Illawarra Health and Medical Research Institute, Wollongong, NSW 2522, Australia.
School of Chemistry and Molecular Bioscience, University of Wollongong, Wollongong, NSW 2522, Australia.
Biosci Rep. 2021 Feb 26;41(2). doi: 10.1042/BSR20204243.
APOE ε4 is the major genetic risk factor for Alzheimer's disease (AD). A precise role for apolipoprotein E (apoE) in the pathogenesis of the disease remains unclear in part due to its expression in multiple cell types of the brain. APOE is highly expressed in astrocytes and microglia, however its expression can also be induced in neurons under various conditions. The neuron-like cell line SK-N-SH is a useful model in the study of the cellular and molecular effects of apoE as it can be differentiated with retinoic acid to express and secrete high levels of apoE and it also shows the same apoE fragmentation patterns observed in the human brain. We previously found that apoE is cleaved into a 25-kDa fragment by high temperature-requirement serine protease A1 (HtrA1) in SK-N-SH cells. To further understand the endogenous functions of apoE, we used CRISPR/Cas9 to generate SK-N-SH cell lines with APOE expression knocked-down (KD). APOE KD cells showed lower APOE and HTRA1 expression than parental SK-N-SH cells but no overt differences in neuritogenesis or cell proliferation compared with the CRISPR/Cas9 control cells. This research shows that the loss of apoE and HtrA1 has a negligible effect on neuritogenesis and cell survival in SK-N-SH neuron-like cells.
载脂蛋白 E（APOE）ε4 是阿尔茨海默病（AD）的主要遗传风险因素。APOE 在疾病发病机制中的精确作用尚不清楚，部分原因是其在大脑的多种细胞类型中表达。APOE 在星形胶质细胞和小胶质细胞中高度表达，但在各种条件下也可以在神经元中诱导其表达。神经元样细胞系 SK-N-SH 是研究 APOE 细胞和分子作用的有用模型，因为它可以用维甲酸分化，表达和分泌高水平的 APOE，并且还显示出与人类大脑中观察到的相同的 APOE 片段模式。我们之前发现 APOE 在 SK-N-SH 细胞中被高温需求丝氨酸蛋白酶 A1（HtrA1）切割成 25kDa 的片段。为了进一步了解 APOE 的内源性功能，我们使用 CRISPR/Cas9 生成 APOE 表达敲低（KD）的 SK-N-SH 细胞系。APOE KD 细胞的 APOE 和 HTRA1 表达低于亲本 SK-N-SH 细胞，但与 CRISPR/Cas9 对照细胞相比，在神经突生成或细胞增殖方面没有明显差异。这项研究表明，在 SK-N-SH 神经元样细胞中，APOE 和 HtrA1 的缺失对神经突生成和细胞存活几乎没有影响。
