脂质体包埋脑源性神经营养因子靶向透过血脑屏障的研究
发布时间：2018-11-07 08:38
【摘要】：目的：采用脂质体包埋脑源性神经营养因子(brain derived neurotrophic factor, BDNF)并进行适当修饰,观察其透过血脑屏障进入脑内的情况,为实现脑内靶向给药提供实验基础。 方法：将前期试验中构建的靶向脂质体Tf-pGFAP-BDNF-PEG、 Tf-pCMV-BDNF-PEG及Tf-H2O-PEG,按照不同浓度10μg/kg和2μg/kg分别经尾静脉注射大鼠体内。将雄性SD大鼠60只,体重(180±30)g,周龄6-8周,随机分为：Tf-pGFAP-BDNF-PEG组,Tf-pCMV-BDNF-PEG组,Tf-H2O-PEG对照组；每组分为高剂量组(10μg)、低剂量组(2μg),共六组,每组10只。将三种不同脂质体分别通过尾静脉注射到大鼠体内,48小时后取脑及肝脏,分别行免疫荧光染色、逆转录PCR (reverse transcript ion-PCR, RT-PCR)及免疫组织化学方法观察BDNF的分布及表达情况。 结果：(1)与Tf-H2O-PEG组相比,Tf-pCMV-BDNF-PEG和Tf-pGFAP-BDNF-PEG两种不同脂质体均可以有效透过血脑屏障,在脑组织不同部位BDNF分布不同,其主要分布于大脑皮层区,而海马及白质区表达不明显。(2)以不同类型脂质体为载体包裹的BDNF在大脑皮层表达水平具有显著性差异,其中Tf-pGFAP-BDNF-PEG组脑组织阳性细胞表达水平显著高于Tf-pCMV-BDNF-PEG组及Tf-H2O-PEG组(P0.05)。(3)相同载体不同剂量脂质体比较时,Tf-pGFAP-BDNF-PEG及Tf-pCMV-BDNF-PEG组的表达也有差异,高剂量组透过血脑屏障的量明显高于低剂量组(.P0.05),而Tf-H2O-PEG组不同剂量间比较无显著性差异(P0.05)。(4)Tf-pCMV-BDNF-PEG组在肝脏中BDNF的表达明显高于Tf-pGFAP-BDNF-PEG组(P-O.0.5),而Tf-H2O-PEG组在肝脏中BDNF的表达不明显。 结论：采用Tf、PEG修饰的脂质体并连接脑特异性启动子GFAP,可实现外源基因BDNF的脑内转染,减少外周器官的捕获,增加脑内的表达含量,一定程度提高了脂质体的靶向性,为中枢神经系统疾病的治疗提供实验依据。
[Abstract]:Aim: to investigate the effect of (brain derived neurotrophic factor, BDNF) entrapment with liposome on the brain through the blood-brain barrier in order to provide the experimental basis for the targeted administration of brain-derived neurotrophic factor (BDNF) in the brain. Methods: the targeted liposomes Tf-pGFAP-BDNF-PEG, Tf-pCMV-BDNF-PEG and Tf-H2O-PEG, were injected into rat caudal vein according to different concentrations of 10 渭 g/kg and 2 渭 g/kg respectively. Sixty male SD rats, weighing (180 卤30) g, aged 6-8 weeks, were randomly divided into Tf-pGFAP-BDNF-PEG group, Tf-pCMV-BDNF-PEG group and Tf-H2O-PEG control group. Each group was divided into high dose group (10 渭 g), low dose group) (2 渭 g), group), 10 rats in each group. Three kinds of liposomes were injected into the rat via tail vein respectively. The brain and liver were taken 48 hours later. Immunofluorescence staining and reverse transcription PCR (reverse transcript ion-PCR, were performed respectively. RT-PCR) and immunohistochemical method were used to observe the distribution and expression of BDNF. Results: (1) compared with Tf-H2O-PEG group, Tf-pCMV-BDNF-PEG and Tf-pGFAP-BDNF-PEG liposomes could effectively penetrate the blood-brain barrier, and the distribution of BDNF in different parts of brain tissue was different. However, the expression of BDNF in hippocampus and white matter was not obvious. (2) the expression level of BDNF coated with different types of liposomes was significantly different in cerebral cortex. The expression level of brain positive cells in Tf-pGFAP-BDNF-PEG group was significantly higher than that in Tf-pCMV-BDNF-PEG group and Tf-H2O-PEG group (P0.05). (3). There were also differences in the expression of Tf-pGFAP-BDNF-PEG and Tf-pCMV-BDNF-PEG. The amount of blood brain barrier in high dose group was significantly higher than that in low dose group (.P0.05). However, the expression of BDNF in liver of Tf-pCMV-BDNF-PEG group was significantly higher than that of Tf-pGFAP-BDNF-PEG group (P-O.0.5), but there was no significant difference between different doses of Tf-H2O-PEG group (P0.05). (4), the expression of BDNF in liver of Tf-pCMV-BDNF-PEG group was significantly higher than that of Tf-pGFAP-BDNF-PEG group (P-O.0.5). However, the expression of BDNF in the liver of Tf-H2O-PEG group was not obvious. Conclusion: Tf,PEG modified liposome and brain specific promoter GFAP, can be used to transfect the exogenous gene BDNF in brain, reduce the capture of peripheral organs, increase the content of brain expression, and improve the targeting of liposome to some extent. To provide experimental basis for the treatment of central nervous system diseases.
【学位授予单位】：延边大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743
本文编号：2315802
[Abstract]:Aim: to investigate the effect of (brain derived neurotrophic factor, BDNF) entrapment with liposome on the brain through the blood-brain barrier in order to provide the experimental basis for the targeted administration of brain-derived neurotrophic factor (BDNF) in the brain. Methods: the targeted liposomes Tf-pGFAP-BDNF-PEG, Tf-pCMV-BDNF-PEG and Tf-H2O-PEG, were injected into rat caudal vein according to different concentrations of 10 渭 g/kg and 2 渭 g/kg respectively. Sixty male SD rats, weighing (180 卤30) g, aged 6-8 weeks, were randomly divided into Tf-pGFAP-BDNF-PEG group, Tf-pCMV-BDNF-PEG group and Tf-H2O-PEG control group. Each group was divided into high dose group (10 渭 g), low dose group) (2 渭 g), group), 10 rats in each group. Three kinds of liposomes were injected into the rat via tail vein respectively. The brain and liver were taken 48 hours later. Immunofluorescence staining and reverse transcription PCR (reverse transcript ion-PCR, were performed respectively. RT-PCR) and immunohistochemical method were used to observe the distribution and expression of BDNF. Results: (1) compared with Tf-H2O-PEG group, Tf-pCMV-BDNF-PEG and Tf-pGFAP-BDNF-PEG liposomes could effectively penetrate the blood-brain barrier, and the distribution of BDNF in different parts of brain tissue was different. However, the expression of BDNF in hippocampus and white matter was not obvious. (2) the expression level of BDNF coated with different types of liposomes was significantly different in cerebral cortex. The expression level of brain positive cells in Tf-pGFAP-BDNF-PEG group was significantly higher than that in Tf-pCMV-BDNF-PEG group and Tf-H2O-PEG group (P0.05). (3). There were also differences in the expression of Tf-pGFAP-BDNF-PEG and Tf-pCMV-BDNF-PEG. The amount of blood brain barrier in high dose group was significantly higher than that in low dose group (.P0.05). However, the expression of BDNF in liver of Tf-pCMV-BDNF-PEG group was significantly higher than that of Tf-pGFAP-BDNF-PEG group (P-O.0.5), but there was no significant difference between different doses of Tf-H2O-PEG group (P0.05). (4), the expression of BDNF in liver of Tf-pCMV-BDNF-PEG group was significantly higher than that of Tf-pGFAP-BDNF-PEG group (P-O.0.5). However, the expression of BDNF in the liver of Tf-H2O-PEG group was not obvious. Conclusion: Tf,PEG modified liposome and brain specific promoter GFAP, can be used to transfect the exogenous gene BDNF in brain, reduce the capture of peripheral organs, increase the content of brain expression, and improve the targeting of liposome to some extent. To provide experimental basis for the treatment of central nervous system diseases.
【学位授予单位】：延边大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R743
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