Sebastian Brandner

Early CSF and Serum S100B Concentrations for Outcome Prediction in Traumatic Brain Injury and Subarachnoid Hemorrhage.

OBJECTIVES S100B has been proposed as a putative biochemical marker in determining the extent of brain injury and corresponding prognosis in neurotrauma. The aim of this study was to evaluate the prognostic value of S100B early concentrations in serum and cerebrospinal fluid (CSF) in traumatic brain injury (TBI) and subarachnoid hemorrhage (SAH), to determine prognostically relevant threshold values and to evaluate fluctuation following EVD placement. PATIENTS AND METHODS In 102 patients (45 SAH and 57 TBI) under intensive care unit (ICU) treated between January 2011 and December 2012 with external ventricular drain (EVD) S100B measurements were performed simultaneously in serum and CSF during the first 5 days and before and after EVD exchange. Glasgow coma scale (GCS) was assessed on admission and Glasgow outcome scale (GOS) 6 months later. RESULTS Peak S100B levels in CSF and serum were measured on the first day after admission and concentrations decreased during the ensuing days post injury gradually. CSF and serum S100B concentrations in TBI patients were significantly higher than in SAH (p<0.005). Both in TBI and SAH patients S100B concentrations in CSF and serum were significantly higher in patients with an unfavorable outcome (GOS 1-3) in comparison to patients with a good outcome (GOS 4-5). Correlation of S100B concentrations in serum and GOS score at 6 months was significant both in TBI and SAH (p<0.05). Serum S100B concentrations >0.7μg/l correlated with 100% mortality. Correlation between S100B in CSF and GOS was significant in SAH (p<0.05), whereas it was not significant in TBI. After EVD exchange (n=53) we found a significant increase of S100B concentration in CSF (p<0.005). CONCLUSION Initial S100B levels have a limited prognostic value in neurotrauma with CSF concentrations being highly sensitive to smallest influences like EVD placement. However, high initial S100B levels of >0.7μg/dl in serum are associated with 100% mortality, which might help to guide therapy strategies in severe neurotrauma.

Neuroprotein dynamics in the cerebrospinal fluid: intraindividual concomitant ventricular and lumbar measurements.

Objective: The measurement of neuromarker/neuroproteins in the cerebrospinal fluid (CSF) is gaining increased popularity. However, insufficient information is available on the rostrocaudal distribution of neuroproteins in the CSF to guarantee an appropriate interpretation of ventricular versus lumbar concentrations. Methods: In 10 patients treated with both an external ventricular and a lumbar CSF drain, we collected concomitant CSF samples. We measured CSF concentrations of the glial S100B protein, the neuron-specific enolase (Cobas e411®; Roche Diagnostics), the leptomeningeal β-trace protein (BN Pro Spec®; Dade Behring/Siemens), and the blood-derived albumin (Immage; Beckman Coulter). Statistical analysis was performed with a paired Wilcoxon signed ranks test. Results: In patients with a free CSF circulation without any recent neurosurgical procedure, S100B and neuron-specific enolase concentrations did not differ between the ventricular and lumbar CSF while β-trace and albumin levels were significantly higher in the lumbar than in the ventricular CSF (p = 0.008 and p = 0.005). Following posterior fossa tumor surgery, all proteins accumulate in the lumbar CSF. Conclusion: For brain-derived proteins, we could not confirm a rostrocaudal CSF gradient while lepto-meningeal and blood-derived proteins accumulate in the lumbar CSF. We conclude that for the interpretation of protein CSF concentrations, the source of the sample is of crucial importance.

Comparison of Different Matrices as Potential Quality Control Samples for Neurochemical Dementia Diagnostics

BACKGROUND Assay-vendor independent quality control (QC) samples for neurochemical dementia diagnostics (NDD) biomarkers are so far commercially unavailable. This requires that NDD laboratories prepare their own QC samples, for example by pooling leftover cerebrospinal fluid (CSF) samples. OBJECTIVE To prepare and test alternative matrices for QC samples that could facilitate intra- and inter-laboratory QC of the NDD biomarkers. METHODS Three matrices were validated in this study: (A) human pooled CSF, (B) Aβ peptides spiked into human prediluted plasma, and (C) Aβ peptides spiked into solution of bovine serum albumin in phosphate-buffered saline. All matrices were tested also after supplementation with an antibacterial agent (sodium azide). We analyzed short- and long-term stability of the biomarkers with ELISA and chemiluminescence (Fujirebio Europe, MSD, IBL International), and performed an inter-laboratory variability study. RESULTS NDD biomarkers turned out to be stable in almost all samples stored at the tested conditions for up to 14 days as well as in samples stored deep-frozen (at - 80°C) for up to one year. Sodium azide did not influence biomarker stability. Inter-center variability of the samples sent at room temperature (pooled CSF, freeze-dried CSF, and four artificial matrices) was comparable to the results obtained on deep-frozen samples in other large-scale projects. CONCLUSION Our results suggest that it is possible to replace self-made, CSF-based QC samples with large-scale volumes of QC materials prepared with artificial peptides and matrices. This would greatly facilitate intra- and inter-laboratory QC schedules for NDD measurements.

Quantification of serial changes in cerebral blood volume and metabolism in patients with recurrent glioblastoma undergoing antiangiogenic therapy.

OBJECTIVES To evaluate the usefulness of quantitative advanced magnetic resonance imaging (MRI) methods for assessment of antiangiogenic therapy (AAT) response in recurrent glioblastoma multiforme (GBM). METHODS Eighteen patients with recurrent GBM received bevacizumab and 18 patients served as control group. Baseline MRI and two follow-up examinations were acquired every 3-5 months using dynamic susceptibility-weighted contrast (DSC) perfusion MRI and (1)H-MR spectroscopic imaging ((1)H-MRSI). Maps of absolute cerebral blood volume (aCBV) were coregistered with choline (Cho) and N-acetyl-aspartate (NAA) concentrations and compared to usually used relative parameters as well as controls. RESULTS Perfusion significantly decreased in responding and pseudoresponding GBMs but also in normal appearing brain after AAT onset. Cho and NAA concentrations were superior to Cr-ratios in lesion differentiation and showed a clear gap between responding and pseudoresponding lesions. Responders to AAT exceptionally frequently (6 out of 8 patients) showed remote GBM progression. CONCLUSIONS Quantification of CBV reveals changes in normal brain perfusion due to AAT, which were not described so far. DSC perfusion MRI seems not to be suitable for differentiation between response and pseudoresponse to AAT. However, absolute quantification of brain metabolites may allow for distinction due to a clear gap at 6-9 months after therapy onset.

Ventricular and Lumbar Cerebrospinal Fluid Concentrations of Alzheimer's Disease Biomarkers in Patients with Normal Pressure Hydrocephalus and Posttraumatic Hydrocephalus

BACKGROUND Little information is available on the rostro-caudal concentration gradient of Alzheimers disease (AD) biomarkers. OBJECTIVE We studied the concentrations of amyloid-β (Aβ) peptides 1-42 and 1-40 as well as the Tau and pTau proteins in simultaneously collected ventricular and lumbar cerebrospinal fluid (CSF) samples. METHODS The samples were simultaneously collected from the ventricle and the lumbar spinal canal in two groups of patients: 10 subjects being treated for normal pressure hydrocephalus (NPH) by the placement of a ventriculo-peritoneal shunt and 5 patients treated simultaneously with an external ventricular drain and a lumbar CSF drain due to posttraumatic hydrocephalus (PTH). RESULTS The ventricular-lumbar (V/L) concentration ratio for Aβ1-40 was 0.81 in NPH patients and 0.71 in PTH patients. The V/L-ratio for Aβ1-42 was 0.84 in NPH, reflecting significantly higher concentrations in lumbar CSF than in ventricular CSF, and 1.02 in PTH patients. The V/L-ratios for Tau and pTau differed significantly depending on the diagnostic group: the median V/L-ratio for Tau was 6.83 in NPH patients but only 0.97 in PTH patients. The median V/L-ratio for pTau was 2.36 in NPH patients and 0.91 in PTH patients. CONCLUSIONS We conclude that the rostro-caudal concentration gradient for brain-derived proteins (Tau and pTau in this study) depends on the diagnosis and clinical status of the patient, which were largely neglected in the previously postulated models.

Shunt-Dependent Hydrocephalus Following Subarachnoid Hemorrhage Correlates with Increased S100B Levels in Cerebrospinal Fluid and Serum

Posthemorrhagic hydrocephalus requiring permanent ventriculoperitoneal shunt placement is a major complication of aneurysmal subarachnoid hemorrhage (SAH). High S100B serum and cerebrospinal fluid (CSF) levels are considered to reflect the severity of brain injury. We prospectively assessed whether S100B levels in serum and CSF were predictive parameters for permanent shunt requirement following aneurysmal SAH. In patients suffering from aneurysmal SAH and treated with an external ventricular drain (EVD), S100B levels in serum and CSF were measured daily as long as the EVD was in place. S100B levels of patients who passed their EVD challenge were compared with those patients who required a permanent ventriculoperitoneal shunt placement. Out of 68 patients included in the study, 43 patients (63.2%) passed the EVD challenge and in 25 patients (36.8%) permanent ventriculoperitoneal shunting was performed. Group comparison revealed that in patients who required shunt placement, S100B was significantly higher in CSF (p < 0.05 at days 2, 4, 6, 10; p < 0.005 at days 1, 3, 5, 7, 8, 9) and serum (p < 0.05 at days 4-7) compared with patients who could be weaned from the EVD. Assessment of S100B levels in CSF and serum may be useful as a predictive parameter for shunt dependency in patients with posthemorrhagic hydrocephalus following aneurysmal SAH.

Brain-derived protein concentrations in the cerebrospinal fluid: contribution of trauma resulting from ventricular drain insertion.

In recent years, the measurement of biomarkers following neurotrauma assisted in improving outcome prediction and guiding therapy. The use of neuroproteins as diagnostic parameters requires a detailed knowledge of their dynamics in biological fluids for an appropriate interpretation. S100B is the most widely studied neuromarker, and its concentration in serum and cerebrospinal fluid (CSF) reflects the extent of brain damage. Neuron-specific enolase (NSE) is considered reflecting neuronal damage, while Beta-Trace is a lepto-meningeal protein used to diagnose CSF leakage. In five patients treated with an external ventricular drain (EVD) because of aneurysmal subarachnoid hemorrhage (SAH, n=3) or postinfectious hydrocephalus (n=2), an EVD exchange was performed 8 to 12 days after initial insertion. S100B and NSE were measured with the Cobas e411(®) electrochemiluminescence assay (Roche Diagnostics, Mannheim, Germany) and Beta-Trace with the BN Pro Spec(®) nephelometer (Dade Behring/Siemens, Germany) 1 h before EVD exchange, upon the insertion of the new drain, and 1, 3, 6, 12, 18, 24 and 48 h after EVD exchange. Before EVD exchange, S100B CSF concentrations were within the normal range in all patients (1.48 ± 0.37 μg/L), while NSE CSF concentrations were normal in four of five patients (6.51 ± 2.98 μg/L). Following EVD exchange, S100B and NSE CSF levels peaked significantly at 3 h after insertion of the new drain (S100B 39.02 ± 9.17 μg/L; NSE 54.80 ± 43.34 μg/L). S100B serum levels were slightly increased 6 to 24 h after EVD exchange. Beta-Trace concentrations in the CSF were not altered by EVD insertion. Our data demonstrate that EVD insertion results in a distinct increase of S100B and NSE concentrations in the CSF. Thus, the tampering of brain-derived protein concentrations in the CSF by diagnostic or therapeutic procedures has to be considered in the interpretation of neuromarker levels.

Non-Phosphorylated Tau as a Potential Biomarker of Alzheimer’s Disease: Analytical and Diagnostic Characterization

BACKGROUND Virtually nothing is known about a potential diagnostic role of non-phospho-epitopes of Tau (Non-P-Tau) in cerebrospinal fluid (CSF). OBJECTIVE To establish and analytically and clinically characterize the first assay capable to measure concentrations of Non-P-Tau in human CSF. METHODS An antibody (1G2) was developed that selectively binds to the Tau molecule non-phosphorylated at the positions T175 and T181, and was used in establishing a sandwich ELISA capable to measure Non-P-Tau in human CSF, following analytical and clinical validation of the method. RESULTS The 1G2 antibody shows decreasing reactivity to tau peptides containing phosphorylation mainly at positions T175 and T181. Detection limit of the assay is 25 pg/ml; the coefficients of variation (CVs) of the optical densities of the repeated standard curves were between 3.6-15.9%. Median intra-assay imprecision of double measurements was 4.8%; inter-assay imprecision was in the range of 11.2% - 15.3%. Non-P-Tau concentrations are stable in the CSF samples sent to distinct laboratories under ambient temperature; inter-laboratory variation was approximately 30%. The Non-P-Tau CSF concentrations were highly significantly increased in patients with Alzheimers disease in stage of mild cognitive impairment or dementia (AD/MCI, n = 58, 109.2±32.0 pg/mL) compared to the non-demented Controls (n = 42, 62.1±9.3 pg/mL, p < 0.001). At the cut-off of 78.3 pg/mL, the sensitivity and the specificity were 94.8% and 97.6%, respectively. CONCLUSION For the first time, an assay is reported to reliably measure concentrations of non-phosphorylated Tau in human CSF.

Plasma Concentrations of the Amyloid-β Peptides in Young Volunteers: The Influence of the APOE Genotype

Changes in the concentrations of amyloid-β (Aβ) in the body fluids are the earliest alterations observed in Alzheimers disease (AD), however, there is a lack of data about how early these alterations occur, before the onset of the clinical symptoms. APOE genotype is the most recognized genetic risk/protective factor of AD, meaning that a group of non-demented persons carrying ε4 allele is enriched in the subjects who will develop AD, compared to the group of non-carriers. Therefore, we studied the plasma concentrations of Aβ peptides (Aβ1-42, Aβ1-40, Aβx-42, and Aβx-40), and the APOE genotype in 173 young volunteers (average age, 28 ± 7.6 years) without memory deficits, in order to see whether the non-demented group of subjects at risk already characterize with Aβ changes three-to-four decades before the age at which dementia usually occurs. We did not find statistically significant differences among the groups of ε4 carriers, ε3 homozygotes, and ε2 carriers. We conclude that the APOE genotype does not influence the metabolism of the Aβ peptides in young persons without memory deficits.

Interlaboratory proficiency processing scheme in CSF aliquoting: implementation and assessment based on biomarkers of Alzheimer’s disease

BackgroundIn this study, we tested to which extent possible between-center differences in standardized operating procedures (SOPs) for biobanking of cerebrospinal fluid (CSF) samples influence the homogeneity of the resulting aliquots and, consequently, the concentrations of the centrally analyzed selected Alzheimer’s disease biomarkers.MethodsProficiency processing samples (PPSs), prepared by pooling of four individual CSF samples, were sent to 10 participating centers, which were asked to perform aliquoting of the PPSs into two secondary aliquots (SAs) under their local SOPs. The resulting SAs were shipped to the central laboratory, where the concentrations of amyloid beta (Aβ) 1–42, pTau181, and albumin were measured in one run with validated routine analytical methods. Total variability of the concentrations, and its within-center and between-center components, were analyzed with hierarchical regression models.ResultsWe observed neglectable variability in the concentrations of pTau181 and albumin across the centers and the aliquots. In contrast, the variability of the Aβ1–42 concentrations was much larger (overall coefficient of variation 31%), with 28% of the between-laboratory component and 10% of the within-laboratory (i.e., between-aliquot) component. We identified duration of the preparation of the aliquots and the centrifugation force as two potential confounders influencing within-center variability and biomarker concentrations, respectively.ConclusionsProficiency processing schemes provide objective evidence for the most critical preanalytical variables. Standardization of these variables may significantly enhance the quality of the collected biospecimens. Studies utilizing retrospective samples collected under different local SOPs need to consider such differences in the statistical evaluations of the data.