Microbubble Mediating Ang-1 Gene Therapy Lower Limb Artery Ischemia By Using Ultrasound

ObjectiveTo discuss the efficacy of ultrasound microbubble contrast mediated angiopoietin--1 gene therapy for lower limb ischemia,and to provide an effective treatment for clinical treatment of lower limb ischemia.Methods(1) Set up an animal model:experimental animals for the health of 15 New Zealand white rabbits,randomly divided into three groups,to ligate left lower limb femoral artery and its branch,resulting in the left lower limb ischemia model.(2) Ultrasound microbubble contrast agents ready:microbubble contrast agent for lipid ultrasound contrast agent.(3) Plasmid preparation:Plasmid used in the experiment are pEGFP/Ang-1,Plasmid transformed and amplified through colibacillus.(4) Plasmid transfection:group A used plasmid mixture of microbubble contrast agent and pEGFP/Ang-1 for intramuscular injection,and at the same time ultrasound irradiated;group B used a simple plasmid pEGFP/Ang-1 for intramuscular injection;group C used normal saline for intramuscular injection. (5) Arterial contrast:abdominal aortic cannulation and visualization after 4 weeks,observed neovascularization and the status of collateral vessels at DSA machine.(6) Immunohistochemistry:cut adductor tissue,10%formalin-fixed, paraffin sections,Ang-1 immunohistochemical staining.(7) HE staining:to observe the neovascularization and capillary count, and statistical analysis by SPSS11.0 software.Results(1) Arterial contrast:the neovascularization and the status of collateral vessels of group A is more than group B in the rabbit lower limb ischemia; group A and group B is more than group C.(2) Immunohistochemistry:group A of muscle cells shows that Ang-1 immunoreactivity in the deep tan,group B intracytoplasmic have some tan, and the group C intracytoplasmic have less brown.(3) Neovascularization count:the neovascularization count of group A is more than group B in the rabbit lower limb ischemia,group A and group B are more than group C,and have statistical significance.Conclusion(1)The angiography of lower limb ischemia showed rich collateral vessels after gene therapy;(2) Transfected muscle cells has Ang-1 protein expression;...