[Show abstract][Hide abstract] ABSTRACT: Apolipoprotein E (ApoE) is a major cholesterol carrier and plays an important role in maintaining lipid homeostasis both in the periphery and brain. Human APOE gene is polymorphic at two single nucleotides (rs429358 and rs7412) resulting in three different alleles (ε2, ε3 and ε4). ApoE isoforms modulate the risk for a variety of vascular and neurodegenerative diseases; thus, APOE genotyping is crucial for predicting disease risk and designing individualized therapy based on APOE genotype.
We have developed an APOE genotyping method that is based on allele-specific PCR methodology adapted to Real Time PCR monitored by TaqMan probe. Rather than using TaqMan probes specific for the two polymorphic sites, only one TaqMan probe is used as the polymorphic alleles are recognized by site-specific PCR primers. Each genotyping assay can be completed within 90 minutes and is applicable to high-throughput analysis. Using this protocol, we genotyped a total of 1158 human DNA samples and obtained a 100 % concordance with the APOE genotype determined by sequencing analysis.
The APOE genotyping assay we have developed is accurate and cost-effective. In addition, our assay can readily be applied to genotyping large sample numbers. Therefore, our APOE genotyping method can be used for assessing the risk for a variety of vascular and neurodegenerative diseases that have been reported to be associated with APOE polymorphism.
Preview · Article · Dec 2016 · Molecular Neurodegeneration
[Show abstract][Hide abstract] ABSTRACT: Apolipoprotein E (apoE) mediates lipid metabolism both in peripheral and in the brain. The human APOE gene has three polymorphic alleles that influence the risk for various types of cancer and neurodegenerative diseases. A potential association between APOE allele and the risk for gastric cancer has been implicated, but the specific allele involved and potential associations with the subtype and the grade of cancer malignancy need further clarification. We screened the APOE genotype in 550 gastric cancer patients and 550 non-cancer control individuals and found that the presence of the APOE ε2 and lower serum total cholesterol are associated with an increased risk for gastric cancer (all P ≤ 0.0005). Interestingly, APOE ε2 is also correlated with increased risk for both intestinal and diffuse histotypes but not with TN classification or stage in gastric cancer patients, suggesting that APOE polymorphic alleles are associated with the risk of development but unlikely the progression of gastric cancer. Since ε2 carriers have lower levels of serum total cholesterol than non-ε2 carriers, our findings suggest that the increased risk for gastric cancer by APOE ε2 allele might be mediated through lowered serum total cholesterol levels.
[Show abstract][Hide abstract] ABSTRACT: Alzheimer's disease (AD) is the most common form of dementia in the elderly. It is generally believed that β-amyloidogenesis, tau-hyperphosphorylation, and synaptic loss underlie cognitive decline in AD. Rps23rg1, a functional retroposed mouse gene, has been shown to reduce Alzheimer's β-amyloid (Aβ) production and tau phosphorylation. In this study, we have identified its human homolog, and demonstrated that RPS23RG1 regulates synaptic plasticity, thus counteracting Aβ oligomer (oAβ)-induced cognitive deficits in mice. The level of RPS23RG1 mRNA is significantly lower in the brains of AD compared to non-AD patients, suggesting its potential role in the pathogenesis of the disease. Similar to its mouse counterpart, human RPS23RG1 interacts with adenylate cyclase, activating PKA/CREB, and inhibiting GSK-3. Furthermore, we show that human RPS23RG1 promotes synaptic plasticity and offsets oAβ-induced synaptic loss in a PKA-dependent manner in cultured primary neurons. Overexpression of Rps23rg1 in transgenic mice consistently prevented oAβ-induced PKA inactivation, synaptic deficits, suppression of long-term potentiation, and cognitive impairment as compared to wild type littermates. Our study demonstrates that RPS23RG1 may reduce the occurrence of key elements of AD pathology and enhance synaptic functions to counteract oAβ-induced synaptic and cognitive deficits in AD.
Full-text · Article · Jan 2016 · Scientific Reports
[Show abstract][Hide abstract] ABSTRACT: Background:
Intelligence is an important human feature that strongly affects many life outcomes, including health, life-span, income, educational and occupational attainments. People at all ages differ in their intelligence but the origins of these differences are much debated. A variety of environmental and genetic factors have been reported to be associated with individual intelligence, yet their nature and contribution to intelligence differences have been controversial.
To investigate the contribution of apolipoprotein E (APOE) genotype, which is associated with the risk for Alzheimer's disease, as well as demographic and lifestyle characteristics, to the variation in intelligence.
A total of 607 Chinese college students aged 18 to 25 years old were included in this prospective observational study. The Chinese revision of Wechsler Adult Intelligence Scale (the fourth edition, short version) was used to determine the intelligence level of participants. Demographic and lifestyle characteristics data were obtained from self-administered questionnaires.
No significant association was found between APOE polymorphic alleles and different intelligence quotient (IQ) measures. Interestingly, a portion of demographic and lifestyle characteristics, including age, smoking and sleep quality were significantly associated with different IQ measures.
Our findings indicate that demographic features and lifestyle characteristics, but not APOE genotype, are associated with intelligence measures among young Chinese college students. Thus, although APOE ε4 allele is a strong genetic risk factor for Alzheimer's disease, it does not seem to impact intelligence at young ages.
[Show abstract][Hide abstract] ABSTRACT: Several heterozygous missense mutations in the triggering receptor expressed on myeloid cells 2 (TREM2) have recently been linked to risk for a number of neurological disorders including Alzheimer's disease (AD), Parkinson's disease (PD) and frontotemporal dementia (FTD). These discoveries have re-ignited interest in the role of neuroinflammation in the pathogenesis of neurodegenerative diseases. TREM2 is highly expressed in microglia, the resident immune cells of the central nervous system. Along with its adaptor protein, DAP12, TREM2 regulates inflammatory cytokine release and phagocytosis of apoptotic neurons. Here, we report apolipoprotein E (apoE) as a novel ligand for TREM2. Using a biochemical assay, we demonstrated high-affinity binding of apoE to human TREM2. The functional significance of this binding was highlighted by increased phagocytosis of apoE-bound apoptotic N2a cells by primary microglia in a manner that depends on TREM2 expression. Moreover, when the AD-associated TREM2-R47H mutant was used in biochemical assays, apoE binding was vastly reduced. Our data demonstrate that apoE-TREM2 interaction in microglia plays critical roles in modulating phagocytosis of apoE-bound apoptotic neurons and establish a critical link between two proteins whose genes are strongly linked to the risk for AD.
No preview · Article · Sep 2015 · Journal of Biological Chemistry
[Show abstract][Hide abstract] ABSTRACT: Myeloid-lineage cells accomplish a myriad of homeostatic tasks including the recognition of pathogens, regulation of the inflammatory milieu, and mediation of tissue repair and regeneration. The innate immune receptor and its adaptor protein-triggering receptor expressed on myeloid cells 2 (TREM2) and DNAX-activating protein of 12 kDa (DAP12)-possess the ability to modulate critical cellular functions via crosstalk with diverse signaling pathways. As such, mutations in TREM2 and DAP12 have been found to be associated with a range of disease phenotypes. In particular, mutations in TREM2 increase the risk for Alzheimer's disease and other neurodegenerative disorders. The leading hypothesis is that microglia, the resident immune cells of the central nervous system, are the major myeloid cells affected by dysregulated TREM2-DAP12 function. Here, we review how impaired signaling by the TREM2-DAP12 pathway leads to altered immune responses in phagocytosis, cytokine production, and microglial proliferation and survival, thus contributing to disease pathogenesis.
[Show abstract][Hide abstract] ABSTRACT: Circulating tumor cells (CTCs) are in limited numbers and heterogeneous, making their detection, isolation, and enumeration a major challenge. To overcome these difficulties, we developed a novel method to detect and enumerate CTCs with invasive property. Our assay consists of three simple steps: enrichment, Matrigel invasion assay, and immunostaining. We have validated this method using mouse xenograft tumor models and confirmed its utility in human cancer patients. Our method does not require special equipment and antigen expression for CTC selection, is less likely to be affected by the heterogeneity of the CTCs, and could be applicable to virtually all cancers. Most important, our method enumerates invasive CTCs, which may allow more accurate correlations with clinical outcome and treatment response compared with other CTC detection methods.
[Show abstract][Hide abstract] ABSTRACT: Amyloid-β (Aβ) peptide plays an essential role in the pathogenesis of Alzheimer's disease (AD) and is generated from amyloid-β precursor protein (APP) through sequential proteolytic cleavages by β-site APP cleaving enzyme 1 (BACE1) and γ-secretase. Trafficking dysregulation of APP, BACE1, and γ-secretase may affect Aβ generation and disease pathogenesis. Sorting nexin 15 (SNX15) is known to regulate protein trafficking. Here, we report that SNX15 is abundantly expressed in mouse neurons and astrocytes. In addition, we show that although not affecting the protein levels of APP, BACE1, and γ-secretase components and the activity of BACE1 and γ-secretase, overexpression and downregulation of SNX15 reduce and promote Aβ production, respectively. Furthermore, we find that overexpression of SNX15 increases APP protein levels in cell surface through accelerating APP recycling, whereas downregulation of SNX15 has an opposite effect. Finally, we show that exogenous expression of human SNX15 in the hippocampal dentate gyrus by adeno-associated virus (AAV) infection can significantly reduce Aβ pathology in the hippocampus and improve short-term working memory in the APPswe/PSEN1dE9 double transgenic AD model mice. Together, our results suggest that SNX15 regulates the recycling of APP to cell surface and, thus, its processing for Aβ generation.
No preview · Article · Jun 2015 · Molecular Neurobiology
[Show abstract][Hide abstract] ABSTRACT: Background
Human apolipoprotein E (apoE) exists in three major isoforms: apoE2, apoE3 and apoE4. In the brain, apoE is produced mostly by astrocytes and transports cholesterol to neurons via apoE receptors. Among the gene alleles encoding the three isoforms, the APOE4 allele is the strongest genetic risk factor for late-onset Alzheimer’s disease (AD), whereas APOE2 is protective. ApoE4 confers a gain of toxic function, a loss of neuroprotective function or a combination of both in AD pathogenesis. Given that therapeutic impacts of modulating apoE expression may be isoform-dependent, we sought to investigate the relationship between overexpressing apoE isoform and apoE-related functions in apoE-targeted replacement (TR) mice. Specifically, apoE isoform expression driven by the astrocyte-specific glial fibrillary acidic protein (GFAP) promoter was built into an adeno-associated virus serotype 8 (AAV8) vector and injected into the ventricles of postnatal day 2 (P2) apoE3-TR or apoE4-TR mice. Upon confirmation of apoE isoform expression, effects on apoE lipidation and the levels of amyloid-β (Aβ) in the brain were assessed.
AAV8-GFAP-apoE isoforms were specifically expressed in astrocytes throughout all brain regions, which led to overall increased apoE levels in the brain. Viral mediated overexpression of apoE4 in the apoE4-TR background increased poorly-lipidated apoE lipoprotein particles and decreased apoE-associated cholesterol in apoE4-TR mice. Conversely, apoE2 overexpression in apoE4-TR mice enhanced apoE lipidation and associated cholesterol. Furthermore, overexpression of apoE4 elevated the levels of endogenous Aβ, whereas apoE2 overexpression trended to lower endogenous Aβ.
Overexpression of apoE isoforms induces differential effects in the apoE4-TR background: apoE4 decreases apoE lipidation and enhances Aβ accumulation, whereas apoE2 has the opposite effects. Our findings suggest that increasing apoE2 in APOE4 carriers is a beneficial strategy to treat AD, whereas increasing apoE4 in APOE4 carriers is likely harmful. We have also established novel methods to express apoE isoforms in mouse brain to study apoE-related pathways in AD and related dementia.
Electronic supplementary material
The online version of this article (doi:10.1186/s13024-015-0001-3) contains supplementary material, which is available to authorized users.
No preview · Article · Mar 2015 · Molecular Neurodegeneration
[Show abstract][Hide abstract] ABSTRACT: Accumulation and deposition of amyloid-β peptide (Aβ) in the brain is a primary cause of the pathogenesis of Alzheimer's disease (AD). Aβ is generated from amyloid-β precursor protein (APP) through sequential cleavages first by β-secretase and then by γ-secretase. Inhibiting β-secretase activity is believed to be one of the most promising strategies for AD treatment. In the present study, we found that a resveratrol trimer, miyabenol C, isolated from stems and leaves of the small-leaf grape (Vitisthunbergii var. taiwaniana), can markedly reduce Aβ and sAPPβ levels in both cell cultures and the brain of AD model mice. Mechanistic studies revealed that miyabenol C affects neither protein levels of APP, the two major α-secretases ADAM10 and TACE, and the γ-secretase component Presenilin 1, nor γ-secretase-mediated Notch processing and TACE activity. In contrast, although miyabenol C has no effect on altering protein levels of the β-secretase BACE1, it can inhibit both in vitro and in vivo β-secretase activity. Together, our results indicate that miyabenol C is a prominent β-secretase inhibitor and lead compound for AD drug development.
[Show abstract][Hide abstract] ABSTRACT: One of the most critical pathological features of Alzheimer's disease (AD) is the accumulation of β-amyloid (Aβ) peptides that form extracellular senile plaques in the brain. Aβ is derived from β-amyloid precursor protein (APP) through sequential cleavage by β- and γ-secretases. γ-secretase is a high molecular weight complex minimally composed of four components: presenilins (PS), nicastrin, anterior pharynx defective 1 (APH-1), and presenilin enhancer 2 (PEN-2). In addition to APP, γ-secretase also cleaves many other type I transmembrane (TM) protein substrates. As a crucial enzyme for Aβ production, γ-secretase is an appealing therapeutic target for AD. Here, we summarize current knowledge on the structure and function of γ-secretase, as well as recent progress in developing γ-secretase targeting drugs for AD treatment.
Full-text · Article · Dec 2014 · Frontiers in Cellular Neuroscience
[Show abstract][Hide abstract] ABSTRACT: Emerging evidence indicates that certain microRNAs (miRNAs) play important roles in epileptogenesis. MiR-219 is a brain-specific miRNA and has been shown to negatively regulate the function of N-methyl-D-aspartate (NMDA) receptors by targeting Ca(2+)/calmodulin-dependent protein kinase II (CaMKII)γ. Herein, we found that the level of miR-219 was decreased in both the kainic acid (KA)-induced epilepsy model and in cerebrospinal fluid specimens of epilepsy patients. Importantly, silencing of miR-219 by its antagomir in vivo resulted in seizure behaviors, abnormal cortical electroencephalogram (EEG) recordings in the form of high-amplitude and high-frequency discharges, and increased levels of CaMKIIγ and an NMDA receptor component, NR1, in a pattern similar to that found in KA-treated mice. Moreover, treatments with the miR-219 agomir in vivo alleviated seizures, abnormal EEG recordings, and decreased levels of CaMKIIγ and NR1 in KA-treated mice. Furthermore, treatment with MK-801, an antagonist of NMDA receptors, significantly alleviated abnormal EEG recordings induced by miR-219 antagomir. Together, these results demonstrate that miR-219 plays a crucial role in suppressing seizure formation in experimental models of epilepsy through modulating the CaMKII/NMDA receptor pathway and that miR-219 supplement may be a potential anabolic strategy for ameliorating epilepsy.
No preview · Article · Nov 2014 · Molecular Neurobiology
[Show abstract][Hide abstract] ABSTRACT: Alzheimer's disease (AD) is a devastating disease characterized by synaptic and neuronal loss in the elderly. Compelling evidence suggests that soluble amyloid-beta peptide (Abeta) oligomers induce synaptic loss in AD. Abeta-induced synaptic dysfunction is dependent on overstimulation of N-methyl-D-aspartate receptors (NMDARs) resulting in aberrant activation of redox-mediated events as well as elevation of cytoplasmic Ca2+, which in turn triggers downstream pathways involving phospho-tau (p-tau), caspases, Cdk5/dynamin-related protein 1 (Drp1), calcineurin/PP2B, PP2A, Gsk-3beta, Fyn, cofilin, and CaMKII and causes endocytosis of AMPA receptors (AMPARs) as well as NMDARs. Dysfunction in these pathways leads to mitochondrial dysfunction, bioenergetic compromise and consequent synaptic dysfunction and loss, impaired long-term potentiation (LTP), and cognitive decline. Evidence also suggests that Abeta may, at least in part, mediate these events by causing an aberrant rise in extrasynaptic glutamate levels by inhibiting glutamate uptake or triggering glutamate release from glial cells. Consequent extrasynaptic NMDAR (eNMDAR) overstimulation then results in synaptic dysfunction via the aforementioned pathways. Consistent with this model of Abeta-induced synaptic loss, Abeta synaptic toxicity can be partially ameliorated by the NMDAR antagonists (such as memantine and NitroMemantine). PSD-95, an important scaffolding protein that regulates synaptic distribution and activity of both NMDA and AMPA receptors, is also functionally disrupted by Abeta. PSD-95 dysregulation is likely an important intermediate step in the pathological cascade of events caused by Abeta. In summary, Abeta-induced synaptic dysfunction is a complicated process involving multiple pathways, components and biological events, and their underlying mechanisms, albeit as yet incompletely understood, may offer hope for new therapeutic avenues.
Preview · Article · Nov 2014 · Molecular Neurodegeneration
[Show abstract][Hide abstract] ABSTRACT: Alzheimer's disease (AD) is an age-related neurological disorder characterized by synaptic loss and dementia. The low-density lipoprotein receptor-related protein 6 (LRP6) is an essential coreceptor for Wnt signaling, and its genetic variants have been linked to AD risk. Here we report that neuronal LRP6-mediated Wnt signaling is critical for synaptic function and cognition. Conditional deletion of Lrp6 gene in mouse forebrain neurons leads to age-dependent deficits in synaptic integrity and memory. Neuronal LRP6 deficiency in an amyloid mouse model also leads to exacerbated amyloid pathology due to increased APP processing to amyloid-β. In humans, LRP6 and Wnt signaling are significantly downregulated in AD brains, likely by a mechanism that depends on amyloid-β. Our results define a critical pathway in which decreased LRP6-mediated Wnt signaling, synaptic dysfunction, and elevated Aβ synergistically accelerate AD progression and suggest that restoring LRP6-mediated Wnt signaling can be explored as a viable strategy for AD therapy.