Multiplex proteomics identifies novel CSF and plasma biomarkers of early Alzheimer's disease.

To date, the development of disease-modifying therapies for Alzheimer's disease (AD) has largely focused on the removal of amyloid beta Aß fragments from the CNS. Proteomic profiling of patient fluids may help identify novel therapeutic targets and biomarkers associated with AD pathology. Here, we applied the Olink™ ProSeek immunoassay to measure 270 CSF and plasma proteins across 415 Aß- negative cognitively normal individuals (Aß- CN), 142 Aß-positive CN (Aß+ CN), 50 Aß- mild cognitive impairment (MCI) patients, 75 Aß+ MCI patients, and 161 Aß+ AD patients from the Swedish BioFINDER study. A validation cohort included 59 Aß- CN, 23 Aß- + CN, 44 Aß- MCI and 53 Aß+ MCI. To compare protein concentrations in patients versus controls, we applied multiple linear regressions adjusting for age, gender, medications, smoking and mean subject-level protein concentration, and corrected findings for false discovery rate (FDR, q < 0.05). We identified, and replicated, altered levels of ten CSF proteins in Aß+ individuals, including CHIT1, SMOC2, MMP-10, LDLR, CD200, EIF4EBP1, ALCAM, RGMB, tPA and STAMBP (- 0.14 < d < 1.16; q < 0.05). We also identified and replicated alterations of six plasma proteins in Aß+ individuals OSM, MMP-9, HAGH, CD200, AXIN1, and uPA (- 0.77 < d < 1.28; q < 0.05). Multiple analytes associated with cognitive performance and cortical thickness (q < 0.05). Plasma biomarkers could distinguish AD dementia (AUC = 0.94, 95% CI = 0.87-0.98) and prodromal AD (AUC = 0.78, 95% CI = 0.68-0.87) from CN. These findings reemphasize the contributions of immune markers, phospholipids, angiogenic proteins and other biomarkers downstream of, and potentially orthogonal to, Aß- and tau in AD, and identify candidate biomarkers for earlier detection of neurodegeneration.

CSF biomarkers of neuroinflammation and cerebrovascular dysfunction in early Alzheimer disease.

OBJECTIVE: To measure CSF levels of biomarkers reflecting microglia and astrocytes activation, neuroinflammation, and cerebrovascular changes and study their associations with the core biomarkers of Alzheimer disease (AD) pathology (ß-amyloid [Aß] and tau), structural imaging correlates, and clinical disease progression over time. METHODS: The study included cognitively unimpaired elderly (n = 508), patients with mild cognitive impairment (MCI, n = 256), and patients with AD dementia (n = 57) from the longitudinal Swedish BioFINDER cohort. CSF samples were analyzed for YKL-40, interleukin (IL)-6, IL-7, IL-8, IL-15, IP-10, monocyte chemoattractant protein 1, intercellular adhesion molecule 1 (ICAM-1), vascular adhesion molecule 1 (VCAM-1), placental growth factor, and fms-related tyrosine kinase 1 (Flt-1). MRI data were available from 677 study participants. Longitudinal clinical assessments were conducted in control individuals and patients with MCI (mean follow-up 3 years, range 1-6 years). RESULTS: CSF levels of YKL-40, ICAM-1, VCAM-1, IL-15, and Flt-1 were increased during the preclinical, prodromal, and dementia stages of AD. High levels of these biomarkers were associated with increased CSF levels of total tau, with the associations, especially for YKL-40, being stronger in Aß-positive individuals. The results were similar for associations between phosphorylated tau and YKL-40, ICAM-1, and VCAM-1. High levels of the biomarkers were also associated with cortical thinning (primarily in the precuneus and superior parietal regions) and with subsequent cognitive deterioration in patients without dementia as measured with Mini-Mental State Examination (YKL-40) and Clinical Dementia Rating Sum of Boxes (YKL-40, ICAM-1, VCAM-1 and IL-15). Finally, higher levels of CSF YKL-40, ICAM-1, and Flt-1 increased risk of development of AD dementia in patients without dementia. CONCLUSIONS: Neuroinflammation and cerebrovascular dysfunction are early events occurring already at presymptomatic stages of AD and contribute to disease progression.

Effects of surgery and propofol-remifentanil total intravenous anesthesia on cerebrospinal fluid biomarkers of inflammation, Alzheimer's disease, and neuronal injury in humans: a cohort study.

BACKGROUND: Surgery and anesthesia have been linked to postoperative cognitive disturbance and increased risk of Alzheimer's disease. It is not clear by which mechanisms this increased risk for cognitive disease is mediated. Further, amyloid ß production has been suggested to depend on the sleep-wake cycle and neuronal activity. The aim of the present study was to examine if cerebrospinal fluid (CSF) concentrations of a number of biomarkers for Alzheimer's disease-related processes, including amyloid ß, neuronal injury, and inflammation, changed over time during intravenous anesthesia in surgical patients. METHODS: We included patients scheduled for hysterectomy via laparotomy during general anesthesia with intravenous propofol and remifentanil. CSF samples were obtained before, during, and after surgery (5 h after induction) and tested for 27 biomarkers. Changes over time were tested with linear mixed effects models. RESULTS: A total of 22 patients, all females, were included. The mean age was 50 years (± 9 SD). The mean duration of the anesthesia was 145 min (± 40 SD). Interleukin (IL)-6, IL-8, monocyte chemoattractant protein 1, and vascular endothelial growth factor A increased over time. IL-15 and IL-7 decreased slightly over time. Macrophage inflammatory protein 1ß and placental growth factor also changed significantly. There were no significant effects on amyloid ß (Aß) or tau biomarkers. CONCLUSIONS: Surgery and general anesthesia with intravenous propofol and remifentanil induce, during and in the short term after the procedure, a neuroinflammatory response which is dominated by monocyte attractants, without biomarker signs of the effects on Alzheimer's disease pathology or neuronal injury.

A novel quantification-driven proteomic strategy identifies an endogenous peptide of pleiotrophin as a new biomarker of Alzheimer's disease.

We present a new, quantification-driven proteomic approach to identifying biomarkers. In contrast to the identification-driven approach, limited in scope to peptides that are identified by database searching in the first step, all MS data are considered to select biomarker candidates. The endopeptidome of cerebrospinal fluid from 40 Alzheimer's disease (AD) patients, 40 subjects with mild cognitive impairment, and 40 controls with subjective cognitive decline was analyzed using multiplex isobaric labeling. Spectral clustering was used to match MS/MS spectra. The top biomarker candidate cluster (215% higher in AD compared to controls, area under ROC curve = 0.96) was identified as a fragment of pleiotrophin located near the protein's C-terminus. Analysis of another cohort (n = 60 over four clinical groups) verified that the biomarker was increased in AD patients while no change in controls, Parkinson's disease or progressive supranuclear palsy was observed. The identification of the novel biomarker pleiotrophin 151-166 demonstrates that our quantification-driven proteomic approach is a promising method for biomarker discovery, which may be universally applicable in clinical proteomics.

Strategic roadmap for an early diagnosis of Alzheimer's disease based on biomarkers.

The diagnosis of Alzheimer's disease can be improved by the use of biological measures. Biomarkers of functional impairment, neuronal loss, and protein deposition that can be assessed by neuroimaging (ie, MRI and PET) or CSF analysis are increasingly being used to diagnose Alzheimer's disease in research studies and specialist clinical settings. However, the validation of the clinical usefulness of these biomarkers is incomplete, and that is hampering reimbursement for these tests by health insurance providers, their widespread clinical implementation, and improvements in quality of health care. We have developed a strategic five-phase roadmap to foster the clinical validation of biomarkers in Alzheimer's disease, adapted from the approach for cancer biomarkers. Sufficient evidence of analytical validity (phase 1 of a structured framework adapted from oncology) is available for all biomarkers, but their clinical validity (phases 2 and 3) and clinical utility (phases 4 and 5) are incomplete. To complete these phases, research priorities include the standardisation of the readout of these assays and thresholds for normality, the evaluation of their performance in detecting early disease, the development of diagnostic algorithms comprising combinations of biomarkers, and the development of clinical guidelines for the use of biomarkers in qualified memory clinics.

Increased amyloidogenic APP processing in APOE ɛ4-negative individuals with cerebral ß-amyloidosis.

Increased APP (amyloid precursor protein) processing causes ß-amyloid (Aß) accumulation in autosomal dominant Alzheimer's disease (AD), but it is unclear if it also affects sporadic Aß accumulation. We tested healthy controls and patients with mild cognitive symptoms (N=331) in the BioFINDER study, using cerebrospinal fluid (CSF) Aß40 as a surrogate for amyloidogenic APP processing. We find that levels of brain Aß fibrils (measured by 18F-flutemetamol PET) are independently associated with high CSF Aß40 (P<0.001) and APOE ɛ4 (P<0.001). The association between CSF Aß40 and brain Aß is stronger in APOE ɛ4-negative than in positive people (P=0.0080). The results are similar for CSF Aß38 and for a combination of CSF Aß38 and CSF Aß40. In conclusion, sporadic Aß accumulation may be partly associated with increased amyloidogenic APP production, especially in APOE ɛ4-negative subjects. The risk for sporadic AD may consequently depend on increased Aß production, in addition to decreased Aß clearance.

Interaction of Cigarette Smoking History With APOE Genotype and Age on Amyloid Level, Glucose Metabolism, and Neurocognition in Cognitively Normal Elders.

INTRODUCTION: Chronic cigarette smoking is associated with increased risk for Alzheimer's disease (AD). The goal of this study was to determine if smoking history moderated the associations of age and APOE genotype (the most robust risk factors for AD) on brain amyloid deposition, glucose metabolism, and neurocognition in cognitively-normal elders. METHODS: Participants (n = 264) were grouped according to their APOE ε4 carrier status (ε4 carrier: APOE4+; non-ε4 carrier: APOE4-) and smoking status (smokers: at least 1 year of smoking during lifetime; never-smokers: no history of smoking). Approximately 89% of the smoking sample was former-smokers. We specifically tested for interactions of smoking status with APOE ε4 carrier status and age on measures of cortical amyloid deposition, glucose metabolism, and neurocognition. RESULTS: (1) smoking status interacted with APOE ε4 carrier status, where smoker APOE4+ showed lower glucose metabolism and poorer auditory-verbal learning and memory than never-smoking APOE4-, never-smoking APOE4+, and smoking APOE4-; (2) smoking status interacted with age on measures of semantic fluency, processing speed/set-shifting and global neurocognition; smokers, irrespective of APOE ε4 carrier status, demonstrated poorer performance with increasing age than never-smokers; and (3) smoking APOE4+ and never-smoking APOE4+ showed greater cortical amyloid deposition than never-smoking APOE4- and smoking APOE4-. CONCLUSIONS: The findings indicate consideration of smoking history is essential to both better understand the factors associated with neurobiological and neurocognitive abnormalities in elders, and the risk for development of AD-related neuropathology.

CSF in Alzheimer's disease.

Alzheimer's disease (AD) is a progressive brain amyloidosis that injures brain regions involved in memory consolidation and other cognitive functions. Neuropathologically, the disease is characterized by accumulation of a 42-amino acid protein called amyloid beta, and N-terminally truncated fragments thereof, in extracellular senile plaques together with intraneuronal inclusions of hyperphosphorylated tau protein in neurofibrillary tangles, and neuronal and axonal degeneration and loss. Clinical chemistry tests for these pathologies have been developed for use on cerebrospinal fluid samples. Here, we review what these markers have taught us on the disease process in AD and how they can be implemented in routine clinical chemistry. We also provide an update on new marker development and ongoing analytical standardization effort.

Exploring Alzheimer molecular pathology in Down's syndrome cerebrospinal fluid.

BACKGROUND: Individuals with Down's syndrome (DS) develop early Alzheimer's disease (AD) with ß-amyloid (Aß) plaque pathology. The extra amyloid precursor protein (APP) gene copy in DS is believed to result in a 50% increase in Aß production, but it is unclear how this relates to the development of other AD hallmarks, including axonal degeneration and microglia cell activation, and to other neurological problems in DS, including disturbed sleep regulation. OBJECTIVE: To evaluate if cerebrospinal fluid (CSF) biomarkers for cerebral amyloidosis, axonal degeneration, microglial activation and sleep regulation were altered in young and old patients with DS, and if these biomarkers were related to altered Aß and APP metabolism, reflected by CSF levels of different Aß and APP peptides. METHODS: CSF from DS patients (n=12) and healthy controls (n=20) were analyzed for Aß peptides (Aß1-42, AßX-38/40/42), secreted APP species (sAPPα/ß), biomarkers for AD-like axonal degeneration [total tau (T-tau), phosphorylated tau], microglial activation (YKL-40, CC chemokine ligand 2) and orexin-A, which is a peptide involved in sleep regulation. We compared biomarker levels between groups and tested for relations between biomarkers, disease stage and age. RESULTS: Several of the markers were specifically increased in DS, including AßX-40, sAPPα and sAPPß. Οrexin-A was significantly decreased in DS and correlated with Aß and sAPP. Orexin-A decreased with age in DS, while T-tau and YKL-40 increased with age. CONCLUSION: Down's patients have increased APP and Aß production and increased microglial activation with age. The orexin-A metabolism is disturbed in DS and may be linked to APP and Aß production. Biomarker studies of DS may contribute to our understanding of the amyloidogenic and neurodegenerative process in AD.

History of cigarette smoking in cognitively-normal elders is associated with elevated cerebrospinal fluid biomarkers of oxidative stress.

BACKGROUND: Cigarette smoking in adults is associated with abnormalities in brain neurobiology. Smoking-induced central nervous system oxidative stress (OxS) is a potential mechanism associated with these abnormalities. The goal of this study was to compare cognitively-normal elders on cerebrospinal fluid (CSF) levels of F2-isoprostane biomarkers of OxS. METHODS: Elders with a lifetime history of smoking (smokers; n=50; 75±5 years of age; 34±28 pack-years; approximately 12% were actively smoking at the time of study) were compared to never-smokers (n=61; 76±6 years of age) on CSF iPF2α-III and 8,12, iso-iPF2α-VI F2-isoprostanes levels. F2-isoprostanes levels were quantitated with HPLC-atmospheric pressure chemical ionization-tandem mass spectrometry. Associations between F2-isoprostanes levels, hippocampal volumes, and cigarette exposure measures were also evaluated. RESULTS: Smokers showed higher iPF2α-III level than never-smokers. An age×smoking status interaction was observed for 8,12, iso-iPF2α-VI, where smokers demonstrate a significantly greater concentration with increasing age than never-smokers. In smokers only, higher 8,12, iso-iPF2α-VI concentration was associated with smaller hippocampal volume, and greater iPF2α-III level was related to greater pack years. CONCLUSIONS: This is the first study to demonstrate that a history of cigarette smoking in cognitively-normal elders was associated with significantly elevated CSF F2-isoprostane levels and greater age-related increases in F2-isoprostanes, and that higher F2-isoprostane levels in smokers were related to smaller hippocampal volume. These findings provide additional novel evidence that a history of chronic smoking during adulthood is associated with adverse effects on the human brain that are potentially enduring even with extended smoking cessation.

Smoking and increased Alzheimer's disease risk: a review of potential mechanisms.

BACKGROUND: Cigarette smoking has been linked with both increased and decreased risk for Alzheimer's disease (AD). This is relevant for the US military because the prevalence of smoking in the military is approximately 11% higher than in civilians. METHODS: A systematic review of published studies on the association between smoking and increased risk for AD and preclinical and human literature on the relationships between smoking, nicotine exposure, and AD-related neuropathology was conducted. Original data from comparisons of smoking and never-smoking cognitively normal elders on in vivo amyloid imaging are also presented. RESULTS: Overall, literature indicates that former/active smoking is related to a significantly increased risk for AD. Cigarette smoke/smoking is associated with AD neuropathology in preclinical models and humans. Smoking-related cerebral oxidative stress is a potential mechanism promoting AD pathology and increased risk for AD. CONCLUSIONS: A reduction in the incidence of smoking will likely reduce the future prevalence of AD.

Understanding the cause of sporadic Alzheimer's disease.

The increasing prevalence of Alzheimer's disease (AD) and a lack of effective prevention or disease-modifying therapies are global challenges with devastating personal, social and economic consequences. The amyloid ß (Aß) hypothesis posits that cerebral ß-amyloidosis is a critical early event in AD pathogenesis. However, failed clinical trials of Aß-centric drug candidates have called this hypothesis into question. Whereas we acknowledge that the Aß hypothesis is far from disproven, we here re-visit the links between Aß, tau and neurodegeneration. We review the genetics, epidemiology and pathology of sporadic AD and give an updated account of what is currently known about the molecular pathogenesis of the disease.

Assessment of peptide chemical modifications on the development of an accurate and precise multiplex selected reaction monitoring assay for apolipoprotein e isoforms.

Apolipoprotein E (ApoE) is a polymorphic protein that plays a major role in lipid metabolism in the central nervous system and periphery. It has three common allelic isoforms, ApoE2, ApoE3, and ApoE4, that differ in only one or two amino acids. ApoE isoforms have been associated with the occurrence and progression of several pathological conditions, such as coronary atherosclerosis and Alzheimer's disease. The aim of this study was to develop a mass spectrometry (MS)-based assay for absolute quantification of ApoE isoforms in cerebrospinal fluid and plasma samples using isotope-labeled peptides. The assay included five tryptic peptides: CLAVYQAGAR (ApoE2), LGADMEDVCGR (ApoE2 and 3), LAVYQAGAR (ApoE3 and 4), LGADMEDVR (ApoE4), and LGPLVEQGR (total ApoE). Both cerebrospinal fluid and plasma samples were assayed to validate the method. The digestion yield and the extension of chemical modifications in selected amino acid residues (methionine oxidation, glutamine deamidation, and cyclization of N-terminus carbamidomethylcysteine) were also studied. The ApoE phenotype was successfully assigned to all samples analyzed in a blinded manner. The method showed good linearity (R(2) > 0.99) and reproducibility (within laboratory imprecision <13%). The comparison of the MS-based assay with an ELISA for total ApoE concentration showed a moderate correlation (R(2) = 0.59). This MS-based assay can serve as an important tool in clinical studies aiming to elucidate the association between ApoE genotype, total ApoE, and ApoE isoform concentrations in various disorders related to ApoE polymorphisms.

Cerebrospinal fluid brain injury biomarkers in children: a multicenter study.

BACKGROUND: Cerebrospinal fluid (CSF) biomarkers reflecting neuronal and astroglial injury, such as total tau (T-tau), glial fibrillary acidic protein (GFAP), and neurofilament light (NFL), have been extensively investigated in neurologic diseases in adults, but no large study has investigated these biomarkers in children. METHODS: This study presents a detailed evaluation of CFS T-tau, GFAP, NFL, and CSF:albumin ratio in a large cohort of pediatric patients. This is a retrospective multicenter study on pediatric patients aged <16 years (n = 607), where neuronal injury biomarkers T-tau, GFAP, NFL, and CSF albumin ratio were analyzed during 2000-2010 at the Clinical Neurochemistry Laboratory, Sahlgrenska University Hospital, Sweden. The patients were grouped into eight categories: epilepsy, infectious and inflammatory central nervous system disorders, progressive encephalopathy, static encephalopathy, tumors, movement disorders, miscellaneous disorders, and a control group. RESULTS: T-tau, GFAP, and NFL were increased in progressive encephalopathy (P < 0.001), epilepsy (P < 0.001), and infectious and inflammatory central nervous system disorders (P < 0.001) compared with controls. T-tau was the biomarker with the highest diagnostic accuracy with the area under the curve of 0.83 (95% confidence interval (CI), 0.77-0.90; P < 0.0001) for progressive encephalopathy followed by epilepsy 0.80 (95% CI, 0.75-0.87; P < 0.0001). The combination of all four biomarkers further improved the area under the curve for the progressive encephalopathy 0.87 (95% CI, 0.77-0.89; P < 0.0001), followed by epilepsy 0.81 (95% CI, 0.74-0.80; P = 0.030). The combination of the biomarkers also separated progressive from static encephalopathy 0.88 (95% CI, 0.83-0.93; P < 0.0001). CONCLUSIONS: CSF T-tau, GFAP, and NFL are differently altered across different neurologic diseases in children. Importantly, the biomarker pattern distinguishes between progressive and static neurologic disorders.

Serum levels of LIGHT in MS.

BACKGROUND: Recently, a polymorphism in the LIGHT gene was shown to increase the risk of multiple sclerosis (MS) in a genome-wide association study (GWAS). OBJECTIVE: Our aim was to investigate if serum levels of LIGHT were affected by this polymorphism and by the disease itself. METHODS: Serum levels of LIGHT were investigated in four cohorts; 1) MS (n = 159) and controls (n = 160) in relation to rs1077667 genotype; 2) MS at relapse (n = 30) vs. healthy controls (n = 26); 3) MS (n = 27) vs. other neurological disease (OND, n = 33); and 4) MS patients before and after one year of treatment with natalizumab (n = 30). RESULTS: Carriers of the GG genotype had the lowest serum levels of LIGHT (p=0.02). Serum levels of LIGHT were increased in MS at relapse in two separate cohorts: vs. healthy controls (p=0.00005) and vs. remission (p=0.00006), other neurological disease (OND) (p=0.002) and OND with signs of inflammation (iOND; p=0.00005). Furthermore, serum levels of LIGHT were decreased by natalizumab treatment (p=0.001). CONCLUSION: Soluble LIGHT is an inhibitor of T-cell activation and GG carriers of rs1077667, with the highest risk for MS, had the lowest serum levels. The increased levels of LIGHT at times of increased MS activity suggest that soluble LIGHT is protective and may act to limit inflammation.

Amyloid-ß metabolism in Niemann-Pick C disease models and patients.

Niemann-Pick type C (NPC) is a progressive neurodegenerative lysosomal disease with altered cellular lipid trafficking. The metabolism of amyloid-ß (Aß) - previously mainly studied in Alzheimer's disease - has been suggested to be altered in NPC. Here we aimed to perform a detailed characterization of metabolic products from the amyloid precursor protein (APP) in NPC models and patients. We used multiple analytical technologies, including immunoassays and immunoprecipitation followed by mass spectrometry (IP-MS) to characterize Aß peptides and soluble APP fragments (sAPP-α/ß) in cell media from pharmacologically (U18666A) and genetically (NPC1 ( -/- ) ) induced NPC cell models, and cerebrospinal fluid (CSF) from NPC cats and human patients. The pattern of Aß peptides and sAPP-α/ß fragments in cell media was differently affected by NPC-phenotype induced by U18666A treatment and by NPC1 ( -/- ) genotype. U18666A treatment increased the secreted media levels of sAPP-α, AßX-40 and AßX-42 and reduced the levels of sAPP-ß, Aß1-40 and Aß1-42, while IP-MS showed increased relative levels of Aß5-38 and Aß5-40 in response to treatment. NPC1 ( -/- ) cells had reduced media levels of sAPP-α and Aß1-16, and increased levels of sAPP-ß. NPC cats had altered CSF distribution of Aß peptides compared with normal cats. Cats treated with the potential disease-modifying compound 2-hydroxypropyl-ß-cyclodextrin had increased relative levels of short Aß peptides including Aß1-16 compared with untreated cats. NPC patients receiving ß-cyclodextrin had reduced levels over time of CSF Aß1-42, AßX-38, AßX-40, AßX-42 and sAPP-ß, as well as reduced levels of the axonal damage markers tau and phosphorylated tau. We conclude that NPC models have altered Aß metabolism, but with differences across experimental systems, suggesting that NPC1-loss of function, such as in NPC1 ( -/- ) cells, or NPC1-dysfunction, seen in NPC patients and cats as well as in U18666A-treated cells, may cause subtle but different effects on APP degradation pathways. The preliminary findings from NPC cats suggest that treatment with cyclodextrin may have an impact on APP processing pathways. CSF Aß, sAPP and tau biomarkers were dynamically altered over time in human NPC patients.

Peptidome analysis of cerebrospinal fluid by LC-MALDI MS.

We report on the analysis of endogenous peptides in cerebrospinal fluid (CSF) by mass spectrometry. A method was developed for preparation of peptide extracts from CSF. Analysis of the extracts by offline LC-MALDI MS resulted in the detection of 3,000-4,000 peptide-like features. Out of these, 730 peptides were identified by MS/MS. The majority of these peptides have not been previously reported in CSF. The identified peptides were found to originate from 104 proteins, of which several have been reported to be involved in different disorders of the central nervous system. These results support the notion that CSF peptidomics may be viable complement to proteomics in the search of biomarkers of CNS disorders.

Cerebrospinal fluid microglial markers in Alzheimer's disease: elevated chitotriosidase activity but lack of diagnostic utility.

Activated microglial cells, which are the resident macrophages of the central nervous system, surround amyloid ß-plaques in Alzheimer's disease (AD) brains. Inflammation including microglial activation may contribute in AD pathogenesis, and biomarkers for this process may thus be of value to study AD pathogenesis and might facilitate development of therapies targeting these cells. We therefore examined cerebrospinal fluid (CSF) biomarkers in patients with AD, other dementias, mild cognitive impairment and in healthy controls. Samples were analyzed for markers with known association to macrophage activity, including chitotriosidase, YKL-40 (CHI3L1, HC gp-39) and chemokine CC motif ligand 2 (CCL2, MCP1). Patients with AD had higher chitotriosidase activity than controls and patients with stable mild cognitive impairment, consistent with the presence of activated microglial cells in AD brains, but with large overlaps between groups. CCL2 and YKL-40 concentrations did not differ among groups. Microglial markers are unlikely to be useful for AD diagnosis, but might be useful for identification of distinct subgroups of patients, and for the development and implementation of drugs targeting microglial pathology.

Converging pathways of chromogranin and amyloid metabolism in the brain.

Much is unknown regarding the regulation of Alzheimer-related amyloid-beta protein precursor (AbetaPP)-processing in the human central nervous system. It has been hypothesized that amyloidogenic AbetaPP-processing preferentially occurs in the regulated secretory pathway of neurons. To test this hypothesis we looked for correlations of AbetaPP-derived molecules in cerebrospinal fluid (CSF) with chromogranin (Cg) derived peptides, representing the regulated secretion. Patients with Alzheimer's disease (AD, N=32), multiple sclerosis (MS, N=50), and healthy controls (N= 70) were enrolled. CSF was analyzed for the amyloid peptides Abeta1-42, Abetax-42, Abetax-40, Abetax-38, alpha-cleaved soluble AbetaPP (sAbetaPPalpha), beta-cleaved soluble AbetaPP (sAbetaPPbeta), and peptides derived from CgB and SgII (Secretogranin-II, CgC). We investigated CSF levels of the protease BACE1, which processes AbetaPP into Abeta, in relation to Cg-levels. Finally, we measured Cg levels in cell media from untreated and BACE1-inhibited SH-SY5Y human neuroblastoma cells. CSF Cg levels correlated to sAbetaPP and Abeta peptides in AD, MS, and controls, and to CSF BACE1. Cell medium from BACE1-inhibited cells had decreased CgB levels. These results suggest that a large part of AbetaPP in the human central nervous system is processed in the regulated secretory pathway of neurons.