Key role of MIF-related neuroinflammation in neurodegeneration and cognitive impairment in Alzheimer's disease.

BACKGROUND: Macrophage Migration Inhibitory Factor (MIF) is a potent proinflammatory cytokine that promotes the production of other immune mediators. MIF is produced by most cell types in the brain including microglia, astrocytes and neurons. Enhanced expression of MIF might contribute to the persistent activation of glial, chronic neuroinflammation and neurodegeneration. Here, we investigated the effect of MIF on inflammatory markers and spatial learning in a mouse model of sporadic AD and on tau pathology in AD patients. METHODS: We examined the effects of MIF deficiency and pharmacological MIF inhibition in vitro and in vivo. In vitro, quantitative PCR and ELISA were used to assess cytokine production of STZ-treated glial cells. In vivo, C57BL/6 mice were subjected to intracerebroventricular streptozotocin injection (3 mg/kg, ICV-STZ). Neuroinflammation and contextual learning performance were assessed using quantitative PCR and fear conditioning, respectively. Pharmacological MIF inhibition was achieved with intraperitoneal injections of ISO-1 (daily, IP, 20 mg/kg in 5% DMSO in 0.9% NaCl) for 4 weeks following ICV-STZ injection. The findings from ISO-1 treated mice were confirmed in MIF knockout C57BL/6. To assess the role of MIF in human AD, cerebrospinal fluid levels of MIF and hyperphosphorylated tau were measured using ELISA. RESULTS: Administration ICV-STZ resulted in hippocampal dependent cognitive impairment. MIF inhibition with ISO-1 significantly improved the STZ-induced impairment in contextual memory performance, indicating MIF-related inflammation as a major contributor to ICV-STZ-induced memory deficits. Furthermore, inhibition of the MIF resulted in reduced cytokine production in vitro and in vivo. In human subjects with AD at early clinical stages, cerebrospinal fluid levels of MIF were increased in comparison with age-matched controls, and correlated with biomarkers of tau hyper-phosphorylation and neuronal injury hinting at MIF levels as a potential biomarker for early-stage AD. CONCLUSIONS: The present study indicates the key role of MIF in controlling the chronic cytokine release in neuroinflammation related to tau hyperphosphorylation, neurodegeneration, and clinical manifestations of AD, suggesting the potential of MIF inhibition as therapeutic strategy to slow down neurodegeneration and clinical disease progression.

Proteomes of Paired Human Cerebrospinal Fluid and Plasma: Relation to Blood-Brain Barrier Permeability in Older Adults.

The systems-level relationship between the proteomes of cerebrospinal fluid (CSF) and plasma has not been comprehensively described so far. Recently developed shotgun proteomic workflows allow for deeper characterization of the proteomes from body fluids in much larger sample size. We deployed state-of-the-art mass spectrometry-based proteomics in paired CSF and plasma samples volunteered by 120 elders with and without cognitive impairment to comprehensively characterize and examine compartmental proteome differences and relationships between both body fluids. We further assessed the influence of blood-brain barrier (BBB) integrity and tested the hypothesis that BBB breakdown can be identified from CSF and plasma proteome alterations in nondemented elders. We quantified 790 proteins in CSF and 422 proteins in plasma, and 255 of the proteins were identified in both compartments. Pearson's statistics determined 28 proteins with associated levels between CSF and plasma. BBB integrity as defined with the CSF/serum albumin index influenced 76 CSF/plasma protein ratios. In least absolute shrinkage and selection operator models, CSF and plasma proteins improved identification of BBB impairment. In conclusion, we provide here a first comprehensive draft map of interacting human CSF and plasma proteomes, in view of their complex and dynamic compositions, and influence of the BBB.

Blood-brain barrier breakdown, neuroinflammation, and cognitive decline in older adults.

INTRODUCTION: Blood-brain barrier (BBB) breakdown is observed in older versus younger adults and in late-onset Alzheimer's disease versus age-matched controls, but its causes and consequences in aging are unclear. We tested the hypothesis that BBB breakdown is associated with cognitive decline and inflammation in nondemented elders. METHODS: Cerebrospinal fluid and serum inflammatory markers were measured using sandwich immunoassays in 120 subjects. Least Absolute Shrinkage and Selection Operator-logistic regression selected cerebrospinal fluid and serum signatures that best classified BBB impairment defined by the cerebrospinal fluid albumin index ≥9. Linear regression examined changes in Clinical Dementia Rating sum of boxes as a function of BBB integrity at baseline. RESULTS: Mean age was 70 years, mean Mini­Mental State Examination was 27, and BBB impairment was recorded in 13.5%. BBB breakdown was associated with cognitive decline (P = .015). Cerebrospinal fluid intercellular adhesion molecule-1, vascular endothelial growth factor, interleukin-8, serum amyloid A, macrophage derived chemokine, and gender generated an area under the curve of 0.95 for BBB impairment, and serum IL-16, VEGF-D, IL-15, and other variables generated an AUC of 0.92 for BBB impairment. DISCUSSION: BBB breakdown is associated with more rapid cognitive decline. Inflammatory mechanisms, including cell adhesion, neutrophil migration, lipid metabolism, and angiogenesis may be implicated.

ICP-MS/MS-Based Ionomics: A Validated Methodology to Investigate the Biological Variability of the Human Ionome.

We here describe the development, validation and application of a quantitative methodology for the simultaneous determination of 29 elements in human serum using state-of-the-art inductively coupled plasma triple quadrupole mass spectrometry (ICP-MS/MS). This new methodology offers high-throughput elemental profiling using simple dilution of minimal quantity of serum samples. We report the outcomes of the validation procedure including limits of detection/quantification, linearity of calibration curves, precision, recovery and measurement uncertainty. ICP-MS/MS-based ionomics was used to analyze human serum of 120 older adults. Following a metabolomic data mining approach, the generated ionome profiles were subjected to principal component analysis revealing gender and age-specific differences. The ionome of female individuals was marked by higher levels of calcium, phosphorus, copper and copper to zinc ratio, while iron concentration was lower with respect to male subjects. Age was associated with lower concentrations of zinc. These findings were complemented with additional readouts to interpret micronutrient status including ceruloplasmin, ferritin and inorganic phosphate. Our data supports a gender-specific compartmentalization of the ionome that may reflect different bone remodelling in female individuals. Our ICP-MS/MS methodology enriches the panel of validated "Omics" approaches to study molecular relationships between the exposome and the ionome in relation with nutrition and health.

Markers of neuroinflammation associated with Alzheimer's disease pathology in older adults.

BACKGROUND: In vitro and animal studies have linked neuroinflammation to Alzheimer's disease (AD) pathology. Studies on markers of inflammation in subjects with mild cognitive impairment or AD dementia provided inconsistent results. We hypothesized that distinct blood and cerebrospinal fluid (CSF) inflammatory markers are associated with biomarkers of amyloid and tau pathology in older adults without cognitive impairment or with beginning cognitive decline. OBJECTIVE: To identify blood-based and CSF neuroinflammation marker signatures associated with AD pathology (i.e. an AD CSF biomarker profile) and to investigate associations of inflammation markers with CSF biomarkers of amyloid, tau pathology, and neuronal injury. DESIGN/METHODS: Cross-sectional analysis was performed on data from 120 older community-dwelling adults with normal cognition (n=48) or with cognitive impairment (n=72). CSF Aß1-42, tau and p-tau181, and a panel of 37 neuroinflammatory markers in both CSF and serum were quantified. Least absolute shrinkage and selection operator (LASSO) regression was applied to determine a reference model that best predicts an AD CSF biomarker profile defined a priori as p-tau181/Aß1-42 ratio >0.0779. It was then compared to a second model that included the inflammatory markers from either serum or CSF. In addition, the correlations between inflammatory markers and CSF Aß1-42, tau and p-tau181 levels were assessed. RESULTS: Forty-two subjects met criteria for having an AD CSF biomarker profile. The best predictive models included 8 serum or 3 CSF neuroinflammatory markers related to cytokine mediated inflammation, vascular injury, and angiogenesis. Both models improved the accuracy to predict an AD biomarker profile when compared to the reference model. In analyses separately performed in the subgroup of participants with cognitive impairment, adding the serum or the CSF neuroinflammation markers also improved the accuracy of the diagnosis of AD pathology. None of the inflammatory markers correlated with the CSF Aß1-42 levels. Six CSF markers (IL-15, MCP-1, VEGFR-1, sICAM1, sVCAM-1, and VEGF-D) correlated with the CSF tau and p-tau181 levels, and these associations remained significant after controlling for age, sex, cognitive impairment, and APOEε4 status. CONCLUSIONS: The identified serum and CSF neuroinflammation biomarker signatures improve the accuracy of classification for AD pathology in older adults. Our results suggest that inflammation, vascular injury, and angiogenesis as reflected by CSF markers are closely related to cerebral tau pathology.

Homocysteine metabolism is associated with cerebrospinal fluid levels of soluble amyloid precursor protein and amyloid beta.

Disturbed homocysteine metabolism may contribute to amyloidogenesis by modulating the amyloid precursor protein (APP) production and processing. The objective of this study was to investigate the relationships between cerebral amyloid production and both blood and cerebrospinal fluid (CSF) markers of the homocysteine metabolism. We assessed CSF concentrations of soluble APPα, soluble APPß, and amyloid ß1-42 (Aß1-42), as well as plasma levels of homocysteine (Hcys), total vitamin B12, and folate, and CSF concentrations of homocysteine (Hcys-CSF), 5-methyltetrahydrofolate (5-MTHF), S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH) in 59 subjects with normal cognition. Linear regression analyses were performed to assess associations between homocysteine metabolism parameters and amyloid production. The study was approved by the Ethical Committee of the University of Bonn. After controlling for age, gender, APOEe4 status, and albumin ratio (Qalb), higher Aß1-42 CSF levels were associated with high Hcys and low vitamin B12 plasma levels as well as with high Hcys, high SAH, and low 5-MTHF CSF levels. Higher CSF concentrations of sAPPα and sAPPß were associated with high SAH levels. The results suggest that disturbed homocysteine metabolism is related to increased CSF levels of sAPP forms and Aß1-42, and may contribute to the accumulation of amyloid pathology in the brain. Disturbed homocysteine metabolism may contribute to amyloidogenesis by modulating the amyloid precursor protein (APP) production and processing. We found associations between CSF levels of soluble APP forms and Aß1-42, and markers of the homocysteine metabolism in both plasma and CSF in adults with normal cognition. Disturbed homocysteine metabolism may represent a target for preventive and early disease-modifying interventions in Alzheimer's disease.

Systemic and central nervous system metabolic alterations in Alzheimer's disease.

BACKGROUND: Metabolic alterations, related to cerebral glucose metabolism, brain insulin resistance, and age-induced mitochondrial dysfunction, play an important role in Alzheimer's disease (AD) on both the systemic and central nervous system level. To study the extent and significance of these alterations in AD, quantitative metabolomics was applied to plasma and cerebrospinal fluid (CSF) from clinically well-characterized AD patients and cognitively healthy control subjects. The observed metabolic alterations were associated with core pathological processes of AD to investigate their relation with amyloid pathology and tau-related neurodegeneration. METHODS: In a case-control study of clinical and biomarker-confirmed AD patients (n = 40) and cognitively healthy controls without cerebral AD pathology (n = 34) with paired plasma and CSF samples, we performed metabolic profiling, i.e., untargeted metabolomics and targeted quantification. Targeted quantification focused on identified deregulated pathways highlighted in the untargeted assay, i.e. the TCA cycle, and its anaplerotic pathways, as well as the neuroactive tryptophan and kynurenine pathway. RESULTS: Concentrations of several TCA cycle and beta-oxidation intermediates were higher in plasma of AD patients, whilst amino acid concentrations were significantly lower. Similar alterations in these energy metabolism intermediates were observed in CSF, together with higher concentrations of creatinine, which were strongly correlated with blood-brain barrier permeability. Alterations of several amino acids were associated with CSF Amyloidß1-42. The tryptophan catabolites, kynurenic acid and quinolinic acid, showed significantly higher concentrations in CSF of AD patients, which, together with other tryptophan pathway intermediates, were correlated with either CSF Amyloidß1-42, or tau and phosphorylated Tau-181. CONCLUSIONS: This study revealed AD-associated systemic dysregulation of nutrient sensing and oxidation and CNS-specific alterations in the neuroactive tryptophan pathway and (phospho)creatine degradation. The specific association of amino acids and tryptophan catabolites with AD CSF biomarkers suggests a close relationship with core AD pathology. Our findings warrant validation in independent, larger cohort studies as well as further investigation of factors such as gender and APOE genotype, as well as of other groups, such as preclinical AD, to identify metabolic alterations as potential intervention targets.

A global HILIC-MS approach to measure polar human cerebrospinal fluid metabolome: Exploring gender-associated variation in a cohort of elderly cognitively healthy subjects.

Cerebrospinal fluid (CSF) is a key body fluid that maintains the homeostasis in central nervous system (CNS). As a biofluid whose content reflects the brain metabolic activity, the CSF has been profiled in the context of neurological diseases to provide novel insights into the disease mechanisms. However, a global high-throughput approach to measure a broad diversity of polar metabolites present in CSF is lacking. Although still perceived as challenging and less reproducible, hydrophilic interaction liquid chromatography (HILIC) has recently evolved to offer the unprecedented coverage capacity of water-soluble metabolome. Here, we present a global HILIC high-resolution mass spectrometry-based (HRMS) approach that combines the profiling in acidic pH ESI (+) and basic pH ESI (-) mode to extend the coverage of CSF polar metabolome. This approach allowed us to annotate and measure a broad range of central carbon metabolites (implicated in glycolysis, TCA cycle, nucleotide, amino acid and fatty acid metabolism) in CSF collected from cognitively healthy elderly volunteers (n = 32), using a single extraction method. Metabolite annotation was achieved using the accurate mass, RT and MS/MS criteria, allowing for the characterization of 146 measurable metabolites. Exploration of characterized individual CSF profiles allowed for a discovery of intriguing gender-associated differences, with significantly higher acylcarnitine levels in men and higher taurine levels women. With this case study, we demonstrate the value of combined HILIC ESI ± HRMS profiling to assess CSF metabolome in clinical research studies.

Alzheimer disease pathology and the cerebrospinal fluid proteome.

BACKGROUND: Altered proteome profiles have been reported in both postmortem brain tissues and body fluids of subjects with Alzheimer disease (AD), but their broad relationships with AD pathology, amyloid pathology, and tau-related neurodegeneration have not yet been fully explored. Using a robust automated MS-based proteomic biomarker discovery workflow, we measured cerebrospinal fluid (CSF) proteomes to explore their association with well-established markers of core AD pathology. METHODS: Cross-sectional analysis was performed on CSF collected from 120 older community-dwelling adults with normal (n = 48) or impaired cognition (n = 72). LC-MS quantified hundreds of proteins in the CSF. CSF concentrations of ß-amyloid 1-42 (Aß1-42), tau, and tau phosphorylated at threonine 181 (P-tau181) were determined with immunoassays. First, we explored proteins relevant to biomarker-defined AD. Then, correlation analysis of CSF proteins with CSF markers of amyloid pathology, neuronal injury, and tau hyperphosphorylation (i.e., Aß1-42, tau, P-tau181) was performed using Pearson's correlation coefficient and Bonferroni correction for multiple comparisons. RESULTS: We quantified 790 proteins in CSF samples with MS. Four CSF proteins showed an association with CSF Aß1-42 levels (p value ≤ 0.05 with correlation coefficient (R) ≥ 0.38). We identified 50 additional CSF proteins associated with CSF tau and 46 proteins associated with CSF P-tau181 (p value ≤ 0.05 with R ≥ 0.37). The majority of those proteins that showed such associations were brain-enriched proteins. Gene Ontology annotation revealed an enrichment for synaptic proteins and proteins originating from reelin-producing cells and the myelin sheath. CONCLUSIONS: We used an MS-based proteomic workflow to profile the CSF proteome in relation to cerebral AD pathology. We report strong evidence of previously reported CSF proteins and several novel CSF proteins specifically associated with amyloid pathology or neuronal injury and tau hyperphosphorylation.

LC-HRMS data as a result of untargeted metabolomic profiling of human cerebrospinal fluid.

Cerebrospinal fluid (CSF) is a key body fluid that maintains the homeostasis in central nervous system (CNS). As a biofluid whose content reflects the brain metabolic activity, the CSF is analyzed in the context of neurological diseases and is rarely collected from healthy subjects. For this reason, the metabolite variation associated with general phenotypic characteristics such as gender and age have hardly ever been studied. Here we present the hydrophilic interaction liquid chromatography-high resolution mass spectrometry (HILIC-HRMS) data as a result of untargeted metabolomics analysis of a cohort of elderly cognitively healthy volunteers (n = 32). 146 unambiguously identified water soluble metabolites (using accurate mass, retention time and MS/MS matching against spectral libraries) were measured and their abundances across all the subjects depending on their gender are provided in this article. Data tables are available at https://data.mendeley.com/datasets/c73xtsd4s5/1. it's published on mendeley, the DOI is DOI:10.17632/c73xtsd4s5.1. The data presented in this article are related to the research article entitled "A global HILIC-MS approach to measure polar human cerebrospinal fluid metabolome: Exploring gender-associated variation in a cohort of elderly cognitively healthy subjects" (Gallart-Ayala et al., 2018, In press).

Macrophage Migration Inhibitory Factor is Associated with Biomarkers of Alzheimer's Disease Pathology and Predicts Cognitive Decline in Mild Cognitive Impairment and Mild Dementia.

BACKGROUND: Macrophage migration inhibitory factor (MIF) is a pro-inflammatory protein playing a regulatory role in the immune response. First evidence from in vitro and animal studies suggests that MIF may be involved in the development of Alzheimer's disease (AD) pathology. OBJECTIVE: To address in older subjects (i) the relationships between AD pathology and MIF plasma and cerebrospinal fluid (CSF) levels; and (ii) to investigate whether increased MIF-related systemic and CNS inflammation is associated with clinical disease progression. METHODS: CSF and plasma concentrations of MIF as well as biomarkers of amyloid, neuronal injury, and tau hyperphosphorylation (CSF Aß1-42, tau, and ptau, respectively) were assessed in 97 subjects with MCI or mild dementia (cognitive impairment, CI) and 52 healthy volunteers with normal cognition. Clinical and neuropsychological evaluations were performed at inclusion and at follow up visits. RESULTS: CSF MIF levels were higher in participants with CI with an AD CSF biomarker profile, but not in CI with a non-AD profile, compared to the healthy controls. Higher MIF CSF levels were associated with higher CSF tau and ptau and lower CSF Aß1-42 after adjusting for potential confounders. In CI, MIF CSF independently predicted cognitive decline at a follow-up visit after controlling for potential confounders including CSF Aß1-42 and tau levels. CONCLUSION: Our study provides evidence that MIF-related inflammation is related to amyloid pathology, tau hyperphosphorylation, and neuronal injury at the early clinical stages of AD. Higher MIF CSF levels are associated with accelerated cognitive decline in MCI and mild dementia.

Plasma Proteomic Profiles of Cerebrospinal Fluid-Defined Alzheimer's Disease Pathology in Older Adults.

BACKGROUND: Cerebrospinal fluid (CSF) biomarkers of the beta-amyloid and microtubule associated protein tau metabolism have proven the capacity to improve classification of subjects developing Alzheimer's disease (AD). The blood plasma proteome was characterized to further elaborate upon the mechanisms involved and identify proteins that may improve classification of older adults developing an AD dementia. OBJECTIVE: Identify and describe plasma protein expressions that best classify subjects with CSF-defined presence of AD pathology and cerebral amyloidosis. METHODS: We performed a cross-sectional analysis of samples collected from community-dwelling elderly with (n = 72) or without (n = 48) cognitive impairment. CSF Aß1-42, tau, and phosphorylated tau (P-tau181) were measured using ELISA, and mass spectrometry quantified the plasma proteomes. Presence of AD pathology was defined as CSF P-tau181/Aß1-42 > 0.0779, and presence of amyloidosis was defined as CSF Aß1-42 < 724 pg/mL. RESULTS: Two hundred and forty-eight plasma proteins were quantified. Plasma proteins did not improve classification of the AD CSF biomarker profile in the whole sample. When the analysis was separately performed in the cognitively impaired individuals, the diagnosis accuracy of AD CSF profile was 88.9% with 19 plasma proteins included. Within the full cohort, there were 16 plasma proteins that improved diagnostic accuracy of cerebral amyloidosis to 92.4%. CONCLUSION: Plasma proteins improved classification accuracy of AD pathology in cognitively-impaired older adults and appeared representative of amyloid pathology. If confirmed, those candidates could serve as valuable blood biomarkers of the preclinical stages of AD or risk of developing AD.

One-carbon metabolism, cognitive impairment and CSF measures of Alzheimer pathology: homocysteine and beyond.

BACKGROUND: Hyperhomocysteinemia is a risk factor for cognitive decline and dementia, including Alzheimer disease (AD). Homocysteine (Hcy) is a sulfur-containing amino acid and metabolite of the methionine pathway. The interrelated methionine, purine, and thymidylate cycles constitute the one-carbon metabolism that plays a critical role in the synthesis of DNA, neurotransmitters, phospholipids, and myelin. In this study, we tested the hypothesis that one-carbon metabolites beyond Hcy are relevant to cognitive function and cerebrospinal fluid (CSF) measures of AD pathology in older adults. METHODS: Cross-sectional analysis was performed on matched CSF and plasma collected from 120 older community-dwelling adults with (n = 72) or without (n = 48) cognitive impairment. Liquid chromatography-mass spectrometry was performed to quantify one-carbon metabolites and their cofactors. Least absolute shrinkage and selection operator (LASSO) regression was initially applied to clinical and biomarker measures that generate the highest diagnostic accuracy of a priori-defined cognitive impairment (Clinical Dementia Rating-based) and AD pathology (i.e., CSF tau phosphorylated at threonine 181 [p-tau181]/ß-Amyloid 1-42 peptide chain [Aß1-42] >0.0779) to establish a reference benchmark. Two other LASSO-determined models were generated that included the one-carbon metabolites in CSF and then plasma. Correlations of CSF and plasma one-carbon metabolites with CSF amyloid and tau were explored. LASSO-determined models were stratified by apolipoprotein E (APOE) ε4 carrier status. RESULTS: The diagnostic accuracy of cognitive impairment for the reference model was 80.8% and included age, years of education, Aß1-42, tau, and p-tau181. A model including CSF cystathionine, methionine, S-adenosyl-L-homocysteine (SAH), S-adenosylmethionine (SAM), serine, cysteine, and 5-methyltetrahydrofolate (5-MTHF) improved the diagnostic accuracy to 87.4%. A second model derived from plasma included cystathionine, glycine, methionine, SAH, SAM, serine, cysteine, and Hcy and reached a diagnostic accuracy of 87.5%. CSF SAH and 5-MTHF were associated with CSF tau and p-tau181. Plasma one-carbon metabolites were able to diagnose subjects with a positive CSF profile of AD pathology in APOE ε4 carriers. CONCLUSIONS: We observed significant improvements in the prediction of cognitive impairment by adding one-carbon metabolites. This is partially explained by associations with CSF tau and p-tau181, suggesting a role for one-carbon metabolism in the aggregation of tau and neuronal injury. These metabolites may be particularly critical in APOE ε4 carriers.