Retinal peri-arteriolar versus peri-venular amyloidosis, hippocampal atrophy, and cognitive impairment: exploratory trial.

The relationship between amyloidosis and vasculature in cognitive impairment and Alzheimer's disease (AD) pathogenesis is increasingly acknowledged. We conducted a quantitative and topographic assessment of retinal perivascular amyloid plaque (AP) distribution in individuals with both normal and impaired cognition. Using a retrospective dataset of scanning laser ophthalmoscopy fluorescence images from twenty-eight subjects with varying cognitive states, we developed a novel image processing method to examine retinal peri-arteriolar and peri-venular curcumin-positive AP burden. We further correlated retinal perivascular amyloidosis with neuroimaging measures and neurocognitive scores. Our study unveiled that peri-arteriolar AP counts surpassed peri-venular counts throughout the entire cohort (P < 0.0001), irrespective of the primary, secondary, or tertiary vascular branch location, with a notable increase among cognitively impaired individuals. Moreover, secondary branch peri-venular AP count was elevated in the cognitively impaired (P < 0.01). Significantly, peri-venular AP count, particularly in secondary and tertiary venules, exhibited a strong correlation with clinical dementia rating, Montreal cognitive assessment score, hippocampal volume, and white matter hyperintensity count. In conclusion, our exploratory analysis detected greater peri-arteriolar versus peri-venular amyloidosis and a marked elevation of amyloid deposition in secondary branch peri-venular regions among cognitively impaired subjects. These findings underscore the potential feasibility of retinal perivascular amyloid imaging in predicting cognitive decline and AD progression. Larger longitudinal studies encompassing diverse populations and AD-biomarker confirmation are warranted to delineate the temporal-spatial dynamics of retinal perivascular amyloid deposition in cognitive impairment and the AD continuum.

Alzheimer's disease pathophysiology in the Retina.

The retina is an emerging CNS target for potential noninvasive diagnosis and tracking of Alzheimer's disease (AD). Studies have identified the pathological hallmarks of AD, including amyloid ß-protein (Aß) deposits and abnormal tau protein isoforms, in the retinas of AD patients and animal models. Moreover, structural and functional vascular abnormalities such as reduced blood flow, vascular Aß deposition, and blood-retinal barrier damage, along with inflammation and neurodegeneration, have been described in retinas of patients with mild cognitive impairment and AD dementia. Histological, biochemical, and clinical studies have demonstrated that the nature and severity of AD pathologies in the retina and brain correspond. Proteomics analysis revealed a similar pattern of dysregulated proteins and biological pathways in the retina and brain of AD patients, with enhanced inflammatory and neurodegenerative processes, impaired oxidative-phosphorylation, and mitochondrial dysfunction. Notably, investigational imaging technologies can now detect AD-specific amyloid deposits, as well as vasculopathy and neurodegeneration in the retina of living AD patients, suggesting alterations at different disease stages and links to brain pathology. Current and exploratory ophthalmic imaging modalities, such as optical coherence tomography (OCT), OCT-angiography, confocal scanning laser ophthalmoscopy, and hyperspectral imaging, may offer promise in the clinical assessment of AD. However, further research is needed to deepen our understanding of AD's impact on the retina and its progression. To advance this field, future studies require replication in larger and diverse cohorts with confirmed AD biomarkers and standardized retinal imaging techniques. This will validate potential retinal biomarkers for AD, aiding in early screening and monitoring.

Distinctive retinal peri-arteriolar versus peri-venular amyloid plaque distribution correlates with the cognitive performance.

Introduction: The vascular contribution to Alzheimer's disease (AD) is tightly connected to cognitive performance across the AD continuum. We topographically describe retinal perivascular amyloid plaque (AP) burden in subjects with normal or impaired cognition. Methods: Using scanning laser ophthalmoscopy, we quantified retinal peri-arteriolar and peri-venular curcumin-positive APs in the first, secondary and tertiary branches in twenty-eight subjects. Perivascular AP burden among cognitive states was correlated with neuroimaging and cognitive measures. Results: Peri-arteriolar exceeded peri-venular AP count (p<0.0001). Secondary branch AP count was significantly higher in cognitively impaired (p<0.01). Secondary small and tertiary peri-venular AP count strongly correlated with clinical dementia rating, hippocampal volumes, and white matter hyperintensity count. Discussion: Our topographic analysis indicates greater retinal amyloid accumulation in the retinal peri-arteriolar regions overall, and distal peri-venular regions in cognitively impaired individuals. Larger longitudinal studies are warranted to understand the temporal-spatial relationship between vascular dysfunction and perivascular amyloid deposition in AD. Highlights: Retinal peri-arteriolar region exhibits more amyloid compared with peri-venular regions.Secondary retinal vascular branches have significantly higher perivascular amyloid burden in subjects with impaired cognition, consistent across sexes.Cognitively impaired individuals have significantly greater retinal peri-venular amyloid deposits in the distal small branches, that correlate with CDR and hippocampal volumes.

Identification of retinal tau oligomers, citrullinated tau, and other tau isoforms in early and advanced AD and relations to disease status.

Importance: This study identifies and quantifies diverse pathological tau isoforms in the retina of both early and advanced-stage Alzheimer's disease (AD) and determines their relationship with disease status. Objective: A case-control study was conducted to investigate the accumulation of retinal neurofibrillary tangles (NFTs), paired helical filament (PHF)-tau, oligomeric tau (oligo-tau), hyperphosphorylated tau (p-tau), and citrullinated tau (Cit-tau) in relation to the respective brain pathology and cognitive dysfunction in mild cognitively impaired (MCI) and AD dementia patients versus normal cognition (NC) controls. Design setting and participants: Eyes and brains from donors diagnosed with AD, MCI (due to AD), and NC were collected (n=75 in total), along with clinical and neuropathological data. Brain and retinal cross-sections-in predefined superior-temporal and inferior-temporal (ST/IT) subregions-were subjected to histopathology analysis or Nanostring GeoMx digital spatial profiling. Main outcomes and measure: Retinal burden of NFTs (pretangles and mature tangles), PHF-tau, p-tau, oligo-tau, and Cit-tau was assessed in MCI and AD versus NC retinas. Pairwise correlations revealed associations between retinal and brain parameters and cognitive status. Results: Increased retinal NFTs (1.8-fold, p=0.0494), PHF-tau (2.3-fold, p<0.0001), oligo-tau (9.1-fold, p<0.0001), CitR 209 -tau (4.3-fold, p<0.0001), pSer202/Thr205-tau (AT8; 4.1-fold, p<0.0001), and pSer396-tau (2.8-fold, p=0.0015) were detected in AD patients. Retinas from MCI patients showed significant increases in NFTs (2.0-fold, p=0.0444), CitR 209 -tau (3.5-fold, p=0.0201), pSer396-tau (2.6-fold, p=0.0409), and, moreover, oligo-tau (5.8-fold, p=0.0045). Nanostring GeoMx quantification demonstrated upregulated retinal p-tau levels in MCI patients at phosphorylation sites of Ser214 (2.3-fold, p=0.0060), Ser396 (1.8-fold, p=0.0052), Ser404 (2.4-fold, p=0.0018), and Thr231 (3.3-fold, p=0.0028). Strong correlations were found between retinal tau forms to paired-brain pathology and cognitive status: a) retinal oligo-tau vs. Braak stage (r=0.60, P=0.0002), b) retinal PHF-tau vs. ABC average score (r=0.64, P=0.0043), c) retinal pSer396-tau vs. brain NFTs (r=0.68, P<0.0001), and d) retinal pSer202/Thr205-tau vs. MMSE scores (r= -0.77, P=0.0089). Conclusions and Relevance: This study reveals increases in immature and mature retinal tau isoforms in MCI and AD patients, highlighting their relationship with brain pathology and cognition. The data provide strong incentive to further explore retinal tauopathy markers that may be useful for early detection and monitoring of AD staging through noninvasive retinal imaging.

The effects of enhancing angiotensin converting enzyme in myelomonocytes on ameliorating Alzheimer's-related disease and preserving cognition.

This review examines the role of angiotensin-converting enzyme (ACE) in the context of Alzheimer's disease (AD) and its potential therapeutic value. ACE is known to degrade the neurotoxic 42-residue long alloform of amyloid ß-protein (Aß42), a peptide strongly associated with AD. Previous studies in mice, demonstrated that targeted overexpression of ACE in CD115+ myelomonocytic cells (ACE10 models) improved their immune responses to effectively reduce viral and bacterial infection, tumor growth, and atherosclerotic plaque. We further demonstrated that introducing ACE10 myelomonocytes (microglia and peripheral monocytes) into the double transgenic APPSWE/PS1ΔE9 murine model of AD (AD+ mice), diminished neuropathology and enhanced the cognitive functions. These beneficial effects were dependent on ACE catalytic activity and vanished when ACE was pharmacologically blocked. Moreover, we revealed that the therapeutic effects in AD+ mice can be achieved by enhancing ACE expression in bone marrow (BM)-derived CD115+ monocytes alone, without targeting central nervous system (CNS) resident microglia. Following blood enrichment with CD115+ ACE10-monocytes versus wild-type (WT) monocytes, AD+ mice had reduced cerebral vascular and parenchymal Aß burden, limited microgliosis and astrogliosis, as well as improved synaptic and cognitive preservation. CD115+ ACE10-versus WT-monocyte-derived macrophages (Mo/MΦ) were recruited in higher numbers to the brains of AD+ mice, homing to Aß plaque lesions and exhibiting a highly Aß-phagocytic and anti-inflammatory phenotype (reduced TNFα/iNOS and increased MMP-9/IGF-1). Moreover, BM-derived ACE10-Mo/MΦ cultures had enhanced capability to phagocytose Aß42 fibrils, prion-rod-like, and soluble oligomeric forms that was associated with elongated cell morphology and expression of surface scavenger receptors (i.e., CD36, Scara-1). This review explores the emerging evidence behind the role of ACE in AD, the neuroprotective properties of monocytes overexpressing ACE and the therapeutic potential for exploiting this natural mechanism for ameliorating AD pathogenesis.

Osteopontin depletion in macrophages perturbs proteostasis via regulating UCHL1-UPS axis and mitochondria-mediated apoptosis.

Introduction: Osteopontin (OPN; also known as SPP1), an immunomodulatory cytokine highly expressed in bone marrow-derived macrophages (BMMΦ), is known to regulate diverse cellular and molecular immune responses. We previously revealed that glatiramer acetate (GA) stimulation of BMMΦ upregulates OPN expression, promoting an anti-inflammatory, pro-healing phenotype, whereas OPN inhibition triggers a pro-inflammatory phenotype. However, the precise role of OPN in macrophage activation state is unknown. Methods: Here, we applied global proteome profiling via mass spectrometry (MS) analysis to gain a mechanistic understanding of OPN suppression versus induction in primary macrophage cultures. We analyzed protein networks and immune-related functional pathways in BMMΦ either with OPN knockout (OPNKO) or GA-mediated OPN induction compared with wild type (WT) macrophages. The most significant differentially expressed proteins (DEPs) were validated using immunocytochemistry, western blot, and immunoprecipitation assays. Results and discussion: We identified 631 DEPs in OPNKO or GA-stimulated macrophages as compared to WT macrophages. The two topmost downregulated DEPs in OPNKO macrophages were ubiquitin C-terminal hydrolase L1 (UCHL1), a crucial component of the ubiquitin-proteasome system (UPS), and the anti-inflammatory Heme oxygenase 1 (HMOX-1), whereas GA stimulation upregulated their expression. We found that UCHL1, previously described as a neuron-specific protein, is expressed by BMMΦ and its regulation in macrophages was OPN-dependent. Moreover, UCHL1 interacted with OPN in a protein complex. The effects of GA activation on inducing UCHL1 and anti-inflammatory macrophage profiles were mediated by OPN. Functional pathway analyses revealed two inversely regulated pathways in OPN-deficient macrophages: activated oxidative stress and lysosome-mitochondria-mediated apoptosis (e.g., ROS, Lamp1-2, ATP-synthase subunits, cathepsins, and cytochrome C and B subunits) and inhibited translation and proteolytic pathways (e.g., 60S and 40S ribosomal subunits and UPS proteins). In agreement with the proteome-bioinformatics data, western blot and immunocytochemical analyses revealed that OPN deficiency perturbs protein homeostasis in macrophages-inhibiting translation and protein turnover and inducing apoptosis-whereas OPN induction by GA restores cellular proteostasis. Taken together, OPN is essential for macrophage homeostatic balance via the regulation of protein synthesis, UCHL1-UPS axis, and mitochondria-mediated apoptotic processes, indicating its potential application in immune-based therapies.

Retinal arterial Aß40 deposition is linked with tight junction loss and cerebral amyloid angiopathy in MCI and AD patients.

INTRODUCTION: Vascular amyloid beta (Aß) protein deposits were detected in retinas of mild cognitively impaired (MCI) and Alzheimer's disease (AD) patients. We tested the hypothesis that the retinal vascular tight junctions (TJs) were compromised and linked to disease status. METHODS: TJ components and Aß expression in capillaries and larger blood vessels were determined in post mortem retinas from 34 MCI or AD patients and 27 cognitively normal controls and correlated with neuropathology. RESULTS: Severe decreases in retinal vascular zonula occludens-1 (ZO-1) and claudin-5 correlating with abundant arteriolar Aß40 deposition were identified in MCI and AD patients. Retinal claudin-5 deficiency was closely associated with cerebral amyloid angiopathy, whereas ZO-1 defects correlated with cerebral pathology and cognitive deficits. DISCUSSION: We uncovered deficiencies in blood-retinal barrier markers for potential retinal imaging targets of AD screening and monitoring. Intense retinal arteriolar Aß40 deposition suggests a common pathogenic mechanism of failed Aß clearance via intramural periarterial drainage.

Retinal pathological features and proteome signatures of Alzheimer's disease.

Alzheimer's disease (AD) pathologies were discovered in the accessible neurosensory retina. However, their exact nature and topographical distribution, particularly in the early stages of functional impairment, and how they relate to disease progression in the brain remain largely unknown. To better understand the pathological features of AD in the retina, we conducted an extensive histopathological and biochemical investigation of postmortem retina and brain tissues from 86 human donors. Quantitative examination of superior and inferior temporal retinas from mild cognitive impairment (MCI) and AD patients compared to those with normal cognition (NC) revealed significant increases in amyloid ß-protein (Aß42) forms and novel intraneuronal Aß oligomers (AßOi), which were closely associated with exacerbated retinal macrogliosis, microgliosis, and tissue atrophy. These pathologies were unevenly distributed across retinal layers and geometrical areas, with the inner layers and peripheral subregions exhibiting most pronounced accumulations in the MCI and AD versus NC retinas. While microgliosis was increased in the retina of these patients, the proportion of microglial cells engaging in Aß uptake was reduced. Female AD patients exhibited higher levels of retinal microgliosis than males. Notably, retinal Aß42, S100 calcium-binding protein B+ macrogliosis, and atrophy correlated with severity of brain Aß pathology, tauopathy, and atrophy, and most retinal pathologies reflected Braak staging. All retinal biomarkers correlated with the cognitive scores, with retinal Aß42, far-peripheral AßOi and microgliosis displaying the strongest correlations. Proteomic analysis of AD retinas revealed activation of specific inflammatory and neurodegenerative processes and inhibition of oxidative phosphorylation/mitochondrial, and photoreceptor-related pathways. This study identifies and maps retinopathy in MCI and AD patients, demonstrating the quantitative relationship with brain pathology and cognition, and may lead to reliable retinal biomarkers for noninvasive retinal screening and monitoring of AD.

Regulating microglial miR-155 transcriptional phenotype alleviates Alzheimer's-induced retinal vasculopathy by limiting Clec7a/Galectin-3+ neurodegenerative microglia.

Single cell RNA sequencing studies identified novel neurodegeneration-associated microglial (MGnD/DAM) subtypes activated around cerebral amyloid plaques. Micro-RNA (miR)-155 of the TREM2-APOE pathway was shown to be a key transcriptional regulator of MGnD microglial phenotype. Despite growing interest in studying manifestations of Alzheimer's disease (AD) in the retina, a CNS organ accessible to noninvasive high-resolution imaging, to date MGnD microglia have not been studied in the AD retina. Here, we discovered the presence and increased populations of Clec7a+ and Galectin-3+ MGnD microglia in retinas of transgenic APPSWE/PS1L166P AD-model mice. Conditionally targeting MGnD microglia by miR-155 ablation via the tamoxifen-inducible CreERT2 system in APPSWE/PS1L166P mice diminished retinal Clec7a+ and Galectin-3+ microglial populations while increasing homeostatic P2ry12+ microglia. Retinal MGnD microglia were often adhering to microvessels; their depletion protected the inner blood-retina barrier and reduced vascular amyloidosis. Microglial miR-155 depletion further limits retinal inflammation. Mass spectrometry analysis revealed enhanced retinal PI3K-Akt signaling and predicted IL-8 and Spp1 decreases in mice with microglia-specific miR-155 knockout. Overall, this study identified MGnD microglia in APPSWE/PS1L166P mouse retina. Transcriptional regulation of these dysfunctional microglia mitigated retinal inflammation and vasculopathy. The protective effects of microglial miR-155 ablation should shed light on potential treatments for retinal inflammation and vascular damage during AD and other ocular diseases.

Label-free hyperspectral imaging and deep-learning prediction of retinal amyloid ß-protein and phosphorylated tau.

Alzheimer's disease (AD) is a major risk for the aging population. The pathological hallmarks of AD-an abnormal deposition of amyloid ß-protein (Aß) and phosphorylated tau (pTau)-have been demonstrated in the retinas of AD patients, including in prodromal patients with mild cognitive impairment (MCI). Aß pathology, especially the accumulation of the amyloidogenic 42-residue long alloform (Aß42), is considered an early and specific sign of AD, and together with tauopathy, confirms AD diagnosis. To visualize retinal Aß and pTau, state-of-the-art methods use fluorescence. However, administering contrast agents complicates the imaging procedure. To address this problem from fundamentals, ex-vivo studies were performed to develop a label-free hyperspectral imaging method to detect the spectral signatures of Aß42 and pS396-Tau, and predicted their abundance in retinal cross-sections. For the first time, we reported the spectral signature of pTau and demonstrated an accurate prediction of Aß and pTau distribution powered by deep learning. We expect our finding will lay the groundwork for label-free detection of AD.

Signature Effects of Vector-Guided Systemic Nano Bioconjugate Delivery Across Blood-Brain Barrier of Normal, Alzheimer's, and Tumor Mouse Models.

The ability to cross the blood-brain barrier (BBB) is critical for targeted therapy of the central nerve system (CNS). Six peptide vectors were covalently attached to a 50 kDa poly(ß-l-malic acid)-trileucine polymer forming P/LLL(40%)/vector conjugates. The vectors were Angiopep-2 (AP2), B6, Miniap-4 (M4), and d-configurated peptides D1, D3, and ACI-89, with specificity for transcytosis receptors low-density lipoprotein receptor-related protein-1 (LRP-1), transferrin receptor (TfR), bee venom-derived ion channel, and Aß/LRP-1 related transcytosis complex, respectively. The BBB-permeation efficacies were substantially increased ("boosted") in vector conjugates of P/LLL(40%). We have found that the copolymer group binds at the endothelial membrane and, by an allosterically membrane rearrangement, exposes the sites for vector-receptor complex formation. The specificity of vectors is indicated by competition experiments with nonconjugated vectors. P/LLL(40%) does not function as an inhibitor, suggesting that the copolymer binding site is eliminated after binding of the vector-nanoconjugate. The two-step mechanism, binding to endothelial membrane and allosteric exposure of transcytosis receptors, is supposed to be an integral feature of nanoconjugate-transcytosis pathways. In vivo brain delivery signatures of the nanoconjugates were recapitulated in mouse brains of normal, tumor (glioblastoma), and Alzheimer's disease (AD) models. BBB permeation of the tumor was most efficient, followed by normal and then AD-like brain. In tumor-bearing and normal brains, AP2 was the top performing vector; however, in AD models, D3 and D1 peptides were superior ones. The TfR vector B6 was equally efficient in normal and AD-model brains. Cross-permeation efficacies are manifested through modulated vector coligation and dosage escalation such as supra-linear dose dependence and crossover transcytosis activities.

Glatiramer Acetate Immunomodulation: Evidence of Neuroprotection and Cognitive Preservation.

Novel, neuroprotective uses of Copaxone (generic name: glatiramer acetate-GA) are being examined, primarily in neurological conditions involving cognitive decline. GA is a well-studied synthetic copolymer that is FDA-approved for immune-based treatment of relapsing remitting multiple sclerosis (RRMS). Clinical studies have explored the potential mechanism of action (MOA) and outcomes of GA immunization in patients. Furthermore, results from these and animal studies suggest that GA has a direct immunomodulatory effect on adaptive and innate immune cell phenotypes and responses. These MOAs have been postulated to have a common neuroprotective impact in several neuroinflammatory and neurodegenerative diseases. Notably, several clinical studies report that the use of GA mitigated MS-associated cognitive decline. Its propensity to ameliorate neuro-proinflammatory and degenerative processes ignites increased interest in potential alternate uses such as in age-related macular degeneration (AMD), amyotrophic lateral sclerosis (ALS), and Alzheimer's disease (AD). Preclinical studies are exploring less frequent subcutaneous administration of GA, such as once weekly or monthly or a single dosing regimen. Indeed, cognitive functions were found to be either preserved, reversed, or improved after the less frequent treatment regimens with GA in animal models of AD. In this systematic review, we examine the potential novel uses of GA across clinical and pre-clinical studies, with evidence for its beneficial impact on cognition. Future investigation in large-size, double-blind clinical trials is warranted to establish the impact of GA immunomodulation on neuroprotection and cognitive preservation in various neurological conditions.

Visual-stimuli Four-arm Maze test to Assess Cognition and Vision in Mice.

Visual impairments, notably loss of contrast sensitivity and color vision, were documented in Alzheimer's disease (AD) patients yet are critically understudied. This protocol describes a novel visual-stimuli four-arm maze (ViS4M; also called visual x-maze), which is a versatile x-shaped maze equipped with spectrum- and intensity-controlled light-emitting diode (LED) sources and dynamic grayscale objects. The ViS4M is designed to allow the assessment of color and contrast vision along with locomotor and cognitive functions in mice. In the color testing mode, the spectral distributions of the LED lights create four homogenous spaces that differ in chromaticity and luminance, corresponding to the mouse visual system. In the contrast sensitivity test, the four grayscale objects are placed in the middle of each arm, contrasting against the black walls and the white floors of the maze. Upon entering the maze, healthy wild-type (WT) mice tend to spontaneously alternate between arms, even under equiluminant conditions of illumination, suggesting that cognitively and visually intact mice use both color and brightness as cues to navigate the maze. Evaluation of the double-transgenic APPSWE/PS1ΔE9 mouse model of AD (AD+ mice) reveals substantial deficits to alternate in both color and contrast modes at an early age, when hippocampal-based memory and learning is still intact. Profiling of timespan, entries, and transition patterns between the different arms uncovers variable aging and AD-associated impairments in color discrimination and contrast sensitivity. The analysis of arm sequences of alternation reveals different pathways of exploration in young WT, old WT, and AD+ mice, which can be used as color and contrast imprints of functionally intact versus impaired mice. Overall, we describe the utility of a novel visual x-maze test to identify behavioral changes in mice related to cognition, as well as color and contrast vision, with high precision and reproducibility. Graphic abstract: Exploratory behavior of AD+ mice versus age- and sex-matched WT mice is tracked (top left: trajectory from a 5-min video file) in a novel visual-stimuli four-arm maze (ViS4M; also named visual x-maze) equipped with spectrum- and intensity-controlled LED sources or grayscale objects. Consecutive arm entries reveal that APPSWE/PS1ΔE9 (AD+) mice alternate less between arms, as opposed to WT mice. Sequence analysis, according to the three alternation pathways (depicted by white, yellow, and brown arrows) under different conditions of illumination, uncovers specific deficits linked to color vision in AD+ mice, evidenced by a color imprint chart.

Retinal Vasculopathy in Alzheimer's Disease.

The retina has been increasingly investigated as a site of Alzheimer's disease (AD) manifestation for over a decade. Early reports documented degeneration of retinal ganglion cells and their axonal projections. Our group provided the first evidence of the key pathological hallmarks of AD, amyloid ß-protein (Aß) plaques including vascular Aß deposits, in the retina of AD and mild cognitively impaired (MCI) patients. Subsequent studies validated these findings and further identified electroretinography and vision deficits, retinal (p)tau and inflammation, intracellular Aß accumulation, and retinal ganglion cell-subtype degeneration surrounding Aß plaques in these patients. Our data suggest that the brain and retina follow a similar trajectory during AD progression, probably due to their common embryonic origin and anatomical proximity. However, the retina is the only CNS organ feasible for direct, repeated, and non-invasive ophthalmic examination with ultra-high spatial resolution and sensitivity. Neurovascular unit integrity is key to maintaining normal CNS function and cerebral vascular abnormalities are increasingly recognized as early and pivotal factors driving cognitive impairment in AD. Likewise, retinal vascular abnormalities such as changes in vessel density and fractal dimensions, blood flow, foveal avascular zone, curvature tortuosity, and arteriole-to-venule ratio were described in AD patients including early-stage cases. A rapidly growing number of reports have suggested that cerebral and retinal vasculopathy are tightly associated with cognitive deficits in AD patients and animal models. Importantly, we recently identified early and progressive deficiency in retinal vascular platelet-derived growth factor receptor-ß (PDGFRß) expression and pericyte loss that were associated with retinal vascular amyloidosis and cerebral amyloid angiopathy in MCI and AD patients. Other studies utilizing optical coherence tomography (OCT), retinal amyloid-fluorescence imaging and retinal hyperspectral imaging have made significant progress in visualizing and quantifying AD pathology through the retina. With new advances in OCT angiography, OCT leakage, scanning laser microscopy, fluorescein angiography and adaptive optics imaging, future studies focusing on retinal vascular AD pathologies could transform non-invasive pre-clinical AD diagnosis and monitoring.

Color and contrast vision in mouse models of aging and Alzheimer's disease using a novel visual-stimuli four-arm maze.

We introduce a novel visual-stimuli four-arm maze (ViS4M) equipped with spectrally- and intensity-controlled LED emitters and dynamic grayscale objects that relies on innate exploratory behavior to assess color and contrast vision in mice. Its application to detect visual impairments during normal aging and over the course of Alzheimer's disease (AD) is evaluated in wild-type (WT) and transgenic APPSWE/PS1∆E9 murine models of AD (AD+) across an array of irradiance, chromaticity, and contrast conditions. Substantial color and contrast-mode alternation deficits appear in AD+ mice at an age when hippocampal-based memory and learning is still intact. Profiling of timespan, entries and transition patterns between the different arms uncovers variable AD-associated impairments in contrast sensitivity and color discrimination, reminiscent of tritanomalous defects documented in AD patients. Transition deficits are found in aged WT mice in the absence of alternation decline. Overall, ViS4M is a versatile, controlled device to measure color and contrast-related vision in aged and diseased mice.

Retinal capillary degeneration and blood-retinal barrier disruption in murine models of Alzheimer's disease.

Extensive effort has been made studying retinal pathology in Alzheimer's disease (AD) to improve early noninvasive diagnosis and treatment. Particularly relevant are vascular changes, which appear prominent in early brain pathogenesis and could predict cognitive decline. Recently, we identified platelet-derived growth factor receptor beta (PDGFRß) deficiency and pericyte loss associated with vascular Aß deposition in the neurosensory retina of mild cognitively impaired (MCI) and AD patients. However, the pathological mechanisms of retinal vascular changes and their possible relationships with vascular amyloidosis, pericyte loss, and blood-retinal barrier (BRB) integrity remain unknown. Here, we evaluated the retinas of transgenic APPSWE/PS1ΔE9 mouse models of AD (ADtg mice) and wild-type mice at different ages for capillary degeneration, PDGFRß expression, vascular amyloidosis, permeability and inner BRB tight-junction molecules. Using a retinal vascular isolation technique followed by periodic acid-Schiff or immunofluorescent staining, we discovered significant retinal capillary degeneration in ADtg mice compared to age- and sex-matched wild-type mice (P < 0.0001). This small vessel degeneration reached significance in 8-month-old mice (P = 0.0035), with males more susceptible than females. Degeneration of retinal capillaries also progressively increased with age in healthy mice (P = 0.0145); however, the phenomenon was significantly worse during AD-like progression (P = 0.0001). A substantial vascular PDGFRß deficiency (~ 50% reduction, P = 0.0017) along with prominent vascular Aß deposition was further detected in the retina of ADtg mice, which inversely correlated with the extent of degenerated capillaries (Pearson's r = - 0.8, P = 0.0016). Importantly, tight-junction alterations such as claudin-1 downregulation and increased BRB permeability, demonstrated in vivo by retinal fluorescein imaging and ex vivo following injection of FITC-dextran (2000 kD) and Texas Red-dextran (3 kD), were found in ADtg mice. Overall, the identification of age- and Alzheimer's-dependent retinal capillary degeneration and compromised BRB integrity starting at early disease stages in ADtg mice could contribute to the development of novel targets for AD diagnosis and therapy.

Parallels between retinal and brain pathology and response to immunotherapy in old, late-stage Alzheimer's disease mouse models.

Despite growing evidence for the characteristic signs of Alzheimer's disease (AD) in the neurosensory retina, our understanding of retina-brain relationships, especially at advanced disease stages and in response to therapy, is lacking. In transgenic models of AD (APPSWE/PS1∆E9; ADtg mice), glatiramer acetate (GA) immunomodulation alleviates disease progression in pre- and early-symptomatic disease stages. Here, we explored the link between retinal and cerebral AD-related biomarkers, including response to GA immunization, in cohorts of old, late-stage ADtg mice. This aged model is considered more clinically relevant to the age-dependent disease. Levels of synaptotoxic amyloid ß-protein (Aß)1-42, angiopathic Aß1-40, non-amyloidogenic Aß1-38, and Aß42/Aß40 ratios tightly correlated between paired retinas derived from oculus sinister (OS) and oculus dexter (OD) eyes, and between left and right posterior brain hemispheres. We identified lateralization of Aß burden, with one-side dominance within paired retinal and brain tissues. Importantly, OS and OD retinal Aß levels correlated with their cerebral counterparts, with stronger contralateral correlations and following GA immunization. Moreover, immunomodulation in old ADtg mice brought about reductions in cerebral vascular and parenchymal Aß deposits, especially of large, dense-core plaques, and alleviation of microgliosis and astrocytosis. Immunization further enhanced cerebral recruitment of peripheral myeloid cells and synaptic preservation. Mass spectrometry analysis identified new parallels in retino-cerebral AD-related pathology and response to GA immunization, including restoration of homeostatic glutamine synthetase expression. Overall, our results illustrate the viability of immunomodulation-guided CNS repair in old AD model mice, while shedding light onto similar retino-cerebral responses to intervention, providing incentives to explore retinal AD biomarkers.

Alzheimer's Retinopathy: Seeing Disease in the Eyes.

The neurosensory retina emerges as a prominent site of Alzheimer's disease (AD) pathology. As a CNS extension of the brain, the neuro retina is easily accessible for noninvasive, high-resolution imaging. Studies have shown that along with cognitive decline, patients with mild cognitive impairment (MCI) and AD often suffer from visual impairments, abnormal electroretinogram patterns, and circadian rhythm disturbances that can, at least in part, be attributed to retinal damage. Over a decade ago, our group identified the main pathological hallmark of AD, amyloid ß-protein (Aß) plaques, in the retina of patients including early-stage clinical cases. Subsequent histological, biochemical and in vivo retinal imaging studies in animal models and in humans corroborated these findings and further revealed other signs of AD neuropathology in the retina. Among these signs, hyperphosphorylated tau, neuronal degeneration, retinal thinning, vascular abnormalities and gliosis were documented. Further, linear correlations between the severity of retinal and brain Aß concentrations and plaque pathology were described. More recently, extensive retinal pericyte loss along with vascular platelet-derived growth factor receptor-ß deficiency were discovered in postmortem retinas of MCI and AD patients. This progressive loss was closely associated with increased retinal vascular amyloidosis and predicted cerebral amyloid angiopathy scores. These studies brought excitement to the field of retinal exploration in AD. Indeed, many questions still remain open, such as queries related to the temporal progression of AD-related pathology in the retina compared to the brain, the relations between retinal and cerebral changes and whether retinal signs can predict cognitive decline. The extent to which AD affects the retina, including the susceptibility of certain topographical regions and cell types, is currently under intense investigation. Advances in retinal amyloid imaging, hyperspectral imaging, optical coherence tomography, and OCT-angiography encourage the use of such modalities to achieve more accurate, patient- and user-friendly, noninvasive detection and monitoring of AD. In this review, we summarize the current status in the field while addressing the many unknowns regarding Alzheimer's retinopathy.

Effects of IL-34 on Macrophage Immunological Profile in Response to Alzheimer's-Related Aß42 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 ß-protein (Aß). Aß42 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 Aß42 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 Aß42 was markedly reduced following exposure to IL-34 compared to M-CSF. Surface expression of type B scavenger receptor CD36, known to facilitate Aß recognition and uptake, was modified following treatment with IL-34. Similarly, IL-34 macrophages expressed lower levels of proteins involved in both Aß uptake (triggering receptor expressed on myeloid cells 2, TREM2) as well as Aß-degradation (matrix metallopeptidase 9, MMP-9). Interestingly, intracellular compartmentalization of Aß 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 Aß42 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 Aß. Given the critical role of macrophage-mediated Aß 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 Aß clearance and prevent disease development.

Activated Bone Marrow-Derived Macrophages Eradicate Alzheimer's-Related Aß42 Oligomers and Protect Synapses.

Impaired synaptic integrity and function due to accumulation of amyloid ß-protein (Aß42) oligomers is thought to be a major contributor to cognitive decline in Alzheimer's disease (AD). However, the exact role of Aß42 oligomers in synaptotoxicity and the ability of peripheral innate immune cells to rescue synapses remain poorly understood due to the metastable nature of oligomers. Here, we utilized photo-induced cross-linking to stabilize pure oligomers and study their effects vs. fibrils on synapses and protection by Aß-phagocytic macrophages. We found that cortical neurons were more susceptible to Aß42 oligomers than fibrils, triggering additional neuritic arborization retraction, functional alterations (hyperactivity and spike waveform), and loss of VGluT1- and PSD95-excitatory synapses. Co-culturing neurons with bone marrow-derived macrophages protected synapses against Aß42 fibrils; moreover, immune activation with glatiramer acetate (GA) conferred further protection against oligomers. Mechanisms involved increased Aß42 removal by macrophages, amplified by GA stimulation: fibrils were largely cleared through intracellular CD36/EEA1+-early endosomal proteolysis, while oligomers were primarily removed via extracellular/MMP-9 enzymatic degradation. In vivo studies in GA-immunized or CD115+-monocyte-grafted APPSWE/PS1ΔE9-transgenic mice followed by pre- and postsynaptic analyses of entorhinal cortex and hippocampal substructures corroborated our in vitro findings of macrophage-mediated synaptic preservation. Together, our data demonstrate that activated macrophages effectively clear Aß42 oligomers and rescue VGluT1/PSD95 synapses, providing rationale for harnessing macrophages to treat AD.