Lower levels of cerebrospinal fluid amyloid beta (Abeta) in non-demented Indian controls

Prevalence of Alzheimer's disease in Indian population is lower than in developed countries. To determine whether limitation of amyloid beta (Abeta) concentration may be responsible for lower rate of incidence, we measured the levels of Abeta in cerebrospinal fluid (CSF) collected from 72 non-demented individuals ranging in the age from 20 years to 65 years. These samples were segregated into three groups ranging from 20-35 years, 36-50 years and 51-65 years of age. Levels of Abeta could be detected in all the age groups and they were much lower than the values reported in literature from the developed countries. No significant difference in the average level of Ass was observed with increase in age.     

Zinc and copper modulate Alzheimer Abeta levels in human cerebrospinal fluid

Abnormal interaction of beta-amyloid 42 (Abeta42) with copper, zinc and iron induce peptide aggregation and oxidation in Alzheimer's disease (AD). However, in health, Abeta degradation is mediated by extracellular metalloproteinases, neprilysin, insulin degrading enzyme (IDE) and matrix metalloproteinases. We investigated the relationship between levels of Abeta and biological metals in CSF. We assayed CSF copper, zinc, other metals and Abeta42 in ventricular autopsy samples of Japanese American men (N=131) from the population-based Honolulu Asia Aging Study. There was a significant inverse correlation of CSF Abeta42 with copper, zinc, iron, manganese and chromium. The association was particularly strong in the subgroup with high levels of both zinc and copper. Selenium and aluminum levels were not associated to CSF Abeta42. In vitro, the degradation of synthetic Abeta substrate added to CSF was markedly accelerated by low levels (2microM) of exogenous zinc and copper. While excessive interaction with copper and zinc may induce neocortical Abeta precipitation in AD, soluble Abeta degradation is normally promoted by physiological copper and zinc concentrations.     

Neurochemical diagnosis of Alzheimer's dementia by CSF Abeta42, Abeta42/Abeta40 ratio and total tau

Cerebrospinal fluid (CSF) concentrations of amyloid beta peptides ending at positions 42 and 40 (Abeta42 and Abeta40, respectively), and total tau (tTau) protein were measured by ELISA in order to compare their accuracy in discriminating patients with Alzheimer's disease (AD, n = 22), non-Alzheimer dementia (nAD, n = 11) and control subjects (CON, n=35). As compared to the other groups, the concentrations of Abeta42 and tTau were decreased (P<0.001) and increased (P<0.001) in AD, respectively, while Abeta40 did not differ significantly among the groups. Receiver operating characteristic (ROC) analysis was performed to define cut-off values for maximized sensitivity and specificity. For all groups compared the Abeta peptide ratio 42/40 classified more patients correctly, as compared to the concentration of Abeta42 alone: AD versus controls, 94 and 86.7%; AD versus nAD, 90 and 85% and AD versus nAD plus controls, 90.8 and 87%, respectively. The percentage of correctly classified patients was further improved when the Abeta ratio was combined with the analysis of the tTau concentration. Presence of the apolipoprotein E 4 allele, age or degree of mental disability did not significantly influence the parameters studied.     

Evaluation of plasma Abeta(40) and Abeta(42) as predictors of conversion to Alzheimer's disease in patients with mild cognitive impairment

Numerous studies have shown a marked decrease of beta-amyloid(42) (Abeta(42)) in the cerebrospinal fluid (CSF) of patients with incipient Alzheimer's disease (AD). However, studies on Abeta in plasma are contradictory, and show very marginal differences between patients and controls. Here, we analyzed plasma samples using a new multiplex immunoassay for simultaneous analysis of Abeta(1-40), Abeta(n-40), Abeta(1-42), and Abeta(n-42). The plasma samples were obtained at baseline from two independent cohorts of patients with mild cognitive impairment (MCI) and age-matched controls. In the first cohort, 41% of the 117 MCI cases converted to AD during a clinical follow-up period of 4-7 years. In the second cohort, 14% of the 110 MCI subjects developed AD during a clinical follow-up period of 2-4 years. None of the plasma Abeta isoforms differed between MCI patients that subsequently developed AD and healthy controls or stable MCI patients. The Cox proportional hazards model did not reveal any differences in the probability of progression from MCI to AD related to plasma Abeta levels. In contrast, low levels of Abeta(1-42) in CSF were strongly associated with increased risk of future AD. The absence of a change in plasma Abeta in incipient AD, despite the marked change in CSF, may be explained by the lack of a correlation between the levels of Abeta(1-42) in CSF and plasma. In conclusion, the results show that CSF biomarkers are better predictors of progression to AD than plasma Abeta isoforms.     

Choline levels are increased in cerebrospinal fluid of Alzheimer patients

We measured choline (Ch), acetylcholinesterase activity (AChE) and total protein in the cerebrospinal fluid (CSF) of 66 Alzheimer patients (ages 54-89 years) and 22 age-matched controls (ages 52-80 years), looking for markers of the well-established cholinergic deficit and neuronal degeneration in Alzheimer disease (AD). Three or more lumbar punctures were performed in 21 patients over a span of 24 months in order to study the changes in these CSF components with disease progression. We found a statistically significant reduction in AChE and an increase in Ch with advancing dementia. These changes were not related to patient age. We suggest that the rise in CSF choline is related to neuronal membrane breakdown and reduced Ch uptake by cholinergic neurons. The reduction in CSF AChE is consistent with the depletion of cholinergic neurons in AD.     

Three-year follow-up of cerebrospinal fluid tau, beta-amyloid 42 and 40 concentrations in Alzheimer's disease

Earlier studies have shown elevated levels of tau protein and decreased levels of amyloid beta42 in cerebrospinal fluid (CSF) from patients with Alzheimer's disease (AD). We investigated the concentrations of Abeta42, Abeta40 and tau in CSF from AD patients on the baseline and after follow-up period of 3 years using ELISA assays. There was a significant decrease of Abeta42 (P<0.05) and Abeta40 (P<0.05) levels with time. AD patients with the duration of the disease 2 years or less at baseline had more pronounced decrease of Abeta42 concentrations compared to those with the duration of the disease more than 2 years at baseline (P<0.05). CSF tau protein concentrations increased in 9/17 but decreased in 8/17 patients. These results suggest that Abeta42 and Abeta40 may be useful in monitoring the long-term progression of AD particularly in the early stages of the disease.     

Amyloid beta protein 1-40 and 1-42 levels in matched cerebrospinal fluid and plasma from patients with Alzheimer disease

We quantitated amyloid beta proteins 1-40 (Abeta40) and 1-42 (Abeta42), and alpha1- antichymotrypsin (ACT) in matched cerebrospinal fluid (CSF) and plasma of 50 patients with probable Alzheimer disease, and analyzed the relationships with age, sex, Mini-Mental State Examination (MMSE), and apolipoprotein E phenotype. There was no relation between CSF Abeta40 and Abeta42 levels with those of plasma. CSF and plasma Abeta40 and Abeta42 levels showed no association with age, sex, and MMSE score. There was a significant correlation between CSF ACT and plasma ACT levels. The data suggest that plasma ACT crosses the blood-brain barrier. However, a lack of correlation between CSF Abeta40 and Abeta42 levels with those of plasma suggests that Abeta in CSF and plasma originates from different sources.     

Matrix metalloproteinases-2 and -3 are reduced in cerebrospinal fluid with low beta-amyloid1–42 levels

Matrix metalloproteinases (MMPs), a family of extracellular soluble or membrane bound endopeptidases, are implicated in many physiological and pathophysiological functions—based on their capability to cleave all protein components of the extracellular matrix. Recent studies have implicated several forms of MMPs in chronic neurodegenerative diseases like Alzheimer's disease (AD), vascular dementia (VD), and Parkinson's disease (PD). The aim of the present study was to analyse eight MMPs (MMP-1, -2, -3, -7, -8, -9, -10, -13) in the human cerebrospinal fluid (CSF) and to correlate with the well established biomarkers beta-amyloid_1–42 (Aβ), total-tau and phospho-tau-181. Our data show a significant decrease of MMP-2 and MMP-3 levels in the CSF in samples with significantly reduced Aβ levels. It is concluded that MMP-2 and MMP-3 are directly linked to Aβ in the brain and a dysfunction may influence the processing of Aβ.     

FDP levels in the cerebrospinal fluid are elevated in patients with meningitis

Since the level of fibrinogen degradation products (FDP) are elevated with severity of inflammation, we assumed that FDP in the cerebrospinal fluid (CSF) could be a marker of meningitis. We, therefore, measured FDP in the CSF of 6 patients with bacterial meningitis and 6 aseptic meningitis. The range of FDP levels in the CSF in patients without meningitis was 0.21 +/- 0.01 microgram/ml. While, the level of FDP in patients with aseptic meningitis (0.43 +/- 0.10 microgram/ml) and in bacterial meningitis (1.78 +/- 0.42 micrograms/ml) was significantly elevated (p less than 0.05). The value was significantly (p less than 0.01) higher in the group of septic meningitis than in aseptic meningitis. In one patient with septic meningitis, we could measure FDP in the CSF several times during the course of the disease, in which the level of FDP got into the high range earlier than the changes in levels of protein, glucose and cell counts in the CSF. FDP in the CSF well correlated to the clinical course of the meningitis. Eventually, we found that FDP in the CSF was definitely elevated in patients with bacterial meningitis, whereas it was slightly elevated in patients with aseptic meningitis. The measurement of FDP in the CSF, therefore, is concluded to be useful for the differential diagnosis of meningitis, and to assess the clinical course of meningitis.     

Selective binding of soluble Abeta1-40 and Abeta1-42 to a subset of senile plaques.

Alzheimer's disease is characterized by the progressive accumulation of amyloid-beta protein (Abeta) in senile plaques and cerebral amyloid angiopathy. It is not known whether the plaque growth is a continuous and homogeneous process or whether some plaques have a more rapid evolution. As plaques grow by the deposition of Abeta, we used an in situ binding technique to analyze the deposition of fluorescein-conjugated and biotinylated Abeta1 40 and Abeta1-42 in cryosections of brains from Alzheimer's disease patients. Only a subset of senile plaques but all cerebrovascular Abeta deposits were labeled by both Abeta1-40 and Abeta1-42. Striking differences in binding were observed among adjacent plaques. Quantitative analysis showed that on average 60% of all plaques were labeled with Abeta1-42 and 31% of all plaques were labeled with Abeta1-40 (n=7; P<0.001). Confocal laser scanning microscopy of double-labeled sections revealed that the newly deposited Abeta was only partially co-localized to pre-existing Abeta and apolipoprotein E and was not co-localized to heparan sulfate proteoglycan. Abeta binding was preserved after glycolytic pretreatment with periodic acid. Our results suggest that at a given time point only a subset of active senile plaques accumulate A(beta) and that plaque growth may be conditioned by the presence of other distinct plaque components different from Abeta, apolipoprotein E or heparan sulfate proteoglycan.     

Mean age-of-onset of familial alzheimer disease caused by presenilin mutations correlates with both increased Abeta42 and decreased Abeta40

The varied ways in which mutations in presenilins (PSEN1 and PSEN2) affect amyloid b precursor protein (APP) processing in causing early-onset familial Alzheimer disease (FAD) are complex and not yet properly understood. Nonetheless, one useful diagnostic marker is an increased ratio of Ab42 to Ab40 (Ab42/Ab40) in patients' brain and biological fluids as well as in transgenic mice and cells. We studied Ab and APP processing for a set of nine clinical PSEN mutations on a novel and highly reproducible enzyme-linked immunosorbent assay (ELISA)-based in vitro method and also sought correlation with brain Ab analyzed by image densitometry and mass spectrometry. All mutations significantly increased Ab42/Ab40 in vitro by significantly decreasing Ab40 with accumulation of APP C-terminal fragments, a sign of decreased PSEN activity. A significant increase in absolute levels of Ab42 was observed for only half of the mutations tested. We also showed that age-of-onset of PSEN1-linked FAD correlated inversely with Ab42/Ab40 (r = -0.89; P = 0.001) and absolute levels of Ab42 (r = -0.83; P = 0.006), but directly with Ab40 levels (r = 0.69; P = 0.035). These changes also partly correlated with brain Ab42 and Ab40 levels. Together, our data suggested that Ab40 might be protective by perhaps sequestering the more toxic Ab42 and facilitating its clearance. Also, the in vitro method we describe here is a valid tool for assaying the pathogenic potential of clinical PSEN mutations in a molecular diagnostic setting.     

Astrocytes containing amyloid beta-protein (Abeta)-positive granules are associated with Abeta40-positive diffuse plaques in the aged human brain.

Amyloid beta-protein (Abeta) is the major component of senile plaques that emerge in the cortex during aging and appear most abundantly in Alzheimer's disease. In the course of our immunocytochemical study on a large number of autopsy cases, we noticed, in many aged nondemented cases, the presence of unique diffuse plaques in the cortex distinct from ordinary diffuse plaques by immunocytochemistry. The former were amorphous, very faintly Abeta-immunoreactive plaques resembling diffuse plaques, but they stained for Abeta40 and were associated with small cells containing Abeta-positive granules. A panel of amino- and carboxyl-terminal-specific Abeta antibodies showed that such Abeta40-positive diffuse plaques and cell-associated granules were composed exclusively of amino-terminally deleted Abeta terminating at Abeta40, -42, and -43. Double immunostaining also showed that those Abeta-immunoreactive granules are located in astrocytes and not in microglia or neurons. Immunoelectron microscopy revealed that nonfibrillar Abeta immunoreactivity was located within lipofuscin-like granules in somewhat swollen astrocytes. These findings raise the possibility that astrocytes take up Abeta and attempt to degrade it in lysosomes in the aged brain.     

Kynurenic acid levels are elevated in the cerebrospinal fluid of patients with schizophrenia.

Kynurenic acid is an endogenous glutamate antagonist with a preferential action at the glycine-site of the N-methyl D-aspartate-receptor. Mounting evidence indicate that the compound is significantly involved in basal neurophysiological processes in the brain. In the present investigation, cerebrospinal fluid (CSF) level of kynurenic acid was analyzed in 28 male schizophrenic patients and 17 male healthy controls by means of high pressure liquid chromatography and fluorescence detection. Schizophrenic patients showed elevated CSF levels of kynurenic acid (1.67+/-0.27 nM) compared to the control group (0.97+/-0.07 nM). Furthermore, CSF levels of kynurenic acid in schizophrenic patients were also found to correlate with age. The present finding is indicative of a contribution of kynurenic acid in the pathogenesis of schizophrenia.     

Full-length amyloid-beta (1-42(43)) and amino-terminally modified and truncated amyloid-beta 42(43) deposit in diffuse plaques.

The amino- and carboxyl-terminal properties of the amyloid-beta (A beta) peptides deposited in diffuse plaques, one of the earliest forms of A beta deposition, were examined in the brains of patients with Down's syndrome and Alzheimer's disease and in aged individuals without dementia by immunocytochemistry. This was done using a panel of antibodies that specifically discriminate the terminal structures and modifications at the amino and carboxyl termini of A beta. Diffuse plaques found in the cerebral and cerebellar cortex, neostriatum, and hypothalamus of Down's syndrome, Alzheimer's disease, and nondemented brains were strongly immunoreactive for A beta N1(L-Asp), A beta N1(L-isoAsp), A beta N1(D-Asp), and A beta N3(pyroGlu) and weakly positive for A beta N11(pyroGlu) and A beta N17(Leu). Diffuse plaques also were positive for A beta 42(43) but negative for A beta 40, using carboxyl-terminal-specific anti-A beta antibodies. These results suggest that the amino termini of the A beta species that initially deposit in diffuse plaques begin with A beta N1(Asp) with or without structural modifications (isomerization and racemization), as well as with A beta N3(pyroGlu), and terminate preferentially at A beta 42(43) rather than A beta 40.     

Uptake of Abeta 1-40- and Abeta 1-42-coated yeast by microglial cells: a role for LRP

Artificial diffuse and amyloid core of neuritic plaques [beta-amyloid peptide (Abeta) deposits] could be prepared using heat-killed yeast particles opsonized with Abeta 1-40 or Abeta 1-42 peptides. Interaction and fate of these artificial deposits with microglial cells could be followed using a method of staining that allows discrimination of adherent and internalized, heat-killed yeast particles. Using this system, it was possible to show that nonfibrillar or fibrillar (f)Abeta peptides, formed in solution upon heating (aggregates), could not impair the internalization of heat-killed yeast particles opsonized with fAbeta 1-40 or fAbeta 1-42. This indicated that depending on their physical state, Abeta peptide(s) do not recognize the same receptors and probably do not follow the same internalization pathway. Using competitive ligands of class A scavenger receptors (SR-A) or low-density lipoprotein-related receptor protein (LRP), it has been shown that SR-A were not involved in the recognition of amyloid peptide deposits, whereas LRP specifically recognized deposits of fAbeta 1-42 (but not fAbeta 1-40) and mediated their phagocytosis.     

Genetic variants in a haplotype block spanning IDE are significantly associated with plasma Abeta42 levels and risk for Alzheimer disease

Risk for late onset Alzheimer disease (LOAD) and plasma amyloid beta levels (Abeta42; encoded by APP), an intermediate phenotype for LOAD, show linkage to chromosome 10q. Several strong candidate genes (VR22, PLAU, IDE) lie within the 1-lod support interval for linkage. Others have independently identified haplotypes in the chromosome 10q region harboring IDE that show highly significant association with intermediate AD phenotypes and with risk for AD. To pursue these associations, we analyzed the same haplotypes for association with plasma Abeta42 in 24 extended LOAD families and for association with LOAD in two independent case-control series. One series (MCR, 188 age-matched case-control pairs) did not show association (p=0.64) with the six haplotypes in the 276-kb region spanning three genes (IDE, KNSL1, and HHEX) previously shown to associate with LOAD. The other series (MCJ, 109 age-matched case-control pairs) showed significant (p=0.003) association with these haplotypes. In the MCJ series, the H4 (odds ratio [OR]=5.1, p=0.003) and H2(H7) haplotypes (OR=0.60, p=0.04) had the same effects previously reported. In this series, the H8 haplotype (OR=2.7, p=0.098) also had an effect similar as in one previous case control series but not in others. In the extended families, the H8 haplotype was associated with significantly elevated plasma Abeta42 (p=0.02). In addition, the H5(H10) haplotype, which is associated with reduced risk for AD in the other study is associated with reduced plasma Abeta42 (p=0.007) in our family series. These results provide strong evidence for pathogenic variant(s) in the 276-kb region harboring IDE that influence intermediate AD phenotypes and risk for AD.     

Reduced levels of nitric oxide metabolites in cerebrospinal fluid are associated with equine protozoal myeloencephalitis

Equine protozoal myeloencephalitis (EPM) is a disease of horses that is primarily associated with infection with the apicomplexan Sarcocystis neurona. Infection with this parasite alone is not sufficient to induce the disease, and the mechanism of neuropathogenesis associated with EPM has not been reported. Nitric oxide (NO) functions as a neurotransmitter, a vasodilator, and an immune effector and is produced in response to several parasitic protozoa. The purpose of this work was to determine if the concentration of NO metabolites (NO(x)(-)) in the cerebrospinal fluid (CSF) is correlated with the development of EPM. CSF NO(x)(-) levels were measured before and after transport-stressed, acclimated, or dexamethasone-treated horses (n = 3 per group) were experimentally infected with S. neurona sporocysts. CSF NO(x)(-) levels were also compared between horses that were diagnosed with EPM after natural infection with S. neurona and horses that did not have clinical signs of disease or that showed no evidence of infection with the parasite (n = 105). Among the experimentally infected animals, the mean CSF NO(x)(-) levels of the transport-stressed group, which had the most severe clinical signs, was reduced after infection, while these values were found to increase after infection in the remaining groups that had less severe signs of EPM. Under natural conditions, horses with EPM (n = 65) had a lower mean CSF NO(x)(-) concentration than clinically normal horses with antibodies (Abs) against S. neurona (n = 15) in CSF, and horses that developed ataxia (n = 81) had a significantly lower mean CSF NO(x)(-) concentration than horses that did not have neurologic signs (n = 24). In conclusion, lower CSF NO(x)(-) levels were associated with clinical EPM, suggesting that measurement of CSF NO(x)(-) levels could improve the accuracy of diagnostic tests that are based upon detection of S. neurona-specific Abs in CSF alone and that reduced NO levels could be causally related to the development of EPM.     

Fibroblast growth factor-2 levels are elevated in the cerebrospinal fluid of multiple sclerosis patients

In recent years a role has been recognized for fibroblast growth factor (FGF)-2 in the pathogenesis of demyelination and the failure of remyelination in experimental models of multiple sclerosis (MS). FGF-2 levels were determined using a sensitive immunoassay in the cerebrospinal fluid (CSF) of 20 patients with clinically isolated syndrome (CIS), 40 patients with relapsing-remitting (R-R) MS, and 30 patients with secondary progressive (SP) MS, correlated with MRI measures. Control CSF samples were obtained from 20 subjects who underwent lumbar puncture for diagnostic purposes and for whom all instrumental and laboratory analyses excluded systemic and nervous system diseases. FGF-2 levels in the CSF of MS and CIS patients were significantly higher than controls (P<0.001 and P<0.05, respectively). The highest levels were detected in R-R MS patients during relapse and in SP MS patients with an increase of 1 point in EDSS scores in the last 6 months. A significant correlation was found in SP MS patients with lesional load (R=0.43, P<0.01) but not with parenchymal fractions as measures of brain atrophy. A slight increase in serum FGF-2 levels was also found in R-R MS patients during relapse with gadolinium enhancing lesions and in SP patients with disability progression. These findings support the implication of FGF-2 in the pathogenesis of MS and concur with recent reports of the involvement of FGF receptor signalling in the disruption of myelin production in differentiated oligodendrocytes and in the loss of adult oligodendrocytes and myelin in vivo due to FGF-2.     

Reduced cerebrospinal fluid estradiol levels are associated with increased beta-amyloid levels in female patients with Alzheimer's disease.

Recent in-vitro studies indicate that estrogens such as 17beta-estradiol (E2) may decrease the production of beta-amyloid 1-42 (Abeta42), a peptide central for the formation of senile plaques in Alzheimer's disease (AD). To test this hypothesis in a clinical study, cerebrospinal fluid levels of E2 were compared between 30 female AD patients and 11 female patients with non-dementing diseases such as major depression and investigated with respect to beta-amyloid 1-40 and Abeta42 levels. E2 levels were significantly (P<0.05) lower in the AD group than in controls; within the AD group E2 levels were inversely correlated with Abeta42 concentrations (r=-0.36, P=0.05). This is the first clinical study providing evidence for an influence of E2 on Abeta42 metabolism in vivo. This observation corresponds to the putative beneficial effects of estrogen replacement therapy on the development and course of AD.