Barbara Ciesielczyk

Tau Protein, Aβ42 and S-100B Protein in Cerebrospinal Fluid of Patients with Dementia with Lewy Bodies

The intra vitam diagnosis of dementia with Lewy bodies (DLB) is still based on clinical grounds. So far no technical investigations have been available to support this diagnosis. As for tau protein and β-amyloid(1–42) (Aβ42), promising results for the diagnosis of Alzheimer’s disease (AD) have been reported; we evaluated these markers and S-100B protein in cerebrospinal fluid (CSF), using a set of commercially available assays, of 71 patients with DLB, 67 patients with AD and 41 nondemented controls (NDC) for their differential diagnostic relevance. Patients with DLB showed significantly lower tau protein values compared to AD but with a high overlap of values. More prominent differences were observed in the comparison of DLB patients with all three clinical core features and AD patients. Aβ42 levels were decreased in the DLB and AD groups versus NDC, without significant subgroup differences. S-100B levels were not significantly different between the groups. Tau protein levels in CSF may contribute to the clinical distinction between DLB and AD, but the value of the markers is still limited especially due to mixed pathology. We conclude that more specific markers have to be established for the differentiation of these diseases.

Total tau protein, phosphorylated tau (181p) protein, β-amyloid1-42, and β-amyloid1-40 in cerebrospinal fluid of patients with dementia with Lewy bodies

Abstract The intra vitam diagnosis of different dementias is still based on clinical grounds. So far, no technical investigations have been available to support these diagnoses. For tau protein and β-amyloid(1–42) in cere-brospinal fluid (CSF), promising results for the diagnosis of Alzheimers disease (AD) have been reported; however, their differential diagnostic spectrum is limited, as was recently shown for dementia with Lewy bodies (DLB) and for AD. Therefore, further marker proteins have to be established to ameliorate, support, and differentiate these clinical diagnoses. We evaluated β-amyloid(1–40) and phosphorylated tau protein (181p), in addition to total tau protein and β-amyloid(1–42), in 20 patients with DLB, 34 AD patients, and 20 non-demented neurological controls (NDCs). All markers could differentiate between the dementia groups (AD, DLB) and the controls. AD and DLB could be differentiated only by levels of total tau protein and by the ratio total tau protein/phosphorylated tau protein. However, values still overlapped markedly. In some cases, tau protein levels in CSF may contribute to the clinical distinction between DLB and AD, but the value of the markers is still limited, especially because of mixed pathology. We conclude that more specific markers have to be established to differentiate between these diseases.

Follow-up investigations in cerebrospinal fluid of patients with dementia with Lewy bodies and Alzheimer’s disease

Summary.Measuring proteins in cerebrospinal fluid (CSF) has gained wide acceptance for the differential diagnosis of dementia. Some groups have already extended these investigations in Alzheimer’s disease (AD) by asking how stable these markers are in follow-up analysis, if they depend on the stage of disease and whether they can be used to monitor the progression and biological effects of treatment. We evaluated 21 patients with dementia with Lewy bodies (DLB) and 19 patients with AD, on two occasions, with regard to levels of tau protein, tau protein phosphorylated at threonine 181 (p-tau), Aβ42, Aβ40 and S-100B protein, using a set of commercially available assays.Tau protein levels were lower in DLB in first and second LP compared to AD and decreased during course of both groups. P-tau levels were increased in AD and DLB and decreased during follow-up. Aβ42 and Aβ40 remained relatively stable during follow-up but we found a slight increase of the median Aβ42 level in DLB, whereas in AD, Aβ42 tends to decrease during follow-up. S-100B protein increased during follow-up in both diseases.The protein dynamics in DLB and AD are relatively similar. S-100B protein may be a useful marker for follow-up in neurodegenerative diseases but has to be analysed in longer follow-up periods. Tau protein may be used to differentiate between DLB and AD.Follow-up CSF analyses are of limited value for the differentiation of AD and DLB. We conclude that more specific markers have to be established for the differentiation and follow-up of these diseases.

Total Prion Protein Levels in the Cerebrospinal Fluid are Reduced in Patients with Various Neurological Disorders

We performed a study on levels of the total prion protein (PrP) in humans affected by different neurological diseases and assessed the influence of several factors such as age, gender, and disease severity on the cerebrospinal fluid PrP levels. PrP-ELISA technique was used to analyze cerebrospinal fluid (CSF) samples. 293 CSF samples of patients with Creutzfeldt-Jakob-disease (CJD), Alzheimers disease, dementia with Lewy-bodies, Parkinsons disease, multiple sclerosis, cerebral ischemia, generalized epileptic seizures, and meningitis and encephalitis in comparison to controls were analyzed. We found a significant reduction of CSF PrP levels in patients suffering from all neurodegenerative disorders analyzed. This group exhibited mean PrP values of 164 ng/ml while non-neurodegenerative disorder patients and healthy controls showed PrP levels of 208 ng/ml and 226 ng/ml, respectively. CSF levels correlated with disease severity in CJD, Alzheimers disease, and dementia with Lewy-bodies. The finding of decreased PrP levels in the CSF of patients not only with CJD but also in other neurodegenerative disorders is intriguing. Age-, gender-, and genetic-specific factors might be involved in the PrP c regulation.

8-OHdG in Cerebrospinal Fluid as a Marker of Oxidative Stress in Various Neurodegenerative Diseases

Background: The 8-hydroxy-2 deoxyguanosine (8-OHdG) is a product of nucleoside oxidation of DNA and a reliable marker of oxidative stress markers. Increased levels of oxidative stress have been reported in the cerebrospinal fluid (CSF) of patients with various neurodegenerative disorders. Objective: In search of a biochemical indicator of Parkinson’s disease (PD), we analyzed the levels 8-OHdG in the CSF of 99 patients, using ELISA to assess the differences between various neurodegenerative disorders. Results: Statistically significant higher CSF levels (p = 0.022) of 8-OHdG in non-demented PD patients as compared to the control group were observed. No differences between CSF 8-OHdG levels and age at the time of lumbar puncture, presence or severity of dementia, or gender were found. Conclusions: 8-OHdG levels could be potentially useful in the neurochemically supported diagnosis of PD.

Brain-derived proteins in the CSF, do they correlate with brain pathology in CJD?

BackgroundBrain derived proteins such as 14-3-3, neuron-specific enolase (NSE), S 100b, tau, phosphorylated tau and Aβ1–42 were found to be altered in the cerebrospinal fluid (CSF) in Creutzfeldt-Jakob disease (CJD) patients. The pathogenic mechanisms leading to these abnormalities are not known, but a relation to rapid neuronal damage is assumed. No systematic analysis on brain-derived proteins in the CSF and neuropathological lesion profiles has been performed.MethodsCSF protein levels of brain-derived proteins and the degree of spongiform changes, neuronal loss and gliosis in various brain areas were analyzed in 57 CJD patients.ResultsWe observed three different patterns of CSF alteration associated with the degree of cortical and subcortical changes. NSE levels increased with lesion severity of subcortical areas. Tau and 14-3-3 levels increased with minor pathological changes, a negative correlation was observed with severity of cortical lesions. Levels of the physiological form of the prion protein (PrPc) and Aβ1–42 levels correlated negatively with cortical pathology, most clearly with temporal and occipital lesions.ConclusionOur results indicate that the alteration of levels of brain-derived proteins in the CSF does not only reflect the degree of neuronal damage, but it is also modified by the localization on the brain pathology. Brain specific lesion patterns have to be considered when analyzing CSF neuronal proteins.

CELLULAR PRION PROTEIN OVEREXPRESSION DISTURBS CELLULAR HOMEOSTASIS IN SH-SY5Y NEUROBLASTOMA CELLS BUT DOES NOT ALTER p53 EXPRESSION: A PROTEOMIC STUDY

The definite physiological role of the cellular prion protein (PrP(c)) remains elusive. There is ample in vitro and in vivo evidence suggesting a neuroprotective role for PrP(c). On the other hand, several in vitro and in vivo studies demonstrated detrimental effects of PrP(c) overexpression through activation of a p53 pathway. Recently, we reported that transient overexpression of PrP(c) in human embryonic kidney 293 cells elicits proteome expression changes which point to deregulation of proteins involved in energy metabolism and cellular homeostasis. Here we report proteome expression changes following stable PrP(c) overexpression in human neuronal SH-SY5Y cells. In total 18 proteins that are involved in diverse biological processes were identified as differentially regulated. The majority of these proteins is involved in cell signaling, cytoskeletal organization and protein folding. Annexin V exhibited a several fold up-regulation following stable PrP(c) overexpression in SH-SY5Y cells. This finding has been reproduced in alternative, mouse N2a and human SK-N-LO neuroblastoma cell lines transiently overexpressing PrP(c). Annexin V plays an important role in maintenance of calcium homeostasis which when disturbed can activate a p53-dependent cell death. Although we did not detect changes in p53 expression between PrP(c) overexpressing SH-SY5Y and control cells, deregulation of several proteins including annexin V, polyglutamine tract-binding protein-1, spermine synthase and transgelin 2 indicates disrupted cellular equilibrium. We conclude that stable PrP(c) overexpression in SH-SY5Y cells is sufficient to perturb cellular balance but insufficient to affect p53 expression.

Follow-Up Investigations of Tau Protein and S-100B Levels in Cerebrospinal Fluid of Patients with Creutzfeldt-Jakob Disease

Background: S-100B and tau protein have a high differential diagnostic potential for the diagnosis of Creutzfeldt-Jakob disease (CJD). So far there has been only limited information available about the dynamics of these parameters in the cerebrospinal fluid (CSF). However, there is a special interest in finding biochemical markers to monitor disease progression for differential diagnosis and treatment. Patients and Methods: We analyzed CSF of 45 patients with CJD and of 45 patients with other neurological diseases for tau protein and S-100B in a follow-up setting. All diagnoses of CJD were later neuropathologically verified. A ratio between tau protein differences and the time between lumbar puncture was calculated. The same was done for S-100B. Results: Tau protein levels of 34 cases were above the cut-off level for CJD (>1,300 pg/ml) in the first CSF sample. In 7 of 11 patients with lower tau levels in the first CSF sample, tau levels rose. The above-mentioned ratio was significantly higher in the CJD group than in the group with other neurological diseases. Similar results were obtained for S-100B. Conclusion: We conclude that follow-up investigations and calculation of ratios is a useful tool in the differential diagnosis of CJD. Variations in this pattern were observed in single cases.

Transcription of Alu DNA elements in blood cells of sporadic Creutzfeldt-Jakob disease (sCJD)

Alu DNA elements were long considered to be of no biological significance and thus have been only poorly defined. However, in the past Alu DNA elements with well-defined nucleotide sequences have been suspected to contribute to disease, but the role of Alu DNA element transcripts has rarely been investigated. For the first time, we determined in a real-time approach Alu DNA element transcription in buffy coat cells isolated from the blood of humans suffering from sporadic Creutzfeldt-Jakob disease (sCJD) and other neurodegenerative disorders. The reverse transcribed Alu transcripts were amplified and their cDNA sequences were aligned to genomic regions best fitted to database genomic Alu DNA element sequences deposited in the UCSC and NCBI data bases. Our cloned Alu RNA/cDNA sequences were widely distributed in the human genome and preferably belonged to the “young” Alu Y family. We also observed that some RNA/cDNA clones could be aligned to several chromosomes because of the same degree of identity and score to resident genomic Alu DNA elements. These elements, called paralogues, have purportedly been recently generated by retrotransposition. Along with cases of sCJD we also included cases of dementia and Alzheimer’s disease (AD). Each group revealed a divergent pattern of transcribed Alu elements. Chromosome 2 was the most preferred site in sCJD cases, besides chromosome 17; in AD cases chromosome 11 was overrepresented whereas chromosomes 2, 3 and 17 were preferred active Alu loci in controls. Chromosomes 2, 12 and 17 gave rise to Alu transcripts in dementia cases. The detection of putative Alu paralogues widely differed depending on the disease. A detailed data search revealed that some cloned Alu transcripts originated from RNA polymerase III transcription since the genomic sites of their Alu elements were found between genes. Other Alu DNA elements could be located close to or within coding regions of genes. In general, our observations suggest that identification and genomic localization of active Alu DNA elements could be further developed as a surrogate marker for differential gene expression in disease. A sufficient number of cases are necessary for statistical significance before Alu DNA elements can be considered useful to differentiate neurodegenerative diseases from controls.

Desmoplakin as a Potential Candidate for Cerebrospinal Fluid Marker to Rule Out 14-3-3 False Positive Rates in Sporadic Creutzfeldt-Jakob Disease Differential Diagnosis

Background: The detection of a 14-3-3 elevated level in cerebrospinal fluid (CSF) is a part of the diagnostic criteria for probable sporadic Creutzfeldt-Jakob disease (sCJD), as defined by the WHO. However, some pathological conditions associated with acute neuronal damage may result in a positive 14-3-3 test and thereby reduce test specificity in sCJD. Objective: Desmoplakin has been previously identified as up-regulated CSF protein in sCJD and these studies aimed to investigate its diagnostic utility and compare it with two known CSF markers, 14-3-3 and tau. Methods and Results: We tested CSF levels of 14-3-3, tau and desmoplakin in 58 sCJD patients and 81 control patients including 45 cases with an elevated 14-3-3 level due to other disease than sCJD. We detected an elevated CSF level of desmoplakin in 78% of the sCJD patients, while 14-3-3 (88%) and tau (91%) showed a higher positive rate. However, the false positive rate of newly tested desmoplakin was significantly lower in comparison to 14-3-3 and tau, and it accounted for only 11% versus 56% and 35%, respectively. Further reduction of false positive rates was achieved by combination of elevated tau level with a positive desmoplakin test. Moreover, in the non-sCJD group, desmoplakin level did not correlate with the level of both above-mentioned CSF markers, whereas a clear correlation was observed in the sCJD group. Conclusion: Desmoplakin showed a low positive rate accompanied by a very low false positive rate. Thus, we conclude that desmoplakin is a promising candidate for supportive CSF marker to rule out 14-3-3 false positive cases in sCJD differential diagnosis.