Ultrasound enhanced gene therapy: Development of ultrasonic methods of gene transfection for cardiovascular system.

点击翻译按钮获取中文摘要

Ultrasound-triggered microbubble destruction has been proposed as a means of targeting gene therapy to specific organs. Successful reporter gene expression using this approach has been reported. However, quantification of therapeutic gene products with this method has not been elucidated yet in cardiovascular cells or tissues. We examined the dose-response relations among the amount of gene, microbubble concentration and the duration of incubation using neonatal rat cardiomyocytes. We employed expression plasmid of Escheririchia Coli β-Galactosidase gene, expression plasmid of rat hepatocyte growth factor (HGF) gene containing CMV-β actin hybrid promoter, and cultured rat neonatal cardiomyocytes for in vitro experimental setups. We used continuous-wave Doppler ultrasound with frequency of 2.5MHz and maximal intensity of 0.5W/cm^2 because of clinical relevance.1.Repetitive exposure to ultrasound-triggered galactose/palmitic acid microbubble destruction may yield efficient transfection of genes to cardiomyocytes.2.Using catheter in the rat left ventricular cavity with precordial ultrasound irradiation, we examined the feasibility of luciferase gene transfection into the myocardium. Luciferase activity was detected only in the anterior myocardium, and the level of expression was similar to that in HUVEC cells for in vitro experiments for transfection, suggesting difficulty to transfect beyond the endothelial barrier.3.We examined the feasibility of luciferase gene transfection into the myocardium with precordial ultrasound irradiation in the setting of intravenous administration of gene and microbubble. We detected no expression of luciferase in the myocardium, and some expression in the liver.4.Taken together, we might as well (1)develop more specific gene targeting method for myocardial transfection with modifying enhancer/promoter ; and (2)improve microbubble shells with higher affinity for both plasmid-gene and myocardial cell membrane.