TREM2 drives microglia response to amyloid-β via SYK-dependent and -independent pathways

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Generation of APOE knock-down SK-N-SH human neuroblastoma cells using CRISPR/Cas9: a novel cellular model relevant to Alzheimer’s disease research

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.     

ApoE4 upregulates the activity of mitochondria‐associated ER membranes

In addition to the appearance of senile plaques and neurofibrillary tangles, Alzheimer''s disease (AD) is characterized by aberrant lipid metabolism and early mitochondrial dysfunction. We recently showed that there was increased functionality of mitochondria‐associated endoplasmic reticulum (ER) membranes (MAM), a subdomain of the ER involved in lipid and cholesterol homeostasis, in presenilin‐deficient cells and in fibroblasts from familial and sporadic AD patients. Individuals carrying the ε4 allele of apolipoprotein E (ApoE4) are at increased risk for developing AD compared to those carrying ApoE3. While the reason for this increased risk is unknown, we hypothesized that it might be associated with elevated MAM function. Using an astrocyte‐conditioned media (ACM) model, we now show that ER–mitochondrial communication and MAM function—as measured by the synthesis of phospholipids and of cholesteryl esters, respectively—are increased significantly in cells treated with ApoE4‐containing ACM as compared to those treated with ApoE3‐containing ACM. Notably, this effect was seen with lipoprotein‐enriched preparations, but not with lipid‐free ApoE protein. These data are consistent with a role of upregulated MAM function in the pathogenesis of AD and may help explain, in part, the contribution of ApoE4 as a risk factor in the disease.     

Post-translational modifications in MeHg-induced neurotoxicity

Mercury (Hg) exposure remains a major public health concern due to its widespread distribution in the environment. Organic mercurials, such as MeHg, have been extensively investigated especially because of their congenital effects. In this context, studies on the molecular mechanism of MeHg-induced neurotoxicity are pivotal to the understanding of its toxic effects and the development of preventive measures. Post-translational modifications (PTMs) of proteins, such as phosphorylation, ubiquitination, and acetylation are essential for the proper function of proteins and play important roles in the regulation of cellular homeostasis. The rapid and transient nature of many PTMs allows efficient signal transduction in response to stress. This review summarizes the current knowledge of PTMs in MeHg-induced neurotoxicity, including the most commonly PTMs, as well as PTMs induced by oxidative stress and PTMs of antioxidant proteins. Though PTMs represent an important molecular mechanism for maintaining cellular homeostasis and are involved in the neurotoxic effects of MeHg, we are far from understanding the complete picture on their role, and further research is warranted to increase our knowledge of PTMs in MeHg-induced neurotoxicity.     

Effects of oxysterols on the blood–brain barrier: Implications for Alzheimer’s disease

Altered brain cholesterol homeostasis plays a key role in neurodegenerative diseases such as Alzheimer’s disease (AD). For a long time, the blood–brain barrier (BBB) was basically considered as a barrier isolating the brain from circulating cholesterol, however, several lines of evidence now suggest that the BBB strictly regulates the exchanges of sterol between the brain and the peripheral circulation. Oxysterols, synthesized by neurons or by peripheral cells, cross the BBB easily and modulate the expression of several enzymes, receptors and transporters which are involved not only in cholesterol metabolism but also in other brain functions. This review article deals with the way oxysterols impact BBB cells. These perspectives open new routes for designing certain therapeutical approaches that target the BBB so that the onset and/or progression of brain diseases such as AD may be modulated.     

Increased effect of the ApoE gene on survival at advanced age in healthy and long‐lived Danes: two nationwide cohort studies

Studies of Nordic twins suggest an increased genetic influence on mortality with age. Contrary to this, the heterogeneity hypothesis predicts that the mortality of individuals carrying a ‘frail’ or ‘risky’ genotype in a population will approach that of noncarriers with age because of selection pressure. The ApoE ε4 allele is associated with an increased mortality risk, and its effect has been suggested to decrease with age. Here, we investigated the effect of ApoE ε4 allele on survival in a sample of the healthiest and long‐lived Danes. The study population comprised Danes born in 1905 and a replicate sample of the 1895 cohort. For the 1905 cohort, a total of 350 carriers and 1256 noncarriers of the ApoE ε4 allele were followed from 1998 until death or end of follow‐up. Cox regression models were used for the analysis. Of the 1606 persons with known ApoE ε4 status in 1998, 1546 had died at the end of the 10‐year follow‐up. Carriers of the ApoE ε4 allele had an increased mortality compared to noncarriers, and the influence of ApoE status on mortality increased in the age interval 92–103. For the covariates sex and independency status, the difference in relative risk of death between groups decreased with advancing age. Our findings of increasing influence of ApoE ε4 allele on mortality with age do not support previous findings of decreased influence ApoE ε4 allele on mortality with age, and alternative models such as the multifactorial threshold models should be considered for understanding the genetic effects on mortality at advanced age.     

Adventitial fibroblasts are activated in the early stages of atherosclerosis in the apolipoprotein E knockout mouse

The role of the adventitia in vascular function and vascular lesion formation has been largely ignored. This study observed the activation of the adventitia and specifically the fibroblasts in the development of atherosclerosis in the apoE(-/-) mouse. The results showed a gradual increase in expression of collagen types I and III after 2, 4, and 8 weeks of hyperlipidic diet. The earliest expression of monocyte chemoattractant protein-1 (MCP-1) protein and mRNA was detected in the adventitial fibroblast before the formation of intimal lesions. Proliferation, too, was first found in the adventitial fibroblasts. We hypothesize that the adventitial fibroblast is activated in the early stage of atherosclerosis. Adventitial inflammation may be an early event in the development of atherosclerotic lesions.     

瑞舒伐他汀对载脂蛋白E基因敲除小鼠动脉粥样硬化中调节性T细胞的影响

目的：探讨瑞舒伐他汀对载脂蛋白E基因敲除（ApoEKO）小鼠动脉粥样硬化中调节性T细胞的影响。方法：首先将30只ApoEKO小鼠建立动脉粥样硬化模型,随机分为高胆固醇饮食组（对照组）、瑞舒伐他汀低剂量组和瑞舒伐他汀高剂量组,各组分别给予蒸馏水或瑞舒伐他汀进行干预8周;将主动脉根部行冰冻切片油红染色,评估粥样硬化斑块面积大小;免疫组织化学法检测主动脉根部粥样硬化斑块处调节性T细胞（Treg）的表达。结果：各组小鼠均有动脉粥样硬化斑块形成,采用瑞舒伐他汀治疗的小鼠动脉粥样硬化斑块的面积明显小于未经治疗的小鼠（P〈0.01）,同时瑞舒伐他汀能明显增加粥样硬化病变处调节性T细胞的表达,且呈现剂效关系。结论：本实验观察到瑞舒伐他汀不仅能减小ApoEKO小鼠的主动脉粥样硬化斑块,且能使调节性T细胞的表达增多,推测瑞舒伐他汀可以通过促进调节性T细胞的生成而起到抑制动脉粥样硬化的作用。     

Structural variation in human apolipoprotein E3 and E4: secondary structure, tertiary structure, and size distribution

Human apolipoprotein E (apoE) is a 299-amino-acid protein with a molecular weight of 34 kDa. The difference between the apoE3 and apoE4 isoforms is a single residue substitution involving a Cys-Arg replacement at residue 112. ApoE4 is positively associated with atherosclerosis and late-onset and sporadic Alzheimer's disease (AD). ApoE4 and its C-terminal truncated fragments have been found in the senile plaques and neurofibrillary tangles in the brain of AD patients. However, detail structural information regarding isoform and domain interaction remains poorly understood. We prepared full-length, N-, and C-terminal truncated apoE3 and apoE4 proteins and studied their structural variation. Sedimentation velocity and continuous size distribution analysis using analytical ultracentrifugation revealed apoE3(72-299) as consisting of a major species with a sedimentation coefficient of 5.9. ApoE4(72-299) showed a wider and more complicated species distribution. Both apoE3 and E4 N-terminal domain (1-191) existed with monomers as the major component together with some tetramer. The oligomerization and aggregation of apoE protein increased when the C-terminal domain (192-271) was incorporated. The structural influence of the C-terminal domain on apoE is to assist self-association with no significant isoform preference. Circular dichroism and fluorescence studies demonstrated that apoE4(72-299) possessed a more alpha-helical structure with more hydrophobic residue exposure. The structural variation of the N-terminal truncated apoE3 and apoE4 protein provides useful information that helps to explain the greater aggregation of the apoE4 isoform and thus has implication for the involvement of apoE4 in AD.