脂質(zhì)體包埋腦源性神經(jīng)營養(yǎng)因子靶向透過血腦屏障的研究
發(fā)布時(shí)間：2018-11-07 08:38
【摘要】：目的：采用脂質(zhì)體包埋腦源性神經(jīng)營養(yǎng)因子(brain derived neurotrophic factor, BDNF)并進(jìn)行適當(dāng)修飾,觀察其透過血腦屏障進(jìn)入腦內(nèi)的情況,為實(shí)現(xiàn)腦內(nèi)靶向給藥提供實(shí)驗(yàn)基礎(chǔ)。 方法：將前期試驗(yàn)中構(gòu)建的靶向脂質(zhì)體Tf-pGFAP-BDNF-PEG、 Tf-pCMV-BDNF-PEG及Tf-H2O-PEG,按照不同濃度10μg/kg和2μg/kg分別經(jīng)尾靜脈注射大鼠體內(nèi)。將雄性SD大鼠60只,體重(180±30)g,周齡6-8周,隨機(jī)分為：Tf-pGFAP-BDNF-PEG組,Tf-pCMV-BDNF-PEG組,Tf-H2O-PEG對(duì)照組；每組分為高劑量組(10μg)、低劑量組(2μg),共六組,每組10只。將三種不同脂質(zhì)體分別通過尾靜脈注射到大鼠體內(nèi),48小時(shí)后取腦及肝臟,分別行免疫熒光染色、逆轉(zhuǎn)錄PCR (reverse transcript ion-PCR, RT-PCR)及免疫組織化學(xué)方法觀察BDNF的分布及表達(dá)情況。 結(jié)果：(1)與Tf-H2O-PEG組相比,Tf-pCMV-BDNF-PEG和Tf-pGFAP-BDNF-PEG兩種不同脂質(zhì)體均可以有效透過血腦屏障,在腦組織不同部位BDNF分布不同,其主要分布于大腦皮層區(qū),而海馬及白質(zhì)區(qū)表達(dá)不明顯。(2)以不同類型脂質(zhì)體為載體包裹的BDNF在大腦皮層表達(dá)水平具有顯著性差異,其中Tf-pGFAP-BDNF-PEG組腦組織陽性細(xì)胞表達(dá)水平顯著高于Tf-pCMV-BDNF-PEG組及Tf-H2O-PEG組(P0.05)。(3)相同載體不同劑量脂質(zhì)體比較時(shí),Tf-pGFAP-BDNF-PEG及Tf-pCMV-BDNF-PEG組的表達(dá)也有差異,高劑量組透過血腦屏障的量明顯高于低劑量組(.P0.05),而Tf-H2O-PEG組不同劑量間比較無顯著性差異(P0.05)。(4)Tf-pCMV-BDNF-PEG組在肝臟中BDNF的表達(dá)明顯高于Tf-pGFAP-BDNF-PEG組(P-O.0.5),而Tf-H2O-PEG組在肝臟中BDNF的表達(dá)不明顯。 結(jié)論：采用Tf、PEG修飾的脂質(zhì)體并連接腦特異性啟動(dòng)子GFAP,可實(shí)現(xiàn)外源基因BDNF的腦內(nèi)轉(zhuǎn)染,減少外周器官的捕獲,增加腦內(nèi)的表達(dá)含量,一定程度提高了脂質(zhì)體的靶向性,為中樞神經(jīng)系統(tǒng)疾病的治療提供實(shí)驗(yàn)依據(jù)。
[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 鹵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.
【學(xué)位授予單位】：延邊大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R743
本文編號(hào)：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 鹵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.
【學(xué)位授予單位】：延邊大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R743
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