Aβ Uptake Research Articles

Probing the Role of a Regional Quantitative Assessment of Amyloid PET

In clinical practice, the amy-PET is globally inspected to provide a binary outcome, but the role of a regional assessment has not been fully investigated yet. To deepen the role of regional amyloid burden and its implication on clinical-neuropsychological features. Amy-PET and a complete neuropsychological assessment (Trail Making Test, Rey Auditory Verbal Learning Test, semantic verbal fluency, Symbol Digit, Stroop, visuoconstruction) were available in 109 patients with clinical suspicion of Alzheimer's disease. By averaging the standardized uptake value ratio and ELBA, a regional quantification was calculated for each scan. Patients were grouped according to their overall amyloid load: correlation maps, based on regional quantification, were calculated and compared. A regression analysis between neuropsychological assessment and the regional amyloid-β (Aβ) load was carried out. Significant differences were observed between the correlation maps of patients at increasing levels of Aβ and the overall dataset. The Aβ uptake of the subcortical gray matter resulted not related to other brain regions independently of the global Aβ level. A significant association of semantic verbal fluency was observed with ratios of cortical and subcortical distribution of Aβ which represent a coarse measure of differences in regional distribution of Aβ. Our observations confirmed the different susceptibility to Aβ accumulation among brain regions. The association between cognition and Aβ distribution deserves further investigations: it is possibly due to a direct local effect or it represents a proxy marker of a more aggressive disease subtype. Regional Aβ assessment represents an available resource on amy-PET scan with possibly clinical and prognostic implications.

Early neuroinflammation is associated with lower amyloid and tau levels in cognitively normal older adults

CNS inflammation is a key factor in Alzheimer’s Disease (AD), but its relation to pathological Aβ, tau, and APOE4 is poorly understood, particularly prior to the onset of cognitive symptoms. To better characterize early relationships between inflammation, APOE4, and AD pathology, we assessed correlations between cerebrospinal fluid (CSF) inflammatory markers and brain levels of Aβ and tau in cognitively normal older adults.Each participant received a lumbar puncture to collect and quantify CSF levels of TNFα, IL-6, IL-8, and IL-10, a T1-weighted MRI, and PET scanning with [18F]flortaucipir (FTP; n = 57), which binds to tau tangles and/or [18F]florbetapir (FBP; n = 58), which binds to Aβ. Parallel voxelwise regressions assessed relationships between each CSF inflammatory marker and FTP and FBP SUVR, as well as APOE4*CSF inflammation interactions.Unexpectedly, we detected significant negative associations between regional Aβ and tau PET uptake and CSF inflammatory markers. For Aβ PET, we detected negative associations with CSF IL-6 and IL-8 in regions known to show early accumulation of Aβ (i.e. lateral and medial frontal lobes). For tau PET, negative relationships were observed with CSF TNFα and IL-8, predominantly in regions known to exhibit early tau accumulation (i.e. medial temporal lobe). In subsequent analyses, significant interactions between APOE4 status and IL-8 on Aβ and tau PET levels were observed in spatially distinct regions from those showing CSF–Aβ/tau relationships.Results from the current cross-sectional study support previous findings that neuroinflammation may be protective against AD pathology at a given stage of the disease, and extend these findings to a cognitively normal aging population. This study provides new insight into a dynamic relationship between neuroinflammation and AD pathology and may have implications for whom and when neuroinflammatory therapies may be appropriate.

IPSC-Derived Microglia for Modeling Human-Specific DAMP and PAMP Responses in the Context of Alzheimer's Disease.

Neuroinflammation in Alzheimer’s disease (AD) has been the focus for identifying targetable pathways for drug development. The role of amyloid beta (Aβ), a prototype of damage-associated molecular patterns (DAMPs), has been implicated in triggering an inflammatory response. As alpha7 nicotinic acetylcholine receptor (α7 nAChR) binds Aβ with high affinity, α7 nAChR may play a role in Aβ-induced neuroinflammation. The conundrum of how α7 nAChR as the mediator of the cholinergic anti-inflammatory response may trigger an inflammatory response has not been resolved. CHRFAM7A, the uniquely human fusion gene between ULK4 and CHRNA7, is a negative regulator of α7 nAChR ionotropic function. To provide the human context, isogenic induced pluripotent stem cell (iPSC) lines were developed from CHRFAM7A null and carrier individuals by genome-editing the null line using TALENs to knock-in CHRFAM7A. In iPSC-derived microglia-like cells, CHRFAM7A mitigated Aβ uptake through the α7 nAChR. Despite the lower Aβ uptake, the presence of CHRFAM7A was associated with an innate immune response that was characterized by NF-κB activation and NF-κB target transcription (TNFA, IL6, and IL1B). LPS, a prototype PAMP, induced a heightened immune response in CHRFAM7A carriers. CHRFAM7A modified the dynamics of NF-κB translocation by prolonging its nuclear presence. CHRFAM7A modified the α7 nAChR metabotropic function, resulting in a human-specific innate immune response. This iPSC model provided an opportunity to elucidate the mechanism and establish high throughput screens.

Amyloid-beta uptake by blood monocytes is reduced with ageing and Alzheimer\u2019s disease

Deficits in the clearance of amyloid β-protein (Aβ) play a pivotal role in the pathogenesis of sporadic Alzheimer’s disease (AD). The roles of blood monocytes in the development of AD remain unclear. In this study, we sought to investigate the alterations in the Aβ phagocytosis function of peripheral monocytes during ageing and in AD patients. A total of 104 cognitively normal participants aged 22–89 years, 24 AD patients, 25 age- and sex-matched cognitively normal (CN) subjects, 15 Parkinson’s disease patients (PD), and 15 age- and sex-matched CN subjects were recruited. The Aβ uptake by blood monocytes was measured and its alteration during ageing and in AD patients were investigated. Aβ1-42 uptake by monocytes decreased during ageing and further decreased in AD but not in PD patients. Aβ1-42 uptake by monocytes was associated with Aβ1-42 levels in the blood. Among the Aβ uptake-related receptors and enzymes, the expression of Toll-like receptor 2 (TLR2) was reduced in monocytes from AD patients. Our findings suggest that monocytes regulate the blood levels of Aβ and might be involved in the development of AD. The recovery of the Aβ uptake function by blood monocytes represents a potential therapeutic strategy for AD.

Elucidating the role of SORL1 as an APOE receptor using iPSC‐derived astrocytes

AbstractBackgroundApolipoprotein E (APOE), the strongest genetic risk factor in late‐onset AD (LOAD), encodes a protein that can interact with Aβ and regulate its clearance. Interestingly, SORL1, another AD risk gene, belongs to the family of APOE receptors involved in Aβ clearance. SORL1 is the only APOE receptor gene that has been identified as a genetic risk factor in LOAD. Recent studies identified SORL1 loss‐of‐function variants in familial AD patients, suggesting that SORL1 may be a causal gene for AD. Here, we investigated the potential convergent mechanisms of APOE and SORL1 function in the brain by examining the novel function of SORL1 in human astrocytes in Aβ clearance.MethodiPSC‐derived astrocytes were generated from individuals with 1) different APOE isoforms 2) SORL1 knock out (KO), risk SNPs and missense mutations and 3) other AD risk SNPs using two astrocyte differentiation protocols. Aβ clearance assays were performed by treating the astrocytes with synthetic or cell derived Aβ, then monitoring the Aβ level in both media and lysate over time to determine the ability of astrocytes to uptake and clear Aβ.ResultWe have generated iPSC‐derived astrocytes from multiple lines to characterize whether SORL1 functions as an APOE receptor in human astrocytes. We determined that SORL1 is highly expressed in astrocytes, both in cytoplasmic and cell surface compartments, and SORL1 can interact with APOE in iPSCs as shown by co‐immunoprecipitation. Further, our preliminary data show that SORL1 KO astrocytes show elevated endogenous extracellular Aβ levels, and impaired Aβ clearance ability when treated with exogenous or cell derived Aβ. Lastly, when Aβ uptake was examined using APOE isogenic astrocytes, APOE KO show impaired clearance, supporting a role for endogenously secreted APOE in Aβ uptake in human astrocytes. Ongoing studies are interrogating the contribution of SORL1 in APOE‐mediated Aβ clearance.ConclusionHere, I have utilized iPSC‐derived astrocytes to examine the novel role of SORL1 as an APOE receptor in astrocytes, and investigate a potential convergent mechanism that APOE and SORL1 play in Aβ clearance.

Therapeutic potential of niacin in Alzheimer's disease

AbstractBackgroundDietary intake of niacin reduces the risk of Alzheimer's disease (AD) and protects against age‐related cognitive decline. Niacin is a ligand for the niacin receptor HCAR2, which, in the brain, is selectively expressed by microglia. HCAR2 activation has been shown to be neuroprotective in stroke and demyelination models. Therefore, we hypothesized that niacin administration could have beneficial effects in AD pathology.Method5 month‐old 5xFAD mice, an animal model of AD, were treated with a FDA‐approved formulation of niacin (Niaspan®) daily, by oral gavage for 30 days with 100 mg niacin/kg. After treatment, the animals were sacrificed and the brains were collected for immunohistochemistry and biochemical analysis. Gene expression and Aβ uptake efficiency were analyzed in primary cultures of microglia incubated with 1.25 nM of Niaspan®.ResultOur results indicate there is an induction of HCAR2 expression in the AD brain, prominently in activated microglia. Niaspan® treatment significantly reduced Aβ burden and number of compact plaques in 5xFAD brain. Accordingly, Niaspan® induced phagocytic genes and Aβ uptake in primary cultures of microglia. Importantly, Niaspan® exerted a neuroprotective effect evidenced by reduced neuronal loss in the subiculum and cortical layer 5 of Niaspan®‐treated mice, accompanied by a rescue of working memory deficits.ConclusionThese preliminary data suggest that Niaspan® treatment has neuroprotective effects in AD, even after the onset of severe amyloid pathology, through an induction of Aβ cleareance by microglia. Niaspan® is a FDA‐approved drug, thus there is a translational potential of this strategy into clinical practice, supporting further study of this therapeutic approach.

Retention Period of Amyloid β1-42 in the Brain Extracellular Fluid as the Toxicological Determinant in Freely Moving Rats.

The pathological significance of amyloid-β1-42 (Aβ1-42) dynamics is poorly understood in the brain extracellular compartment. Here we test which of the concentration or the retention is critical for Aβ1-42 toxicity after injection of equal dose into dentate granule cell layer of freely moving rats. The toxicity of Aβ1-42 (25 µM) was compared between injections at the rate of 0.25 µL/min for 4 min (fast injection) and 0.025 µL/min for 40 min (slow injection). Dentate gyrus long-term potentiation (LTP) was affected 1 and 2 h after the fast injection, but not 4 h. In contrast, LTP was affected even 72 h after the slow injection. Aβ1-42 staining 5 min after finish of the slow injection was more intense in the dentate granule cell layer than of the fast injection. The present study indicates that the retention of Aβ1-42 in the extracellular fluid is correlated with neuronal Aβ1-42 uptake and plays a key role in Aβ1-42 neurotoxicity. In the extracellular fluid of the dentate gyrus, the retention period of Aβ1-42 is much more critical for Aβ1-42 toxicity than Aβ1-42 concentration. It is likely that Aβ1-42 toxicity is accelerated by the disturbance of Aβ1-42 metabolism in the dentate gyrus.

Cerebrovascular amyloid angiopathy in bioengineered vessels is reduced by high‐density lipoprotein particles enriched in apolipoprotein E

AbstractBackgroundSeveral lines of evidence suggest that high‐density lipoprotein (HDL) reduces Alzheimer’s disease (AD) risk by decreasing vascular beta‐amyloid (Aβ) deposition and inflammation, however, the mechanisms by which HDL improve cerebrovascular functions relevant to AD remain poorly understood.MethodsHere we use a human bioengineered model of cerebral amyloid angiopathy (CAA) to define several mechanisms by which HDL reduces Aβ deposition within the vasculature and attenuates endothelial inflammation as measured by monocyte binding.ResultsWe demonstrate that HDL reduces vascular Aβ accumulation independently of its principal binding protein, scavenger receptor (SR)‐BI, in contrast to the SR‐BI‐dependent mechanism by which HDL prevents Aβ‐induced vascular inflammation. We describe multiple novel mechanisms by which HDL acts to reduce CAA, namely: i) altering Aβ binding to collagen‐I, ii) forming a complex with Aβ that maintains its solubility, iii) lowering collagen‐I protein levels produced by smooth‐muscle cells (SMC), and iv) attenuating Aβ uptake into SMC that associates with reduced low density lipoprotein related protein 1 (LRP1) levels. Furthermore, we show that HDL particles enriched in apolipoprotein (apo)E appear to be the major drivers of these effects, providing new insights into the peripheral role of apoE in AD, in particular, the fraction of HDL that contains apoE.ConclusionThe findings in this study identify new mechanisms by which circulating HDL, particularly HDL particles enriched in apoE, may provide vascular resilience to Aβ and shed new light on a potential role of peripherally‐acting apoE in AD.

Maresin 1 attenuates pro-inflammatory activation induced by β-amyloid and stimulates its uptake.

Alzheimer's disease (AD) is the most common dementia, characterized by pathological accumulation of β‐amyloid (Aβ) and hyperphosphorylation of tau protein, together with a damaging chronic inflammation. The lack of effective treatments urgently warrants new therapeutic strategies. Resolution of inflammation, associated with beneficial and regenerative activities, is mediated by specialized pro‐resolving lipid mediators (SPMs) including maresin 1 (MaR1). Decreased levels of MaR1 have been observed in AD brains. However, the pro‐resolving role of MaR1 in AD has not been fully investigated. In the present study, human monocyte‐derived microglia (MdM) and a differentiated human monocyte cell line (THP‐1 cells) exposed to Aβ were used as models of AD neuroinflammation. We have studied the potential of MaR1 to inhibit pro‐inflammatory activation of Aβ and assessed its ability to stimulate phagocytosis of Aβ42. MaR1 inhibited the Aβ42‐induced increase in cytokine secretion and stimulated the uptake of Aβ42 in both MdM and differentiated THP‐1 cells. MaR1 was also found to decrease chemokine secretion and reduce the associated increase in the activation marker CD40. Activation of kinases involved in transduction of inflammation was not affected by MaR1, but the activity of nuclear factor (NF)‐κB was decreased. Our data show that MaR1 exerts effects that indicate a pro‐resolving role in the context of AD and thus presents itself as a potential therapeutic target for AD.

Severe Obstructive Sleep Apnea and Increased Cortical Amyloid-β Deposition.

The suggested association between severe obstructive sleep apnea (OSA) and risk of Alzheimer's disease (AD) needs further study. Only few recent reports exist on associations between brain amyloid-β (Aβ) burden and severe OSA in middle-aged patients. Examine the possible presence of cortical Aβ accumulation in middle-aged patients with severe OSA. We performed detailed multimodal neuroimaging in 19 cognitive intact patients (mean 44.2 years) with severe OSA (Apnea-Hypopnea Index >30 h-1). Known etiological factors for possible Aβ accumulation were used as exclusion criteria. Aβ uptake was studied with [11C]-PiB-PET, glucose metabolism with [18F]-FDG-PET, and structural imaging with 3.0T MRI. When analyzed individually, in [11C]-PiB-PET a substantial number (∼32%) of the patients exhibited statistically significant evidence of increased cortical Aβ uptake based on elevated regional Z-score values, mostly seen bilaterally in the precuneus and posterior cingulum regions. Cortical glucose hypometabolism in [18F]-FDG-PET was seen in two patients. MRI did not show structural changes suggestive of AD-related pathology. Increased [11C]-PiB uptake was seen in middle-aged cognitively intact patients with severe OSA. These findings are similar to those described in cognitive unimpaired older OSA patients. The changes in cortical Aβ uptake suggest that severe OSA itself may predispose to alterations related to AD already in middle-age. Aβ clearance may be compromised without simultaneous evidence of metabolic or structural alterations. The results emphasize the importance of early diagnostics and proper treatment of severe OSA in cognitively intact middle-aged subjects, possibly diminishing the individual risk for later cognitive dysfunction.

Evidence for aggregation-independent, PrPC-mediated Aβ cellular internalization

Evidence linking amyloid beta (Aβ) cellular uptake and toxicity has burgeoned, and mechanisms underlying this association are subjects of active research. Two major, interconnected questions are whether Aβ uptake is aggregation-dependent and whether it is sequence-specific. We recently reported that the neuronal uptake of Aβ depends significantly on peptide chirality, suggesting that the process is predominantly receptor-mediated. Over the past decade, the cellular prion protein (PrPC) has emerged as an important mediator of Aβ-induced toxicity and of neuronal Aβ internalization. Here, we report that the soluble, nonfibrillizing Aβ (1-30) peptide recapitulates full-length Aβ stereoselective cellular uptake, allowing us to decouple aggregation from cellular, receptor-mediated internalization. Moreover, we found that Aβ (1-30) uptake is also dependent on PrPC expression. NMR-based molecular-level characterization identified the docking site on PrPC that underlies the stereoselective binding of Aβ (1-30). Our findings therefore identify a specific sequence within Aβ that is responsible for the recognition of the peptide by PrPC, as well as PrPC-dependent cellular uptake. Further uptake stereodifferentiation in PrPC-free cells points toward additional receptor-mediated interactions as likely contributors for Aβ cellular internalization. Taken together, our results highlight the potential of targeting cellular surface receptors to inhibit Aβ cellular uptake as an alternative route for future therapeutic development for Alzheimer's disease.

CHRFAM7A: A human specific fusion gene, accounts for the translational gap for cholinergic strategies in Alzheimer's disease.

BackgroundCholinergic neuronal loss is one of the hallmarks of AD related neurodegeneration; however, preclinical promise of α7 nAChR drugs failed to translate into humans. CHRFAM7A, a uniquely human fusion gene, is a negative regulator of α7 nAChR and was unaccounted for in preclinical models.MethodsMolecular methods: Function of CHRFAM7A alleles was studied in vitro in two disease relevant phenotypic readouts: electrophysiology and Aβ uptake. Genome edited human induced pluripotent stem cells (iPSC) were used as a model system with the human context. Double blind pharmacogenetic study: We performed double-blind pharmacogenetic analysis on the effect of AChEI therapy based on CHRFAM7A carrier status in two paradigms: response to drug initiation and DMT effect. Mini Mental Status Examination (MMSE) was used as outcome measure. Change in MMSE score from baseline was compared by 2-tailed T-test. Longitudinal analysis of clinical outcome (MMSE) was performed using a fitted general linear model, based on an assumed autoregressive covariance structure. Model independent variables included age, sex, and medication regimen at the time of the first utilized outcome measure (AChEI alone or AChEI plus memantine), APOE4 carrier status (0, 1 or 2 alleles as categorical variables) and CHRFAM7A genotype.FindingsThe direct and inverted alleles have distinct phenotypes. Functional CHRFAM7A allele classifies the population as 25% non-carriers and 75% carriers. Induced pluripotent stem cell (iPSC) models α7 nAChR mediated Aβ neurotoxicity. Pharmacological readout translates into both first exposure (p = 0.037) and disease modifying effect (p = 0.0048) in two double blind pharmacogenetic studies.InterpretationCHRFAM7A accounts for the translational gap in cholinergic strategies in AD. Clinical trials not accounting for this uniquely human genetic factor may have rejected drug candidates that would benefit 25% of AD. Reanalyses of the completed trials using this pharmacogenetic paradigm may identify effective therapy.Funding:

The Chemokine-like Receptor 1 Deficiency Improves Cognitive Deficits of AD Mice and Attenuates Tau Hyperphosphorylation via Regulating Tau Seeding.

Pathologic features of Alzheimer's disease (AD) include accumulation of amyloid β (Aβ) and hyperphosphorylated tau protein. We have shown previously that the chemokine-like receptor 1 (CMKLR1) is a functional receptor for Aβ, and CMKLR1 contributes to the uptake of Aβ. However, it is unclear whether CMKLR1 ameliorates or aggravates the process of AD. Here, we show that deletion of the gene coding for CMKLR1 significantly increased Aβ deposits in brains of both male and female amyloid β precursor protein/presenilin-1 mice. However, it markedly decreased the mortality of these mice. Behavioral studies found that CMKLR1 deficiency improved cognitive impairment of male and female amyloid β precursor protein/presenilin-1 mice and intracerebroventricular-streptozotocin injection AD mice. We further explored the effect of CMKLR1 on tau pathology. We found that CMKLR1 deficiency or inhibition attenuated the hyperphosphorylation of tau in brains of AD mice in vivo and in the neuronal cells in vitro The expression of CMKLR1 on the neurons affected tau phosphorylation by participating in tau seeding. Together, these results uncover a novel mechanism of CMKLR1 in the pathologic process of AD and suggest that inhibiting the promotion effect of CMKLR1 on tau seeding may provide a new strategy for the treatment of AD.SIGNIFICANCE STATEMENT Evidence suggests that inflammation is involved in the pathologic progression of AD. The chemokine-like receptor 1 (CMKLR1), belonging to the family of GPCRs, is able to bind and uptake amyloid β. We show here, for the first time, that, although CMKLR1 deficiency increased amyloid β deposits in AD mice, it reduced the mortality and improved the cognitive deficits of AD mice. We furthermore show that CMKLR1 deficiency or inhibition attenuated tau hyperphosphorylation in brains of AD model mice in vivo and in neuronal cells in vitro Finally, we first discovered that the expression of CMKLR1 on neurons affected tau phosphorylation by participating in tau seeding. These findings suggest that inhibition of CMKLR1 may provide a new strategy for the treatment of AD.

The Impact of Ag Nanoparticles and CdTe Quantum Dots on Expression and Function of Receptors Involved in Amyloid-β Uptake by BV-2 Microglial Cells.

Microglial cells clear the brain of pathogens and harmful debris, including amyloid-β (Aβ) deposits that are formed during Alzheimer’s disease (AD). We studied the expression of Msr1, Ager and Cd36 receptors involved in Aβ uptake and expression of Cd33 protein, which is considered a risk factor in AD. The effect of silver nanoparticles (AgNP) and cadmium telluride quantum dots (CdTeQD) on the expression of the above receptors and Aβ uptake by microglial cells was investigated. Absorption of Aβ and NP was confirmed by confocal microscopy. AgNP, but not CdTeQD, caused a decrease in Aβ accumulation. By using a specific inhibitor—polyinosinic acid—we demonstrated that Aβ and AgNP compete for scavenger receptors. Real-time PCR showed up-regulation of Cd33 and Cd36 gene expression after treatment with CdTeQD for 24 h. Analysis of the abundance of the receptors on the cell surface revealed that AgNP treatment significantly reduced the presence of Msr1, Cd33, Ager and Cd36 receptors (6 and 24 h), whereas CdTeQD increased the levels of Msr1 and Cd36 (24 h). To summarize, we showed that AgNP uptake competes with Aβ uptake by microglial cells and consequently can impair the removal of the aggregates. In turn, CdTeQD treatment led to the accumulation of proinflammatory Cd36 protein on the cell surface.

Effects of IL-34 on Macrophage Immunological Profile in Response to Alzheimer's-Related Aβ42 Assemblies.

Interleukin-34 (IL-34) is a recently discovered cytokine that acts as a second ligand of the colony stimulating factor 1 receptor (CSF1R) in addition to macrophage colony-stimulating factor (M-CSF). Similar to M-CSF, IL-34 also stimulates bone marrow (BM)-derived monocyte survival and differentiation into macrophages. Growing evidence suggests that peripheral BM-derived monocyte/macrophages (BMMO) play a key role in the physiological clearance of cerebral amyloid β-protein (Aβ). Aβ42 forms are especially neurotoxic and highly associated with Alzheimer's disease (AD). As a ligand of CSF1R, IL-34 may be relevant to innate immune responses in AD. To investigate how IL-34 affects macrophage phenotype in response to structurally defined and stabilized Aβ42 oligomers and preformed fibrils, we characterized murine BMMO cultured in media containing M-CSF, IL-34, or regimens involving both cytokines. We found that the immunological profile and activation phenotype of IL-34-stimulated BMMO differed significantly from those cultured with M-CSF alone. Specifically, macrophage uptake of fibrillar or oligomeric Aβ42 was markedly reduced following exposure to IL-34 compared to M-CSF. Surface expression of type B scavenger receptor CD36, known to facilitate Aβ recognition and uptake, was modified following treatment with IL-34. Similarly, IL-34 macrophages expressed lower levels of proteins involved in both Aβ uptake (triggering receptor expressed on myeloid cells 2, TREM2) as well as Aβ-degradation (matrix metallopeptidase 9, MMP-9). Interestingly, intracellular compartmentalization of Aβ visualized by staining of early endosome antigen 1 (EEA1) was not affected by IL-34. Macrophage characteristics associated with an anti-inflammatory and pro-wound healing phenotype, including processes length and morphology, were also quantified, and macrophages stimulated with IL-34 alone displayed less process elongation in response to Aβ42 compared to those cultured with M-CSF. Further, monocytes treated with IL-34 alone yielded fewer mature macrophages than those treated with M-CSF alone or in combination with IL-34. Our data indicate that IL-34 impairs monocyte differentiation into macrophages and reduces their ability to uptake pathological forms of Aβ. Given the critical role of macrophage-mediated Aβ clearance in both murine models and patients with AD, future work should investigate the therapeutic potential of modulating IL-34 in vivo to increase macrophage-mediated Aβ clearance and prevent disease development.

The Sigma-2 Receptor/TMEM97, PGRMC1, and LDL Receptor Complex Are Responsible for the Cellular Uptake of Aβ42 and Its Protein Aggregates

Our lab has recently shown that the Sigma-2 Receptor/Transmembrane Protein 97 (TMEM97) and Progesterone Receptor Membrane Component 1 (PGRMC1) form a complex with the Low Density Lipoprotein Receptor (LDLR), and this intact complex is required for efficient uptake of lipoproteins such as LDL and apolipoprotein E (apoE). These receptors are expressed in the nervous system where they have implications in neurodegenerative diseases such as Alzheimer's disease (AD), where apoE is involved in neuronal uptake and accumulation of Aβ42, eventually cascading into neurodegeneration, synaptic dysfunction, and ultimately, dementia. We hypothesize that the intact Sigma-2 receptor complex-TMEM97, PGRMC1, and LDLR-is necessary for internalization of apoE and Aβ42 monomers (mAβ42) and oligomers (oAβ42), and the disruption of the receptor complex inhibits uptake. The results of this study suggest that the intact Sigma-2 receptor complex is a binding site for mAβ42 and oAβ42, in the presence or absence of apoE2, apoE3, and apoE4. The loss or pharmacological inhibition of one or both of these proteins results in the disruption of the complex leading to decreased uptake of mAβ42 and oAβ42 and apoE in primary neurons. The TMEM97, PGRMC1, and LDLR complex is a pathway for the cellular uptake of Aβ42 via apoE dependent and independent mechanisms. This study suggests that the complex may potentially be a novel pharmacological target to decrease neuronal Aβ42 internalization and accumulation, which may represent a new strategy for inhibiting the rate of neurotoxicity, neurodegeneration, and progression of AD.

Aquaporin-4 polymorphisms predict amyloid burden and clinical outcome in the Alzheimer's disease spectrum

Clearance of amyloid-β (Aβ) from the brain is hypothesized to be mediated by the glymphatic system through aquaporin-4 (AQP4) water channels. Genetic variation of AQP4 may impact water channel function, Aβ clearance, and clinical outcomes. We examined whether single-nucleotide polymorphisms (SNPs) of the AQP4 gene were related to Aβ neuropathology on [18F]Florbetapir PET in 100 Aβ positive late mild cognitive impairment (LMCI) or Alzheimer's disease (AD) patients and were predictive of clinical outcome in prodromal AD patients. AQP4 SNP rs72878794 was associated with decreased Aβ uptake, whereas rs151244 was associated with increased Aβ uptake, increased risk of conversion from MCI and LMCI to AD, and an increased 4-year rate of cognitive decline in LMCI. AQP4 genetic variation was associated with Aβ accumulation, disease stage progression, and cognitive decline. This variation may correspond to changes in glymphatic system functioning and brain Aβ clearance and could be a useful biomarker in predicting disease burden for those on the dementia spectrum.

Role of Membrane Tension Sensitive Endocytosis and Rho GTPases in the Uptake of the Alzheimer's Disease Peptide Aβ(1-42).

Intraneuronal accumulation of amyloid-β (Aβ) is an early pathological signum of Alzheimer’s disease, and compartments of the endolysosomal system have been implicated in both seeding and cell–cell propagation of Aβ aggregation. We have studied how clathrin-independent mechanisms contribute to Aβ endocytosis, exploring pathways that are sensitive to changes in membrane tension and the regulation of Rho GTPases. Using live cell confocal microscopy and flow cytometry, we show the uptake of monomeric Aβ(1-42) into endocytic vesicles and vacuole-like dilations, following relaxation of osmotic pressure-induced cell membrane tension. This indicates Aβ(1-42) uptake via clathrin independent carriers (CLICs), although overexpression of the bar-domain protein GRAF1, a key regulator of CLICs, had no apparent effect. We furthermore report reduced Aβ(1-42) uptake following overexpression of constitutively active forms of the Rho GTPases Cdc42 and RhoA, whereas modulation of Rac1, which is linked to macropinosome formation, had no effect. Our results confirm that uptake of Aβ(1-42) is clathrin- and dynamin-independent and point to the involvement of a new and distinct clathrin-independent endocytic mechanism which is similar to uptake via CLICs or macropinocytosis but that also appear to involve yet uncharacterized molecular players.

A Role of Low-Density Lipoprotein Receptor-Related Protein 4 (LRP4) in Astrocytic Aβ Clearance.

Amyloid-β (Aβ) deposition occurs years before cognitive symptoms appear and is considered a cause of Alzheimer's disease (AD). The imbalance of Aβ production and clearance leads to Aβ accumulation and Aβ deposition. Increasing evidence indicates an important role of astrocytes, the most abundant cell type among glial cells in the brain, in Aβ clearance. We explored the role of low-density lipoprotein receptor-related protein 4 (LRP4), a member of the LDLR family, in AD pathology. We show that Lrp4 is specifically expressed in astrocytes and its levels in astrocytes were higher than those of Ldlr and Lrp1, both of which have been implicated in Aβ uptake. LRP4 was reduced in postmortem brain tissues of AD patients. Genetic deletion of the Lrp4 gene augmented Aβ plaques in 5xFAD male mice, an AD mouse model, and exacerbated the deficits in neurotransmission, synchrony between the hippocampus and PFC, and cognition. Mechanistically, LRP4 promotes Aβ uptake by astrocytes likely by interacting with ApoE. Together, our study demonstrates that astrocytic LRP4 plays an important role in Aβ pathology and cognitive function.SIGNIFICANCE STATEMENT This study investigates how astrocytes, a type of non-nerve cells in the brain, may contribute to Alzheimer's disease (AD) development. We demonstrate that the low-density lipoprotein receptor-related protein 4 (LRP4) is reduced in the brain of AD patients. Mimicking the reduced levels in an AD mouse model exacerbates cognitive impairment and increases amyloid aggregates that are known to damage the brain. We show that LRP4 could promote the clearance of amyloid protein by astrocytes. Our results reveal a previously unappreciated role of LRP4 in AD development.

Involvement of a Na+-coupled Oligopeptide Transport System for β-amyloid Peptide (Aβ1–42) in Brain Cells

A Na+-coupled transport system in mammalian cells is responsible for the uptake of oligopeptides consisting of 5 or more amino acids. Here we investigated if this transport system is expressed in brain cells and transports the 42-amino-acid β-amyloid peptide (Aβ1-42). The human and mouse neuronal cell lines SK-N-SH and HT22, human microglial cell line HMC-3, and human blood-brain barrier endothelial cell line hCMEC/D3 were used to monitor the uptake of [3H]-deltorphin II (a heptapeptide) and fluorescence-labeled Aβ1-42. All four cell lines exhibited Na+-coupled uptake of deltorphin II. Aβ1-42 competed with deltorphin II for the uptake. Uptake of fluorescence-labeled Aβ1-42 was detectable in these cell lines, and the uptake was Na+-dependent and inhibitable by deltorphin II. The Na+-coupled uptake disappeared at high concentrations of Aβ1-42 due to oligomerization of the peptide. Exposure of the cells to excess iron abolished the uptake. In hCMEC/D3 cells cultured on Transwell filters, the uptake was localized preferentially to the abluminal membrane. A Na+-coupled transport system mediates the uptake of Aβ1-42 monomers in neuronal and microglial cells. The same system is also responsible for the uptake of Aβ1-42 from brain into blood-brain barrier endothelial cells. These findings have relevance to Alzheimer's disease.