[1]罗静,赵艳,谢婧雯,等.敲低SORL1表达构建模拟阿尔茨海默病的细胞模型[J].南方医科大学学报,2018,(01):8.
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敲低SORL1表达构建模拟阿尔茨海默病的细胞模型()
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《南方医科大学学报》[ISSN:1673-4254/CN:44-1627/R]

卷:
期数:
2018年01期
页码:
8
栏目:
出版日期:
2018-01-20

文章信息/Info

Title:
Establishment of a new cell model mimicking Alzheimer’s disease by knocking down SORL1 expression
作者:
罗静赵艳谢婧雯刘鑫林芳波侯德仁
关键词:
分拣蛋白相关受体1阿尔茨海默病N2a细胞转染
Keywords:
SORL1 Alzheimer’s disease N2a cell transfection
摘要:
目的通过比较分拣蛋白相关受体1(SORL1)敲低细胞模型和传统的阿尔茨海默病(AD)细胞模型在细胞活力、细胞凋亡 率、TNF-α和IL-1β表达的变化,构建一种模拟AD的细胞模型。方法(1)传统的AD细胞模型的构建:用Aβ25-35诱导N2a细胞 并通过检测细胞活力确定最佳的AD细胞模型。(2)SORL1敲低细胞模型的构建:RNA干扰慢病毒载体及空病毒载体分别转染 给N2a细胞,采用qRT—PCR和Western blot检测和鉴定细胞内SORL1转染的成功。(3)实验分组:对照组:未经干预的野生型 N2a细胞;AD 组:用Aβ25-35诱导的N2a细胞;空白组:用慢病毒空载体转染的N2a细胞;SORL1敲低组:SORL1-shRNA慢病毒 载体转染的N2a细胞。(4)MTT测定细胞活力,流式细胞仪检测细胞凋亡率,ELISA检测细胞培养上清液中TNF-α、IL-1β的含 量。结果(1)AD 组与对照组比较、SORL1敲低组与空白组比较、AD组和SORL1敲低组的细胞活力均降低、细胞凋亡率均增 加,差异有统计学意义(P<0.05);AD 组与SORL1 敲低组比较、对照组与空白组比较,细胞活力和细胞凋亡率均无差异(P> 0.05)。(2)AD组与对照组比较、SORL1敲低组与空白组比较,AD组和SORL1敲低组的TNF-α和IL-1β含量均增加,差异有统计 学意义(P<0.05);AD 组与SORL1敲低组比较、对照组与空白组比较,TNF-α和IL-1β含量均无差异(P>0.05)。结论敲低N2a细 胞SORL1的表达可能构建一种类似于Aβ诱导的AD细胞模型。
Abstract:
Objective To establish a cell model mimicking Alzheimer’s disease (AD) by knocking down SORL1 gene and compare the viability, apoptosis, and expressions of tumor necrosis factor-α ( TNF-α) and interleukin-1β (IL-1β) in this model with a traditional Alzheimer’s disease cell model. Methods A traditional cell model of AD was established by inducing N2a cells with Aβ25-35, and the optimal Aβ25-35 concentration was determined by assessing the cell viability changes. Another cell model of AD was established by transfecting N2a cells with SORL1-shRNA lentiviral vector, and SORL1 expression in the transfected cells were detected using Western blotting and qRT-PCR. With wild-type N2a cells without any treatment and cells transfected with a scramble shRNA as the control groups, the two cell models were examined for cell viability with MTT assay, cell apoptosis with flow cytometry, and TNF-α and IL - 1β levels in the culture supernatant with ELISA. Results The two cell models of AD showed obviously decreased viability and increased cell apoptosis compared with the untreated control cells or cells transfected with a scramble shRNA (P<0.05); no significant difference was found in the cell viability and apoptosis rate between the two AD cell models or between the two control groups (P>0.05). Significantly increased expressions of TNF-α and IL-1β were observed in both of the two cell models compared with their respective control groups (P<0.05) without significant differences between the two cell models or between the two control groups (P>0.05). Conclusion A new AD cell model similar to Aβ25-35-induced AD model can be established by SORL1 knockdown in N2a cells.

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更新日期/Last Update: 1900-01-01