vessels and RBC exudation could be observed; In 2.5 W/cm2 group,the lesion of skeletal muscles could be seen. Conclusions Different energy of ultrasound mediated microbubble destruction had no effects on the agarose gel electrophoresis of plasmid, and ultrasound with the frequency of 1 MHz, intensity of 1.0 - 2.0 W/cm 2 is suitable for gene delivery.