Association of APOE-ε4 and GAP-43-related presynaptic loss with ß-amyloid, tau, neurodegeneration, and cognitive decline.

Apolipoprotein E-ε4 (APOE-ε4) carriers had elevated cerebrospinal fluid (CSF) presynaptic protein growth-associated protein-43 (GAP-43), but the underlying mechanism is not fully understood. We investigated how the APOE-ε4 genotype affects the baseline and longitudinal changes in CSF GAP-43 and their associations with ß-amyloid positron emission tomography (Aß PET), CSF phosphorylated tau 181 (p-Tau181), neurodegeneration, and cognitive decline. Compared to APOE-ε4 non-carriers, APOE-ε4 carriers had higher baseline levels and faster rates of increases in Aß PET, CSF p-Tau181, and CSF GAP-43. Both higher baseline levels and faster rates of increase in CSF GAP-43 were associated with greater baseline Aß PET and CSF p-Tau181, which fully mediated the APOE-ε4 effect on CSF GAP-43 elevations. Independent of Aß PET and CSF p-Tau181, APOE-ε4 carriage was associated with exacerbated GAP-43-related longitudinal hippocampal atrophy and cognitive decline, especially in Aß+ participants (GAP-43 × time × APOE-ε4). These findings suggest that the APOE-ε4 effect on GAP-43-related presynaptic dysfunction is mediated by primary Alzheimer's pathologies and independently correlates to hippocampal atrophy and cognitive decline in the future.

Cerebrospinal fluid growth-associated protein 43 levels in patients with progressive and stable mild cognitive impairment.

BACKGROUND: Cerebrospinal fluid (CSF) growth-associated protein 43 (GAP-43) is prominently elevated in Alzheimer's disease (AD) dementia patients in comparison to normal controls. CSF GAP-43 levels in mild cognitive impairment (MCI) individuals who have different clinical trajectories need to be studied. METHODS: We examined 137 cognitively normal (CN) controls, 218 stable MCI patients (sMCI), 99 progressive MCI (pMCI) patients, and 120 AD dementia patients. Associations between the CSF GAP-43 levels and the four diagnosis groups were evaluated with multiple-variable linear regression. The relationships between CSF GAP-43 and core CSF biomarkers were assessed by Spearman correlations. Cox regression analysis was performed to assess the values of GAP-43 in predicting MCI conversion. We examined associations between baseline CSF GAP-43 levels and longitudinal cognitive function, hippocampal volumes, and brain glucose metabolism using linear mixed-effects models. RESULTS: CSF GAP-43 was elevated in the pMCI and AD groups in comparison to the CN group and in the pMCI and AD groups in comparison to the sMCI group. CSF GAP-43 significantly predicted conversion from MCI to AD. CSF GAP-43 was a significant predictor of cognitive decline, hippocampal atrophy, and brain hypometabolism over time. Furthermore, elevated CSF GAP-43 levels were associated with accelerated deterioration in cognition and neurodegeneration. CONCLUSIONS: CSF GAP-43 is increased in the predementia stage of AD, and it may enhance the neurodegenerative process. Future efforts on pharmacological interventions targeting synaptic dysfunction could be promising in AD treatment.

The Trajectory of Cerebrospinal Fluid Growth-Associated Protein 43 in the Alzheimer's Disease Continuum: A Longitudinal Study.

BACKGROUND: Synaptic degeneration has been suggested as an early pathological event that strongly correlates with severity of dementia in Alzheimer's disease (AD). However, changes in longitudinal cerebrospinal fluid (CSF) growth-associated protein 43 (GAP-43) as a synaptic biomarker in the AD continuum remain unclear. OBJECTIVE: To assess the trajectory of CSF GAP-43 with AD progression and its association with other AD hallmarks. METHODS: CSF GAP-43 was analyzed in 788 participants from the Alzheimer's Disease Neuroimaging Initiative (ADNI), including 246 cognitively normal (CN) individuals, 415 individuals with mild cognitive impairment (MCI), and 127 with AD dementia based on cognitive assessments. The associations between a multimodal classification scheme with amyloid-ß (Aß), tau, and neurodegeneration, and changes in CSF GAP-43 over time were also analyzed. RESULTS: CSF GAP-43 levels were increased at baseline in MCI and dementia patients, and increased significantly over time in the preclinical (Aß-positive CN), prodromal (Aß-positive MCI), and dementia (Aß-positive dementia) stages of AD. Higher levels of CSF GAP-43 were also associated with higher CSF phosphorylated tau (p-tau) and total tau (t-tau), cerebral amyloid deposition and hypometabolism on positron emission tomography, the hippocampus and middle temporal atrophy, and cognitive performance deterioration at baseline and follow-up. Furthermore, CSF GAP-43 may assist in effectively predicting the probability of dementia onset at 2- or 4-year follow-up. CONCLUSION: CSF GAP-43 can be used as a potential biomarker associated with synaptic degeneration in subjects with AD; it may also be useful for tracking the disease progression and for monitoring the effects of clinical trials.

Multi-cohort profiling reveals elevated CSF levels of brain-enriched proteins in Alzheimer's disease.

OBJECTIVE: Decreased amyloid beta (Aß) 42 together with increased tau and phospho-tau in cerebrospinal fluid (CSF) is indicative of Alzheimer's disease (AD). However, the molecular pathophysiology underlying the slowly progressive cognitive decline observed in AD is not fully understood and it is not known what other CSF biomarkers may be altered in early disease stages. METHODS: We utilized an antibody-based suspension bead array to analyze levels of 216 proteins in CSF from AD patients, patients with mild cognitive impairment (MCI), and controls from two independent cohorts collected within the AETIONOMY consortium. Two additional cohorts from Sweden were used for biological verification. RESULTS: Six proteins, amphiphysin (AMPH), aquaporin 4 (AQP4), cAMP-regulated phosphoprotein 21 (ARPP21), growth-associated protein 43 (GAP43), neurofilament medium polypeptide (NEFM), and synuclein beta (SNCB) were found at increased levels in CSF from AD patients compared with controls. Next, we used CSF levels of Aß42 and tau for the stratification of the MCI patients and observed increased levels of AMPH, AQP4, ARPP21, GAP43, and SNCB in the MCI subgroups with abnormal tau levels compared with controls. Further characterization revealed strong to moderate correlations between these five proteins and tau concentrations. INTERPRETATION: In conclusion, we report six extensively replicated candidate biomarkers with the potential to reflect disease development. Continued evaluation of these proteins will determine to what extent they can aid in the discrimination of MCI patients with and without an underlying AD etiology, and if they have the potential to contribute to a better understanding of the AD continuum.

Dissection of synaptic pathways through the CSF biomarkers for predicting Alzheimer disease.

OBJECTIVE: To assess the ability of a combination of synaptic CSF biomarkers to separate Alzheimer disease (AD) and non-AD disorders and to help in the differential diagnosis between neurocognitive diseases. METHODS: This was a retrospective cross-sectional monocentric study. All participants explored with CSF assessments for neurocognitive decline were invited to participate. After complete clinical and imaging evaluations, 243 patients were included. CSF synaptic (GAP-43, neurogranin, SNAP-25 total, SNAP-25aa40, synaptotagmin-1) and AD biomarkers were blindly quantified with ELISA or mass spectrometry. Statistical analysis compared CSF levels between the various groups of AD dementias (n = 81), mild cognitive impairment (MCI)-AD (n = 30), other MCI (n = 49), other dementias (OD) (n = 49), and neurologic controls (n = 35) and their discriminatory powers. RESULTS: All synaptic biomarkers were significantly increased in patients with MCI-AD and AD-dementia compared to the other groups. All synaptic biomarkers could efficiently discriminate AD dementias from OD (AUC ≥0.80). All but synaptotagmin were also able to discriminate patients with MCI-AD from controls (area under the curve [AUC] ≥0.85) and those with AD dementias from controls (AUC ≥0.80). Overall, CSF SNAP-25aa40 had the highest discriminative power (AUC 0.93 between patients with AD dementias and controls or OD, AUC 0.90 between those with MCI-AD and controls). Higher levels were associated with 2 alleles of APOE ε4. CONCLUSION: All synaptic biomarkers tested had a good discriminatory power to distinguish patients with AD abnormal CSF from those with non-AD disorders. SNAP25aa40 demonstrated the highest power to discriminate AD CSF-positive patients from patients without AD and neurologic controls in this cohort. CLASSIFICATION OF EVIDENCE: This retrospective study provides Class II evidence that CSF synaptic biomarkers discriminate patients with AD from those without AD.

Cerebrospinal fluid growth-associated protein 43 in multiple sclerosis.

Neurodegeneration in multiple sclerosis (MS) correlates with disease progression and reparative processes may be triggered. Growth-associated protein 43 (GAP-43) exhibits induced expression during axonal growth and reduced expression during MS progression. We aimed to evaluate if GAP-43 can serve as a biomarker of regeneration in relapsing-remitting MS (RRMS) and whether disease-modifying therapies (DMTs) influence GAP-43 concentration in cerebrospinal fluid (CSF). GAP-43 was measured using an enzyme-linked immunosorbent assay in 105 MS patients (73 RRMS, 12 primary progressive MS, 20 secondary progressive MS) and 23 healthy controls (HCs). In 35 of the patients, lumbar puncture, clinical assessment, and magnetic resonance imaging was performed before initiation of therapeutic intervention, and at follow-up. CSF GAP-43 concentration was significantly lower in progressive MS compared with HCs (p = 0.004) and RRMS (p = < 0.001) and correlated negatively with disability (p = 0.026). However, DMTs did not alter CSF GAP-43. Interestingly, in RRMS CSF GAP-43 levels were higher in patients with signs of active inflammatory disease than in patients in remission (p = 0.042). According to CSF GAP-43 concentrations, regeneration seems reduced in progressive MS, increased during disease activity in RRMS but is unaffected by treatment of highly active DMTs.

Elevated CSF GAP-43 is Alzheimer's disease specific and associated with tau and amyloid pathology.

INTRODUCTION: The level of the presynaptic protein growth-associated protein 43 (GAP-43) in cerebrospinal fluid (CSF) has previously been shown to be increased in Alzheimer's disease (AD) and thus may serve as an outcome measure in clinical trials and facilitate earlier disease detection. METHODS: We developed an enzyme-linked immunosorbent assay for CSF GAP-43 and measured healthy controls (n = 43), patients with AD (n = 275), or patients with other neurodegenerative diseases (n = 344). In a subpopulation (n = 93), CSF GAP-43 concentrations from neuropathologically confirmed cases were related to Aß plaques, tau, α-synuclein, and TDP-43 pathologies. RESULTS: GAP-43 was significantly increased in AD compared to controls and most neurodegenerative diseases and correlated with the magnitude of neurofibrillary tangles and Aß plaques in the hippocampus, amygdala, and cortex. GAP-43 was not associated to α-synuclein or TDP-43 pathology. DISCUSSION: The presynaptic marker GAP-43 is associated with both diagnosis and neuropathology of AD and thus may be useful as a sensitive and specific biomarker for clinical research.

Transient increase in CSF GAP-43 concentration after ischemic stroke.

BACKGROUND: Cerebrospinal fluid (CSF) biomarkers reflect ongoing processes in the brain. Growth-associated protein 43 (GAP-43) is highly upregulated in brain tissue shortly after experimental ischemia suggesting the CSF GAP-43 concentration may be altered in ischemic brain disorders. CSF GAP-43 concentration is elevated in Alzheimer's disease patients; however, patients suffering from stroke have not been studied previously. METHODS: The concentration of GAP-43 was measured in longitudinal CSF samples from 28 stroke patients prospectively collected on days 0-1, 2-4, 7-9, 3 weeks, and 3-5 months after ischemia and cross-sectionally in 19 controls. The stroke patients were clinically evaluated using a stroke severity score system. The extent of the brain lesion, including injury size and degrees of white matter lesions and atrophy were evaluated by CT and magnetic resonance imaging. RESULTS: Increased GAP-43 concentration was detected from day 7-9 to 3 weeks after stroke, compared to day 1-4 and to levels in the control group (P = 0.02 and P = 0.007). At 3-5 months after stroke GAP-43 returned to admission levels. The initial increase in GAP-43 during the nine first days was associated to stroke severity, the degree of white matter lesions and atrophy and correlated positively with infarct size (rs = 0.65, P = 0.001). CONCLUSIONS: The transient increase of CSF GAP-43 is important to take into account when used as a biomarker for other neurodegenerative diseases such as Alzheimer's disease. Furthermore, GAP-43 may be a marker of neuronal responses after stroke and additional studies confirming the potential of CSF GAP-43 to reflect severity and outcome of stroke in larger cohorts are warranted.

CSF YKL-40 and GAP-43 are related to suicidal ideation in older women.

OBJECTIVE: To investigate possible relationships between suicidal ideation and cerebrospinal fluid (CSF) levels of glial markers YKL-40 (also known as chitinase-3-like protein 1), growth-associated protein-43 (GAP-43) and myelin basic protein (MBP). METHOD: The sample was obtained from the Prospective Population Study of Women and included 86 women without dementia who underwent both psychiatric examinations and lumbar puncture (LP). Eight of these women reported past-month suicidal ideation. RESULTS: Significantly, higher CSF levels of both YKL-40 and GAP-43 were detected in women with past-month suicidal ideation. Associations with suicidal ideation remained for both YKL-40 and GAP-43 in regression models adjusted for smoking status, BMI and age. CSF levels of YKL-40, GAP-43 and MBP did not differ by depression status. Higher levels of CSF GAP-43 were associated with feelings of worthlessness; a strong relationship was demonstrated in the fully adjusted model (OR 5.95 CI [1.52-23.20], P = 0.01). CONCLUSION: Our findings of elevated CSF concentrations of both YKL-40 and GAP-43 in women with suicidal ideation, compared to those without, suggest that a disrupted synaptic glial functioning and inflammation may be related to the aetiology of suicidal ideation in older adults.

CSF profiling of the human brain enriched proteome reveals associations of neuromodulin and neurogranin to Alzheimer's disease.

PURPOSE: This study is part of a larger effort aiming to expand the knowledge of brain-enriched proteins in human cerebrospinal fluid (CSF) and to provide novel insight into the relation between such proteins and different neurodegenerative diseases. EXPERIMENTAL DESIGN: Here 280 brain-enriched proteins in CSF from patients with Alzheimer's disease (AD), Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are profiled. In total, 441 human samples of ventricular CSF collected post mortem and lumbar CSF collected ante mortem are analyzed using 376 antibodies in a suspension bead array setup, utilizing a direct labelling approach. RESULTS: Among several proteins displaying differentiated profiles between sample groups, we focus here on two synaptic proteins, neuromodulin (GAP43) and neurogranin (NRGN). They are both found at elevated levels in CSF from AD patients in two independent cohorts, providing disease-associated profiles in addition to verifying and strengthening previously observed patterns. Increased levels are also observed for patients for whom the AD diagnosis was not established at the time of sampling. CONCLUSIONS AND CLINICAL RELEVANCE: These findings indicate that analyzing the brain-enriched proteins in CSF is of particular interest to increase the understanding of the CSF proteome and its relation to neurodegenerative disorders. In addition, this study lends support to the notion that measurements of these synaptic proteins could potentially be of great relevance in future diagnostic tests for AD.

Antibody-based profiling of cerebrospinal fluid within multiple sclerosis.

Antibody suspension bead arrays have proven to enable multiplexed and high-throughput protein profiling in unfractionated plasma and serum samples through a direct labeling approach. We here describe the development and application of an assay for protein profiling of cerebrospinal fluid (CSF). While setting up the assay, systematic intensity differences between sample groups were observed that reflected inherent sample specific total protein amounts. Supplementing the labeling reaction with BSA and IgG diminished these differences without impairing the apparent sensitivity of the assay. We also assessed the effects of heat treatment on the analysis of CSF proteins and applied the assay to profile 43 selected proteins by 101 antibodies in 339 CSF samples from a multiple sclerosis (MS) cohort. Two proteins, GAP43 and SERPINA3 were found to have a discriminating potential with altered intensity levels between sample groups. GAP43 was detected at significantly lower levels in secondary progressive MS compared to early stages of MS and the control group of other neurological diseases. SERPINA3 instead was detected at higher levels in all MS patients compared to controls. The developed assay procedure now offers new possibilities for broad-scale protein profiling of CSF within neurological disorders.

Treatment of Alzheimer's disease with clioquinol.

As heavy metal ions may be implicated in the formation of senile plaques in Alzheimer-afflicted brains, treatment with clioquinol was tested in 20 patients with Alzheimer's disease. Clioquinol is a chelator that crosses the blood-brain barrier and has greater affinity for zinc and copper ions than for calcium and magnesium ions. Treatment was given for 21 days at doses of 20 mg/day to 10 patients and 80 mg/day to another 10 patients. The study was blind to the dosages but included no controls. Cerebrospinal fluid (CSF) investigations revealed a significant increase at day 7 and a decrease at day 21 in Tau protein and growth-associated protein (GAP43). These proteins are increased in Alzheimer's disease and considered as rather stable markers. The initial increase may indicate a temporary cytotoxicity to the brain and/or an increased release into the CSF from stores in the tissue, possibly from senile plaques where the proteins are accumulated. The levels of CSF-Tau protein correlated positively and significantly with the serum levels of copper and also with the serum copper/zinc ratio. Clinical ratings showed slight improvement after 3 weeks treatment with clioquinol in this open study.

The cerebrospinal fluid levels of tau, growth-associated protein-43 and soluble amyloid precursor protein correlate in Alzheimer's disease, reflecting a common pathophysiological process.

Cerebrospinal fluid (CSF) levels of tau (total tau), growth-associated protein-43 (GAP-43), soluble amyloid precursor protein (sAPP; i.e. total sAPP), and beta-amyloid(42) (Abeta(42)) were studied in patients with frontotemporal dementia (FTD; n = 14), Alzheimer's disease (AD; n = 47) and vascular dementia (VAD; n = 16), and in age-matched controls (n = 12). CSF-tau was increased in AD compared to controls and FTD (p < 0.001 for both). CSF-GAP-43 was increased in AD compared to controls (p < 0.05), and both CSF-GAP-43 and CSF-sAPP were increased in AD compared to FTD (p < 0.01). Positive and highly significant correlations were found between CSF-tau and CSF-GAP-43 in all groups and between CSF-tau, CSF-GAP-43 and CSF-sAPP in AD. The correlations found may reflect a common pathophysiologic process such as axonal degeneration.

CSF levels of tau, beta-amyloid(1-42) and GAP-43 in frontotemporal dementia, other types of dementia and normal aging.

Cerebrospinal fluid (CSF) levels of tau, beta-amyloid(1-42) and growth-associated protein 43 (GAP-43) were studied in patients with frontotemporal dementia (FTD; n = 17), Alzheimer's disease (AD; n = 60), subcortical white-matter dementia (SWD; n = 24), Parkinson's disease (PD; n = 23) and dysthymia (n = 19) and in age-matched controls (n = 32). CSF-tau was significantly increased only in AD, and CSF-beta-amyloid(1-42) was significantly decreased in AD and SWD as compared to controls, and in AD compared to FTD. CSF-GAP-43 was significantly decreased only in PD. The GAP-43/tau ratio was decreased in all the patient groups except the dysthymia group compared to controls. A positive correlation was found between CSF-GAP-43 and CSF-tau in all groups. The results suggest normal levels of CSF-tau and CSF-beta-amyloid(1-42) in FTD, which will aid in the clinical separation of FTD from AD. In SWD, decreased levels of CSF-beta-amyloid(1-42) suggest concomitant involvement of vascular and amyloid protein mechanisms.

Identification of synaptic vesicle, pre- and postsynaptic proteins in human cerebrospinal fluid using liquid-phase isoelectric focusing.

Synaptic pathology is central in the pathogenesis of several psychiatric disorders, for example in Alzheimer's disease (AD) and schizophrenia. Quantification of specific synaptic proteins has proved to be a useful method to estimate synapitc density in the brain. Using this approach, several synaptic proteins have been demonstrated to be altered in both AD and schizophrenia. Until recently, the analysis of synaptic pathology has been limited to postmortem tissue. In living subjects, these synaptic proteins may be studied through analysis of cerebrospinal fluid (CSF). In an earlier study performed by us, one synaptic vesicle specific protein, synaptotagmin, was detected in CSF for the first time using a procedure based on affinity chromatography, reversed-phase chromatography, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and chemiluminescence immunoblotting. However, other synaptic proteins were not detectable with this procedure. Therefore, we have developed a procedure including precipitation of CSF proteins with trichloroacetic acid, followed by liquid-phase isoelectric focusing using the Rotofor Cell, and finally analysis of Rotofor fractions by Western blotting for identification of synaptic proteins in CSF. Five synaptic proteins, rab3a, synaptotagmin, growth-associated protein (GAP-43), synaptosomal-associated protein (SNAP-25) and neurogranin, have been demonstrated in CSF using this method. The major advantage of liquid-phase isoelectric focusing (IEF) using the Rotofor cell is that it provides synaptic proteins from CSF in sufficient quantities for identification. This method may also be suitable for identification of other types of trace amounts of brain-specific proteins in CSF. These results demonstrate that several synaptic proteins can be identified and measured in CSF to study synaptic function and pathology in degenerative disorders.