背景：超支化支链淀粉衍生物作为非病毒基因载体具有低毒性、转染率好的特点，但如何高效递送其进入机体内使有治疗作用基因成功表达的方式仍在探索中。 目的：探讨包裹绿色荧光蛋白的新型基因运载载体经改进颈动脉注射法到达大鼠脑缺血损伤区的表达情况。方法：取雄性SD大鼠制作大脑中动脉梗死模型，24 h后随机分为2组，对照组以尾静脉注射包裹绿色荧光蛋白的超支化支链淀粉衍生物，实验组以改进型颈内动脉注射包裹绿色荧光蛋白的超支化支链淀粉衍生物。7 d后处死大鼠，取出脑组织，qPCR和Western blot检测大鼠脑组织中绿色荧光蛋白的基因和蛋白水平，免疫荧光法观察脑组织冰冻切片中血管内壁附近绿色荧光蛋白的表达。 结果与结论：qPCR和Western blot检测结果均显示实验组大鼠脑组织中绿色荧光蛋白的表达量显著高于对照组(P<0.05)，免疫荧光观察结果显示绿色荧光蛋白在实验组血管内壁附近的表达高于对照组。在SD大鼠大脑中动脉梗死模型中，相比尾静脉注射法，改进型颈动脉注射法可显著促进脑缺血损伤区的超支化支链淀粉衍生物及绿色荧光蛋白在脑组织中表达的量。%BACKGROUND:Hyperbranched cationic amylopectin is a kind of nonviral gene vectors with low toxicity and good transfection efficiency. However, searching for more efficient methods to delivery it into the body and making the genes expressed are being explored. OBJECTIVE:To study the expression of DMAPA-Amp wrapped green fluorescent protein (GFP) transferred by modified carotid injection into cerebral ischemic area. METHODS:Male Sprague-Dawley rats subjected to middle cerebral artery infarction were randomly divided into two groups after 24 hours:experimental group (injected with GFP entrapped DMAPA-Amp via the internal carotid artery) and control group (injected with GFP entrapped DMAPA-Amp via the tail vein). These rats were put to death and their brain tissue was removed after 7 days. The expression of GFP was detected by quantitative PCR and western blot assay, and immunofluorescence staining was performed to detect the expression of GFP located near cerebrovascular endothelial cel s by frozen section. RESULTS AND CONCLUSION:Compared with the control group, the expression of GFP was much higher in the experimental group detected by quantitative PCR and western blot (P< 0.05). Additional y, the expression of GFP located near cerebrovascular endothelial cel s by frozen section was also higher than that in the control group. Modified carotid injection could significantly promote the expression of hyperbranched cationic amylopectin derivates and GFP in the brain tissue of Sprague-Dawley rats undergoing middle cerebral artery infarction compared with tail vein injection, which indicates DMAPA-Amp and modified carotid injection may cast new lights on the therapy for angiogenesis of ischemic stroke.
展开▼
