A population-based study of tau protein and ubiquitin in cerebrospinal fluid in 85-year-olds: relation to severity of dementia and cerebral atrophy, but not to the apolipoprotein E4 allele.

Alzheimer's disease (AD) is the most common form of dementia, and is characterized by a degeneration of neurones and their synapses, and a higher number of senile plaques (SP) and neurofibrillary tangles (NFT) compared with that found in non-demented individuals of the same age. NFT are composed of a hyperphosphorylated and ubiquitinated form of tau protein. Previous studies have found that in the cerebrospinal fluid (CSF) both tau and ubiquitin are increased in AD. We examined CSF-tau and CSF-ubiquitin in a population based sample of 85-year-olds, 26 demented (11 with probable Alzheimer's disease (AD), 13 with probable vascular dementia (VAD) and 2 with mixed (AD/VAD) type of dementia) and 35 non-demented individuals. CSF-tau was significantly higher both in the probable AD group (254 +/- 113 pg/mL; P < 0.01), and in the probable VAD group (247 +/- 75 pg/mL; P < 0.005), than in the non-demented group (171 +/- 78 pg/mL), but did not significantly differ between the probable AD and probable VAD groups. In contrast, CSF-ubiquitin did not significantly differ between the probable AD (100 +/- 24 ng/mL), probable VAD (102 +/- 16 ng/mL), and non-demented (97 +/- 27 ng/mL) groups. CSF-tau increased with increasing severity of dementia (P < 0.001), though no such relation was found for CSF-ubiquitin. Neither CSF-tau nor CSF-ubiquitin differed between patients with or without the apolipoprotein E E4 isoform. Higher CSF-tau and CSF-ubiquitin levels were also associated with increasing degree of cortical and central brain atrophy as measured by computerized tomography. The relationships between CSF-tau and severity of dementia and to brain atrophy suggest that CSF-tau may be used as a measure of neuronal/axonal degeneration in patients with dementia. We have previously shown a marked increase in both CSF-tau and CSF-ubiquitin in younger patients with AD and VAD. The less pronounced increase in CSF-tau and the lack of difference in CSF-ubiquitin in older patients suggest that the severity of the degenerative process is less in older than in younger demented patients.

Ubiquitin in cerebrospinal fluid: a rapid competitive enzyme-linked immunoflow assay.

The aims of this study were to develop a rapid immunoassay to determine the levels of ubiquitin in cerebrospinal fluid and to establish the ubiquitin levels in the spinal fluid of normal aged individuals. A competitive enzyme-linked immunoflow assay was developed. In this assay, ubiquitin is bound to nitrocellulose membrane, after which the primary antibody-test sample mixture and the enzyme-labeled secondary antibody under vacuum are applied sequentially. The final reaction product is collected in a microtiter plate by suction. This competitive assay requires only approximately 4 h and is potentially useful for determining in biological fluids the levels of any antigen or its antibodies that might be present. Employing this immunoassay, cerebrospinal fluid ubiquitin levels were found to be 147.5 +/- 5.2 ng ml-1 in non-neurological aged cases.

Alzheimer disease: correlation of cerebro-spinal fluid and brain ubiquitin levels.

Neurofibrillary degeneration is one of the histopathological hallmarks of Alzheimer disease (AD). Previous studies have shown an association of ubiquitin with the cytoskeletal protein pathology in AD. In the present study, we report (i) the measurement of ubiquitin levels in cerebrospinal fluid (CSF) from histopathologically confirmed AD and control cases, using a new rapid immunoassay, the competitive enzyme-linked immunoflow assay (CELIFA), (ii) the determination of ubiquitin levels in brain tissue taken from the same cases, using a competitive enzyme-linked immunosorbent assay (ELISA), and (iii) an evaluation of the correlation between levels of ubiquitin in CSF and in brain tissue. Ubiquitin levels in CSF of AD and neurological control groups are significantly higher than those of non-neurological aged controls. Ubiquitin levels in brain homogenates of the AD group are significantly higher than those of both non-neurological aged and neurological control groups. The source of this increase in brain ubiquitin in AD is the particulate fraction, because ubiquitin levels in the brain cytosol fraction are the same among the three groups. In AD and non-neurological aged controls, there is a significant positive correlation between ubiquitin levels in CSF and in homogenate of cerebral white matter. In contrast, the correlation in the non-AD neurological control group has a negative tendency. These studies suggest that in AD, elevated levels of ubiquitin in the CSF reflect the increased amount of the protein in the brain and, therefore, can serve as a biomarker of the neuropathology in this disease.

Marked increase in cerebrospinal fluid ubiquitin in Creutzfeldt-Jakob disease.

We have established the radioimmunoassay for ubiquitin in the cerebrospinal fluid (CSF) and measured the ubiquitin concentration in CSF from 4 cases of neuropathologically verified Creutzfeldt-Jakob disease (CJD), 10 cases of multi-infarct dementia (MID), 7 cases of senile dementia of Alzheimer type (SDAT), and 18 controls. The normal values were determined to range from 7.3 to 21.0 ng/ml, 14.3 +/- 1.1 ng/ml in the mean +/- S.E.M. The CSF ubiquitin levels in the cases of MID and SDAT were 16.6 +/- 6.4 ng/ml and 21.3 +/- 6.1 ng/ml, respectively. In the cases of CJD, the CSF ubiquitin was markedly increased at the early and middle stages of the disease (230.6 ng/ml in Case 1, 107.6 ng/ml in Case 2, 212.5 ng/ml in Case 3, and 377.0 ng/ml in Case 4) and these gradually decreased as the disease progressed. The measurement of CSF ubiquitin seems useful to make an early diagnosis of CJD.

[A case of Creutzfeldt-Jakob disease with markedly elevated ubiquitin concentration in the cerebrospinal fluid].

A 56-year-old woman initially noticed dizziness in October, 1988, and later dementia and gait disturbance developed, associated with myoclonus and periodic synchronous discharge in the electroencephalogram. On the basis of these clinical findings we made a diagnosis of Creutzfeldt-Jakob disease (CJD). Using RIA for ubiquitin (signal peptide of the ATP dependent proteolytic system), we measured the cerebrospinal fluid (CSF) ubiquitin levels. The CSF level of ubiquitin was markedly elevated in this case five months after the initial symptoms (230.0 ng/ml) compared with normal values (14.3 +/- 1.1 ng/ml) and values in patients with senile dementia of Alzheimer type (21.3 +/- 6.1 ng/ml) and vascular dementia (16.6 +/- 6.4 ng/ml). With progression of brain atrophy in this case, CSF levels of ubiquitin rapidly decreased to near the normal values. These findings suggest that CSF ubiquitin concentration reflects the activity of the disease process in CJD, and it may be useful in the diagnosis of CJD.

Alzheimer's disease: paired helical filament immunoreactivity in cerebrospinal fluid.

A competitive enzyme-linked immunosorbent assay with high sensitivity has been developed for measuring ubiquitin reactivity of paired helical filaments (PHF). Using the assay, ubiquitin immunoreactivity was estimated in the cerebrospinal fluid (CSF) of 44 patients who had been clinically diagnosed as having Alzheimer's disease (AD) and of 38 control patients, including 20 neurological cases. Monoclonal antibody (mAb) 5-25 to isolated paired helical filaments was used. This mAb recognizes amino acids 64-76 of ubiquitin. The levels of ubiquitin immunoreactivity measured in CSF (twice diluted) were significantly higher in the AD (p less than 0.001) than in the control group. In only a minority of instances were values for ubiquitin levels the same in AD and control groups: on PHF-coated plates, immunoreactivity values for 77% of the AD CSF specimens were higher than those for 92% of the controls, and on ubiquitin-coated plates, values for 85% of the AD CSF specimens were higher than those for 88% of the controls. On immunoblots of both AD and control CSF, mAb 5-25 stained a series of protein bands. The free ubiquitin that is also present in the CSF was not labeled. No striking differences were detected in the immunoblot pattern of AD and control CSF. This study demonstrates the presence of quantitative differences in the conjugated ubiquitin in AD and control CSF.