Objective: To study the safety and efficiency of the transfection of antisense oligonucletide into kidney mediated by lipid microbubbles, and to evaluate its potential clinical application.
Methods: The potential and conditions regarding the transfection self-made lipid microbubbles (CY5)-labeled-oligonucleotide (ODN) or CY5-labeled-ODN connective tissue growth factor (CTGF) into the rat kidney were evaluated. Th e safety was evaluated by HE staining, liver and renal function tests. The transfection efficiency was evaluated by fluorescence microscopy. Th e expression of CTGF was detected by RT-PCR and Western blot.
Results: Self-made lipid microbubble and/or ultrasound significantly enhanced the efficiency of gene transfer and expression in the kidney. Especially, 85%-90% of total glomerular could be transfected. CY5-labeled-ODN expression could be observed in glomerular, tubular and interstitial area. Th ere was no significant change in blood tests aft er gene transfer. Levels of LDH in 7 days were decreased compared with that at the fi rst day aft er the transfection (P<0.05). CTGF expression was successfully suppressed by transfection of CTGF-antisense-ODN into kidney.
Conclusion: The ultrasound-mediated gene transfer by self-made lipid microbubble could enhance the efficiency of ODN and expression in the rat kidney. Th is self-made lipid microbubbles supplement may be use for transfection of target genes.
目的：探讨超声联合自制脂质体微泡(lipid contrast agent，LCA)转染肾脏的条件和效率。方法：HE染色、肝肾功能检测评价其安全性；荧光显微镜评价其转染效率；RT-PCR和Western印迹检测其对结缔组织生长因子(connective tissue growth factor，CTGF)表达的影响。结果：超声联合LCA显著增强基因在肾脏内的转导效率，CY5标记的寡核苷酸主要表达在85%~90%的肾小球、肾小管和间质区；血液生化指标无明显改变；转染后第7天的LDH水平均低于0天(P<0.05)；转染至肾脏的CTGF反义寡核苷酸成功抑制了CTGF的表达。结论：超声联合LCA的基因转移系统显著增强基因转移和表达的效率，该方法可以用来转导其他外源性的基因。.