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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 β-protein (Aβ42) forms and novel intraneuronal Aβ oligomers (AβOi), 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 Aβ uptake was reduced. Female AD patients exhibited higher levels of retinal microgliosis than males. Notably, retinal Aβ42, S100 calcium-binding protein B+ macrogliosis, and atrophy correlated with severity of brain Aβ pathology, tauopathy, and atrophy, and most retinal pathologies reflected Braak staging. All retinal biomarkers correlated with the cognitive scores, with retinal Aβ42, far-peripheral AβOi 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.

SummaryDuring vertebrate embryogenesis, axial tendons develop from the paraxial mesoderm and differentiate through specific developmental stages to reach the syndetome stage. While the main roles of signaling pathways in the earlier stages of the differentiation have been well established, pathway nuances in syndetome specification from the sclerotome stage have yet to be explored. Here, we show stepwise differentiation of human iPSCs to the syndetome stage using chemically defined media and small molecules that were modified based on single cell RNA-sequencing and pathway analysis. We identified a significant population of branching off-target cells differentiating towards a neural phenotype overexpressing Wnt. Further transcriptomics post-addition of a WNT inhibitor at the somite stage and onwards revealed not only total removal of the neural off-target cells, but also increased syndetome induction efficiency. Fine-tuning tendon differentiationin vitrois essential to address the current challenges in developing a successful cell-based tendon therapy.

Low back pain (LBP) is often associated with the degeneration of human intervertebral discs (IVDs). However, the pain-inducing mechanism in degenerating discs remains to be elucidated. Here, we identified a subtype of locally residing nucleus pulposus cells (NPCs), generated by the environmental stress in degenerating discs, that triggered the onset of discogenic LBP. Single-cell transcriptomic analysis of human tissues showed a strong correlation between this specific pain-triggering subtype and the pain conditions in human degenerated discs. Next, we recreated this pain-triggering subtype by applying known exogenous stressors to healthy NPCsin vitro. The recreated pain phenotype activated functional sensory neurons responsein vitroand induced local inflammatory responses, hyperalgesia, and mechanical sensitivity in a healthy rat IVDin vivo. Our findings provide strong evidence of a previously unknown pain-inducing mechanism mediated by NPCs in degenerating IVDs. This newly defined pathway will aid in the development of NPC-targeted therapeutic strategies for clinically unmet need to attenuate discogenic LBP.One Sentence SummaryDiscogenic low back pain can be initiated by a stress-induced subtype of nucleus pulposus cells present in human degenerating intervertebral discs

Alzheimer’s disease (AD) pathologies were discovered in the easily accessible neurosensory retina. Yet, their specific nature, topographical distribution, and relationship with disease status remain undefined. Here, we histologically determined burden and spatial distribution of amyloid β-protein (Aβ42), intraneuronal scFvA13+-Aβ species, macro- and microgliosis, and atrophy in superior- and inferior-temporal retinas of human donors with mild cognitive impairment (MCI) or AD versus normal cognition. AD and MCI patients had enhanced retinopathy, predominantly affecting inner layers and peripheral subregions, which quantitatively correlated with severity of cerebral amyloid, tau, and neurodegeneration, and cognitive scores. In advanced clinical stages AD retinopathy further affected central outer segments. Increased retinal macrogliosis and Aβ-phagocytosing microglia were detected in MCI and AD patients. Further, distinct proteome profiles of AD retinas were identified, displaying greater overlap with the temporal cortices than with hippocampi or cerebella. AD retinas exhibited upregulated inflammatory and neurodegenerative processes and downregulated oxidative-phosphorylation/mitochondrial, and photoreceptor-related pathways. This study identifies and maps AD retinopathy, demonstrating the quantitative relationship with brain pathology and cognition.

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, phagocytic, pro-healing phenotype, whereas OPN inhibition triggered a pro-inflammatory phenotype. Here, we applied a global proteome profiling via mass spectrometry analysis to gain a mechanistic understanding of OPN suppression versus induction in macrophages. We identified over 630 differentially expressed proteins (DEPs) in OPN knockout (OPNKO) or GA-stimulated versus wild type (WT) macrophages. Two topmost downregulated DEPs in OPN-deficient macrophages were ubiquitin C-terminal hydrolase L1 (UCHL1), a crucial component of ubiquitin-proteasome system (UPS), and the anti-inflammatory Heme oxygenase 1 (HMOX-1), whereas GA stimulation upregulated their expression. We confirmed UCHL1 expression in BMMΦ, which was previously recognized as a neuronal-specific protein. Further, immunoprecipitation assays suggest a direct interaction of OPN and UCHL1 proteins. Functional pathway analyses revealed two inversely regulated pathways in OPN-deficient macrophages: activated oxidative stress and lysosome-mitochondria-mediated apoptosis (ROS, Lamp1/2, ATP-synthase subunits, cathepsins, and cytochrome C and B subunits) and inhibited translation and proteolytic pathways (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 (proteostasis) in macrophages—inducing apoptosis and inhibiting translation—whereas GA-stimulated OPN induction restores cellular proteostasis. Taken together, OPN is essential for macrophage homeostasis via regulation of cell viability, UCHL1-UPS, and protein synthesis, indicating its potential application in immunotherapy.

Pericyte loss and deficient vascular platelet-derived growth factor receptor-β (PDGFRβ) signaling are prominent features of the blood–brain barrier breakdown described in Alzheimer’s disease (AD) that can predict cognitive decline yet have never been studied in the retina. Recent reports using noninvasive retinal amyloid imaging, optical coherence tomography angiography, and histological examinations support the existence of vascular-structural abnormalities and vascular amyloid β-protein (Aβ) deposits in retinas of AD patients. However, the cellular and molecular mechanisms of such retinal vascular pathology were not previously explored. Here, by modifying a method of enzymatically clearing non-vascular retinal tissue and fluorescent immunolabeling of the isolated blood vessel network, we identified substantial pericyte loss together with significant Aβ deposition in retinal microvasculature and pericytes in AD. Evaluation of postmortem retinas from a cohort of 56 human donors revealed an early and progressive decrease in vascular PDGFRβ in mild cognitive impairment (MCI) and AD compared to cognitively normal controls. Retinal PDGFRβ loss significantly associated with increased retinal vascular Aβ40 and Aβ42 burden. Decreased vascular LRP-1 and early apoptosis of pericytes in AD retina were also detected. Mapping of PDGFRβ and Aβ40 levels in pre-defined retinal subregions indicated that certain geometrical and cellular layers are more susceptible to AD pathology. Further, correlations were identified between retinal vascular abnormalities and cerebral Aβ burden, cerebral amyloid angiopathy (CAA), and clinical status. Overall, the identification of pericyte and PDGFRβ loss accompanying increased vascular amyloidosis in Alzheimer’s retina implies compromised blood–retinal barrier integrity and provides new targets for AD diagnosis and therapy. Electronic supplementary material The online version of this article (10.1007/s00401-020-02134-w) contains supplementary material, which is available to authorized users.

INTRODUCTION: Retinal imaging is a non-invasive tool to study retinal vasculature and neurodegeneration. Patients with mild cognitive impairment (MCI) and Alzheimer’s disease (AD)-related cognitive disorder exhibit both retinal vascular abnormalities and intraretinal accumulation of amyloid beta-protein (Aβ) plaque. Curcumin-enhanced retinal fluorescence imaging (CRFI) was recently translated as a safe imaging tool for retinal Aβ plaque quantification, holding promise as an early-stage pathological biomarker of AD. In this exploratory study, we sought to determine whether retinal vascular features combined with retinal amyloid burden correlate with the neurocognitive status.METHODS: We used scanning laser ophthalmoscopy to assess quantitative CRFI in a cohort of patients with cognitive impairment that underwent standard neuropsychological testing. Retinal blood vessels were segmented in a predefined circumpapillary region of interest. For each centerline, vessel tortuosity index, vessel inflection index and branching angle was quantified. Additionally, we automatically quantified retinal amyloid count in the supero-temporal quadrant and its subregions: posterior pole, proximal mid-periphery, and distal mid-periphery. Investigators performing quantifications were blinded to the subjects’ clinical characteristics. Linear regression models were used to assess the correlations between retinal vascular and amyloid parameters and cognitive domain Z-scores.RESULTS: In this pilot study, 34 subjects underwent automated retinal amyloid imaging and 29 subjects (55% female, mean age 64±6 years) had the combined retinal amyloid and peripapillary vascular analysis. Eleven subjects had normal cognition, 15 MCI, 2 probable AD and 1 non-AD dementia. CRFI was increased in individuals with impaired versus normal cognitive function (p 0.0012). Venous VTI was the most significant vascular parameter that differ across levels of CDR. Branching angle correlated with amyloid count in the distal mid-periphery (p=0.03), whereas vessel inflexion index correlated with posterior pole amyloid count (p=0.02). The combined proximal mid-periphery amyloid count – venous tortuosity index was found to exhibit highly significant group differences between cognitively impaired and cognitively normal subjects (0.49 ± 1.1 vs 0.91 ± 1.4, p=0.006). The combined proximal mid-periphery amyloid-venous tortuosity index also correlated with verbal memory (Wechsler Memory Scale-IV; p=0.001) and cognitive-related quality of life (SF-36 mental component score Z-scores; p=0.039).CONCLUSION: Retinal venular tortuosity discriminates across cognitive scores and in combination with proximal mid-periphery amyloid count predicts verbal memory and cognitive-related quality-of-life. Future research is needed to confirm the clinical utility of this integrated retinal imaging-based methodology.

Introduction: Retinal vascular features predict cognitive performance and retinal imaging is a non-invasive tool to study brain health. Patients with Alzheimer’s disease (AD)-related cognitive disorders exhibit both retinal vascular abnormalities and intraretinal accumulation of amyloid-beta (Aβ) plaque. Curcumin-enhanced retinal fluorescence imaging (CRFI) was recently translated as a safe imaging tool for retinal Aβ plaque quantification. In this study, we sought to determine whether and which retinal arterial and venous parameters are predictive of retinal Aβ burden and cognitive Z-scores. Methods: We used scanning laser ophthalmoscopy with RetiaTM (Centervue, Inc) to obtain CRFI in a cohort of patients with cognitive decline. Retinal blood vessels were segmented in a predefined circumpapillary region of interest. For each centerline, vessel tortuosity index (VTI), vessel inflection index (VII), and branching angle (VBA) were quantified, independently for arteries and veins. Additionally, we automatically quantified retinal amyloid count in the supero-temporal quadrant and its subregions: posterior pole (PP), proximal mid-periphery (PMP), and distal mid-periphery (DMP). All quantifications were done blinded to the subjects’ clinical characteristics. Linear regression models were used to assess the correlations between retinal vascular and amyloid parameters. Results: 29 subjects (55% female, mean age 64±6 years) were included in the analysis: 11 had normal cognition, 16 mild cognitive impairment and 2 probable AD. Venous VTI was the only vascular parameter significantly different across levels of Clinical Dementia Rate scores and between cognitively normal and impaired. Venous VBA was a significant predictor of DMP amyloid count (p=0.03). Arterial VII significantly predicted PP amyloid count (p=0.02). VBA predicted Wechsler Adult Intelligence Scale Z-score (p=0.08). The combined VTI-PMP amyloid index significantly predicted Wechsler Memory Scale, California Verbal Learning Test and SF-36 mental component score Z-scores (p Conclusion: Retinal venous tortuosity discriminates across cognitive scores and in combination with PMP amyloid count may predict verbal memory and related quality-of-life scores.

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 (PDGFRβ) deficiency and pericyte loss associated with vascular Aβ 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, PDGFRβ 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 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 PDGFRβ deficiency (~ 50% reduction, P = 0.0017) along with prominent vascular Aβ 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.

Introduction Despite advances in imaging retinal amyloidosis, a quantitative and topographical investigation of retinal amyloid beta burden in patients with cognitive decline has never been reported. Methods We used the specific amyloid‐binding fluorophore curcumin and laser ophthalmoscopy to assess retinal amyloid imaging (RAI) in 34 patients with cognitive decline. We automatically quantified retinal amyloid count (RAC) and area in the superotemporal retinal sub‐regions and performed correlation analyses with cognitive and brain volumetric parameters. Results RAC significantly and inversely correlated with hippocampal volume (HV; r = ‐0.39, P = .04). The proximal mid‐periphery (PMP) RAC and RA areas were significantly greater in patients with Montreal Cognitive Assessment (MOCA) score < 26 (P = .01; Cohen d = 0.83 and 0.81, respectively). PMP showed significantly more RAC and area in subjects with amnestic mild cognitive impairment (MCI) and Alzheimer's disease (AD) compared to cognitively normal (P = .04; Cohen d = 0.83). Conclusion Quantitative RAI is a feasible technique and PMP RAC may predict HV. Future larger studies should determine RAI's potential as a biomarker of early AD.

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 β‐protein (Aβ)1–42, angiopathic Aβ1–40, non‐amyloidogenic Aβ1–38, and Aβ42/Aβ40 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 Aβ burden, with one‐side dominance within paired retinal and brain tissues. Importantly, OS and OD retinal Aβ 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 Aβ 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., In this study, Doustar et al. revealed that retinal Abeta burden predicts its brain levels in old, late‐stage murine models of Alzheimer's disease and further in response to immunotherapy. Substantial therapeutic effects are detected even at such advanced disease stage; immunomodulation effectively mitigates vascular and parenchymal amyloid‐beta deposition, diminishes neuroinflammation, as well as restores synaptic density and retino‐cerebral glutamine synthetase levels.

Targeted overexpression of angiotensin-converting enzyme (ACE), an amyloid-β protein degrading enzyme, to brain resident microglia and peripheral myelomonocytes (ACE10 model) substantially diminished Alzheimer’s-like disease in double-transgenic APPSWE/PS1ΔE9 (AD+) mice. In this study, we explored the impact of selective and transient angiotensin-converting enzyme overexpression on macrophage behaviour and the relative contribution of bone marrow-derived ACE10 macrophages, but not microglia, in attenuating disease progression. To this end, two in vivo approaches were applied in AD+ mice: (i) ACE10/GFP+ bone marrow transplantation with head shielding; and (ii) adoptive transfer of CD115+-ACE10/GFP+ monocytes to the peripheral blood. Extensive in vitro studies were further undertaken to establish the unique ACE10-macrophage phenotype(s) in response to amyloid-β1-42 fibrils and oligomers. The combined in vivo approaches showed that increased cerebral infiltration of ACE10 as compared to wild-type monocytes (∼3-fold increase; P < 0.05) led to reductions in cerebral soluble amyloid-β1-42, vascular and parenchymal amyloid-β deposits, and astrocytosis (31%, 47–80%, and 33%, respectively; P < 0.05–0.0001). ACE10 macrophages surrounded brain and retinal amyloid-β plaques and expressed 3.2-fold higher insulin-like growth factor-1 (P < 0.01) and ∼60% lower tumour necrosis factor-α (P < 0.05). Importantly, blood enrichment with CD115+-ACE10 monocytes in symptomatic AD+ mice resulted in pronounced synaptic and cognitive preservation (P < 0.05–0.001). In vitro analysis of macrophage response to well-defined amyloid-β1-42 conformers (fibrils, prion rod-like structures, and stabilized soluble oligomers) revealed extensive resistance to amyloid-β1-42 species by ACE10 macrophages. They exhibited 2–5-fold increased surface binding to amyloid-β conformers as well as substantially more effective amyloid-β1-42 uptake, at least 8-fold higher than those of wild-type macrophages (P < 0.0001), which were associated with enhanced expression of surface scavenger receptors (i.e. CD36, scavenger receptor class A member 1, triggering receptor expressed on myeloid cells 2, CD163; P < 0.05–0.0001), endosomal processing (P < 0.05–0.0001), and ∼80% increased extracellular degradation of amyloid-β1-42 (P < 0.001). Beneficial ACE10 phenotype was reversed by the angiotensin-converting enzyme inhibitor (lisinopril) and thus was dependent on angiotensin-converting enzyme catalytic activity. Further, ACE10 macrophages presented distinct anti-inflammatory (low inducible nitric oxide synthase and lower tumour necrosis factor-α), pro-healing immune profiles (high insulin-like growth factor-1, elongated cell morphology), even following exposure to Alzheimer’s-related amyloid-β1-42 oligomers. Overall, we provide the first evidence for therapeutic roles of angiotensin-converting enzyme-overexpressing macrophages in preserving synapses and cognition, attenuating neuropathology and neuroinflammation, and enhancing resistance to defined pathognomonic amyloid-β forms.

Osteopontin (OPN), a matricellular immunomodulatory cytokine highly expressed by myelomonocytic cells, is known to regulate immune cell migration, communication, and response to brain injury. Enhanced cerebral recruitment of monocytes achieved through glatiramer acetate (GA) immunization or peripheral blood enrichment with bone marrow (BM)-derived CD115+ monocytes (MoBM) curbs amyloid β-protein (Aβ) neuropathology and preserves cognitive function in murine models of Alzheimer's disease (ADtg mice). To elucidate the beneficial mechanisms of these immunomodulatory approaches in AD, we focused on the potential role of OPN in macrophage-mediated Aβ clearance. Here, we found extensive OPN upregulation along with reduction of vascular and parenchymal Aβ burden in cortices and hippocampi of GA-immunized ADtg mice. Treatment combining GA with blood-grafted MoBM further increased OPN levels surrounding residual Aβ plaques. In brains from AD patients and ADtg mice, OPN was also elevated and predominantly expressed by infiltrating GFP+- or Iba1+-CD45high monocyte-derived macrophages engulfing Aβ plaques. Following GA immunization, we detected a significant increase in a subpopulation of inflammatory blood monocytes (CD115+CD11b+Ly6Chigh) expressing OPN, and subsequently, an elevated population of OPN-expressing CD11b+Ly6C+CD45high monocyte/macrophages in the brains of these ADtg mice. Correlogram analyses indicate a strong linear correlation between cerebral OPN levels and macrophage infiltration, as well as a tight inverse relation between OPN and Aβ-plaque burden. In vitro studies corroborate in vivo findings by showing that GA directly upregulates OPN expression in BM-derived macrophages (MФBM). Further, OPN promotes a phenotypic shift that is highly phagocytic (increased uptake of Aβ fibrils and surface scavenger receptors) and anti-inflammatory (altered cell morphology, reduced iNOS, and elevated IL-10 and Aβ-degrading enzyme MMP-9). Inhibition of OPN expression in MФBM, either by siRNA, knockout (KOOPN), or minocycline, impairs uptake of Aβ fibrils and hinders GA's neuroprotective effects on macrophage immunological profile. Addition of human recombinant OPN reverses the impaired Aβ phagocytosis in KOOPN-MФBM. This study demonstrates that OPN has an essential role in modulating macrophage immunological profile and their ability to resist pathogenic forms of Aβ.

Introduction: Retinal imaging is a non-invasive tool to study both retinal vasculature and neurodegeneration. In this exploratory retinal curcumin-fluorescence imaging (RFI) study, we sought to determine whether retinal vascular features combined with retinal amyloid burden correlate with the neurocognitive status. Methods: We used quantitative RFI in a cohort of patients with cognitive impairment to automatically compute retinal amyloid burden. Retinal blood vessels were segmented, and the vessel tortuosity index (VTI), inflection index, and branching angle were quantified. We assessed the correlations between retinal vascular and amyloid parameters, and cognitive domain Z-scores using linear regression models. Results: Thirty-four subjects were enrolled and twenty-nine (55% female, mean age 64 ± 6 years) were included in the combined retinal amyloid and vascular analysis. Eleven subjects had normal cognition and 18 had impaired cognition. Retinal VTI was discriminated among cognitive scores. The combined proximal mid-periphery amyloid count and venous VTI index exhibited significant differences between cognitively impaired and cognitively normal subjects (0.49 ± 1.1 vs. 0.91 ± 1.4, p = 0.006), and correlated with both the Wechsler Memory Scale-IV and SF-36 mental component score Z-scores (p &lt, 0.05). Conclusion: This pilot study showed that retinal venular VTI combined with the proximal mid-periphery amyloid count could predict verbal memory loss. Future research is needed to finesse the clinical application of this retinal imaging-based technology.

Background Mechanical ventilation is strongly associated with cognitive decline after critical illness. This finding is particularly evident among older individuals who have pre-existing cognitive impairment, most commonly characterized by varying degrees of cerebral amyloid-β accumulation, neuroinflammation, and blood-brain barrier dysfunction. We sought to test the hypothesis that short-term mechanical ventilation contributes to the neuropathology of cognitive impairment by (i) increasing cerebral amyloid-β accumulation in mice with pre-existing Alzheimer’s disease pathology, (ii) increasing neurologic and systemic inflammation in wild-type mice and mice with pre-existing Alzheimer’s disease pathology, and (iii) increasing hippocampal blood-brain barrier permeability in wild-type mice and mice with pre-existing Alzheimer’s disease pathology. Methods We subjected double transgenic Alzheimer’s disease (APP/PSEN1) and wild-type mice to mechanical ventilation for 4 h and compared to non-mechanically ventilated Alzheimer’s disease model and wild-type mice. Cerebral soluble/insoluble amyloid-β1–40/amyloid-β1–42 and neurological and systemic markers of inflammation were quantified. Hippocampal blood-brain barrier permeability was quantified using a novel methodology that enabled assessment of small and large molecule permeability across the blood-brain barrier. Results Mechanical ventilation resulted in (i) a significant increase in cerebral soluble amyloid-β1–40 (p = 0.007) and (ii) significant increases in neuroinflammatory cytokines in both wild-type and Alzheimer’s disease mice which, in most cases, were not reflected in the plasma. There were (i) direct correlations between polymorphonuclear cells in the bronchoalveolar fluid and cerebral soluble amyloid-β1–40 (p = 0.0033), and several Alzheimer’s disease-relevant neuroinflammatory biomarkers including cerebral TNF-α and IL-6; (iii) significant decreases in blood-brain barrier permeability in mechanically ventilated Alzheimer’s disease mice and a trend towards increased blood-brain barrier permeability in mechanically ventilated wild-type mice. Conclusions These results provide the first evidence that short-term mechanical ventilation independently promotes the neuropathology of Alzheimer’s disease in subjects with and without pre-existing cerebral Alzheimer’s disease pathology. Future studies are needed to further clarify the specific mechanisms by which this occurs and to develop neuroprotective mechanical ventilation strategies that mitigate the risk of cognitive decline after critical illness. Electronic supplementary material The online version of this article (10.1186/s13054-019-2356-2) contains supplementary material, which is available to authorized users.

Cognitive decline in patients with Alzheimer's disease (AD) is associated with elevated brain levels of amyloid β protein (Aβ), particularly neurotoxic Aβ(1-42). Angiotensin-converting enzyme (ACE) can degrade Aβ(1-42), and ACE overexpression in myelomonocytic cells enhances their immune function. To examine the effect of targeted ACE overexpression on AD, we crossed ACE(10/10) mice, which overexpress ACE in myelomonocytes using the c-fms promoter, with the transgenic APP(SWE)/PS1(ΔE9) mouse model of AD (AD⁺). Evaluation of brain tissue from these AD⁺ACE(10/10) mice at 7 and 13 months revealed that levels of both soluble and insoluble brain Aβ(1-42) were reduced compared with those in AD⁺ mice. Furthermore, both plaque burden and astrogliosis were drastically reduced. Administration of the ACE inhibitor ramipril increased Aβ levels in AD⁺ACE(10/10) mice compared with the levels induced by the ACE-independent vasodilator hydralazine. Overall, AD⁺ACE(10/10) mice had less brain-infiltrating cells, consistent with reduced AD-associated pathology, though ACE-overexpressing macrophages were abundant around and engulfing Aβ plaques. At 11 and 12 months of age, the AD⁺ACE(10/WT) and AD⁺ACE(10/10) mice were virtually equivalent to non-AD mice in cognitive ability, as assessed by maze-based behavioral tests. Our data demonstrate that an enhanced immune response, coupled with increased myelomonocytic expression of catalytically active ACE, prevents cognitive decline in a murine model of AD.

Weekly glatiramer acetate immunization of transgenic mice modelling Alzheimer's disease resulted in retained cognition (Morris water maze test), decreased amyloid-β plaque burden, and regulation of local inflammation through a mechanism involving enhanced recruitment of monocytes. Ablation of bone marrow-derived myeloid cells exacerbated plaque pathology, whereas weekly administration of glatiramer acetate enhanced cerebral recruitment of innate immune cells, which dampened the pathology. Here, we assessed the therapeutic potential of grafted CD115(+) monocytes, injected once monthly into the peripheral blood of transgenic APPSWE/PS1ΔE9 Alzheimer's disease mouse models, with and without weekly immunization of glatiramer acetate, as compared to glatiramer acetate alone. All immune-modulation treatment groups were compared with age-matched phosphate-buffered saline-injected control transgenic and untreated non-transgenic mouse groups. Two independent cohorts of mice were assessed for behavioural performance (6-8 mice/group); treatments started in 10-month-old symptomatic mice and spanned a total of 2 months. For all three treatments, our data suggest a substantial decrease in cognitive deficit as assessed by the Barnes maze test (P < 0.0001-0.001). Improved cognitive function was associated with synaptic preservation and reduction in cerebral amyloid-β protein levels and astrogliosis (P < 0.001 and P < 0.0001), with no apparent additive effects for the combined treatment. The peripherally grafted, green fluorescent protein-labelled and endogenous monocytes, homed to cerebral amyloid plaques and directly engulfed amyloid-β; their recruitment was further enhanced by glatiramer acetate. In glatiramer acetate-immunized mice and, moreover, in the combined treatment group, monocyte recruitment to the brain was coupled with greater elevation of the regulatory cytokine IL10 surrounding amyloid-β plaques. All treated transgenic mice had increased cerebral levels of MMP9 protein (P < 0.05), an enzyme capable of degrading amyloid-β, which was highly expressed by the infiltrating monocytes. In vitro studies using primary cultures of bone marrow monocyte-derived macrophages, demonstrated that glatiramer acetate enhanced the ability of macrophages to phagocytose preformed fibrillar amyloid-β1-42 (P < 0.0001). These glatiramer acetate-treated macrophages exhibited increased expression of the scavenger receptors CD36 and SCARA1 (encoded by MSR1), which can facilitate amyloid-β phagocytosis, and the amyloid-β-degrading enzyme MMP9 (P < 0.0001-0.001). Overall, our studies indicate that increased cerebral infiltration of monocytes, either by enrichment of their levels in the circulation or by weekly immunization with glatiramer acetate, resulted in substantial attenuation of disease progression in murine Alzheimer's models by mechanisms that involved enhanced cellular uptake and enzymatic degradation of toxic amyloid-β as well as regulation of brain inflammation.

The cognitive decline in Alzheimer's disease (AD) is associated with elevated brain levels of amyloid β-protein (Aβ), particularly Aβ 1-42 . Angiotensin-converting enzyme (ACE) can enzymatically degrade Aβ 1-42 and its overexpression in myelomonocytic cells enhances their immune function. To examine the effect of targeted ACE overexpression on AD, we crossed ACE 10/10 mice, which over express ACE in myelomonocytic cells, with the double-transgenic APP SWE /PS1 ΔE9 mouse model of AD (AD + ). At 7 months, soluble Aβ 1-42 was reduced by 44% (136 vs 245 pg/mg) and Aβ 1-40 was reduced by 32% (21 vs 31 pg/mg). Plaque burden was reduced by as much as 79% (13 vs 62 10 3 mm 2 ) and insoluble Aβ 1-42 by 64% (189 vs 521 pg/mg). There was also a substantial reduction in astrogliosis (49-57% at 7 months, 50% at 13 months). AD + ACE 10/10 mice demonstrated less overall brain infiltrating cells, consistent with less AD pathology, though ACE-overexpressing monocytes and macrophages were increasingly abundant surrounding and engulfing Aβ plaque. At 11 and 12 months, AD + ACE 10/WT and AD + ACE 10/10 mice were virtually equivalent to non-AD mice in cognitive ability as assessed by maze-based behavioral tests. This study shows that an enhanced immune response, coupled with increased myelomonocytic cell expression of ACE, resulted in essentially complete prevention of the cognitive decline observed in a murine model of AD.

Background: Previous studies from our group and others have demonstrated that peripheral immune cells and especially infiltrating bloodborne monocytes can home to pathological beta-amyloid protein aggregates (Ab plaques) and facilitate their clearance by phagocytosis and secretion of Ab-degrading enzymes. One of the enzymes that was shown capable of degrading Ab and convert the deleterious Ab (1-42) to shorter nontoxic isoforms is angiotensin-converting enzyme (ACE). Results: We found a marked reduction in Ab plaque number and size in the cortex and hippocampus of 5, 7 and 12 month-old AD mice; above 40% decrease in mice having one ACE10 allele and above 80% in mice having two ACE10 alleles. Moreover, the levels of soluble neurotoxic Ab (1-42) isoforms were substantially decreased in brain and plasma of ACE10xADtg mice. Importantly, ADtg mice carrying either one or two ACE10 alleles had a considerable improvement in learning and memory functions as assessed by a panel of behavioral tests such as the Barnes maze, and were indistinguishable than the wild type non-AD mice. These effects were accompanied by an extensive reduction in cortical astrogliosis as assessed by number and area of reactive GFAP-positive astrocytes. Flow cytometry analysis of immune cell profiles in the brain revealed a significant increase in migration of ACE10-overexpessing monocytes as compared to wt monocytes in ADtg mice. Conclusions: These studies indicate that selective ACE-overexpression in microglia and monocytes leads to a remarkable attenuation of AD-like progression in mouse models. Therefore, targeted overexpression of ACE on monocytes may present a highly promising and novel disease-modifying approach for AD treatment.