Disease biomarkers in cerebrospinal fluid of patients with first-onset psychosis

BACKGROUND: Psychosis is a severe mental condition that is characterized by a loss of contact with reality and is typically associated with hallucinations and delusional beliefs. There are numerous psychiatric conditions that present with psychotic symptoms, most importantly schizophrenia, bipolar affective disorder, and some forms of severe depression referred to as psychotic depression. The pathological mechanisms resulting in psychotic symptoms are not understood, nor is it understood whether the various psychotic illnesses are the result of similar biochemical disturbances. The identification of biological markers (so-called biomarkers) of psychosis is a fundamental step towards a better understanding of the pathogenesis of psychosis and holds the potential for more objective testing methods. METHODS AND FINDINGS: Surface-enhanced laser desorption ionization mass spectrometry was employed to profile proteins and peptides in a total of 179 cerebrospinal fluid samples (58 schizophrenia patients, 16 patients with depression, five patients with obsessive-compulsive disorder, ten patients with Alzheimer disease, and 90 controls). Our results show a highly significant differential distribution of samples from healthy volunteers away from drug-naïve patients with first-onset paranoid schizophrenia. The key alterations were the up-regulation of a 40-amino acid VGF-derived peptide, the down-regulation of transthyretin at approximately 4 kDa, and a peptide cluster at approximately 6,800-7,300 Da (which is likely to be influenced by the doubly charged ions of the transthyretin protein cluster). These schizophrenia-specific protein/peptide changes were replicated in an independent sample set. Both experiments achieved a specificity of 95% and a sensitivity of 80% or 88% in the initial study and in a subsequent validation study, respectively. CONCLUSIONS: Our results suggest that the application of modern proteomics techniques, particularly mass spectrometric approaches, holds the potential to advance the understanding of the biochemical basis of psychiatric disorders and may in turn allow for the development of diagnostics and improved therapeutics. Further studies are required to validate the clinical effectiveness and disease specificity of the identified biomarkers

Metabolic Profiling of CSF: Evidence That Early Intervention May Impact on Disease Progression and Outcome in Schizophrenia

BACKGROUND: The identification of schizophrenia biomarkers is a crucial step towards improving current diagnosis, developing new presymptomatic treatments, identifying high-risk individuals and disease subgroups, and assessing the efficacy of preventative interventions at a rate that is not currently possible. METHODS AND FINDINGS: (1)H nuclear magnetic resonance spectroscopy in conjunction with computerized pattern recognition analysis were employed to investigate metabolic profiles of a total of 152 cerebrospinal fluid (CSF) samples from drug-naïve or minimally treated patients with first-onset paranoid schizophrenia (referred to as “schizophrenia” in the following text) and healthy controls. Partial least square discriminant analysis showed a highly significant separation of patients with first-onset schizophrenia away from healthy controls. Short-term treatment with antipsychotic medication resulted in a normalization of the disease signature in over half the patients, well before overt clinical improvement. No normalization was observed in patients in which treatment had not been initiated at first presentation, providing the first molecular evidence for the importance of early intervention for psychotic disorders. Furthermore, the alterations identified in drug-naïve patients could be validated in a test sample set achieving a sensitivity and specificity of 82% and 85%, respectively. CONCLUSIONS: Our findings suggest brain-specific alterations in glucoregulatory processes in the CSF of drug-naïve patients with first-onset schizophrenia, implying that these abnormalities are intrinsic to the disease, rather than a side effect of antipsychotic medication. Short-term treatment with atypical antipsychotic medication resulted in a normalization of the CSF disease signature in half the patients well before a clinical improvement would be expected. Furthermore, our results suggest that the initiation of antipsychotic treatment during a first psychotic episode may influence treatment response and/or outcome

Determination of anandamide and other fatty acyl ethanolamides in human serum by electrospray tandem mass spectrometry.

We developed a new selective liquid chromatography-electrospray ionization-tandem mass spectrometry method for the identification and quantification of anandamide (AEA), an endogenous cannabinoid receptor ligand, and other bioactive fatty acid ethanolamides (FAEs) in biological samples. Detection limit (0.025 pmol for AEA and 0.1 pmol for palmitoylethanolamide (PEA) and oleoylethanolamide (OEA)) and quantification limit (0.2 pmol for AEA and 0.4 pmol for OEA and PEA) were in the high fmol to low pmol range for all analytes. Linear correlations (r(2)=0.99) were observed in the calibration curves for standard AEA over the range of 0.025-25 pmol and for standard PEA and OEA over the range of 0.1-500 pmol. This method provides a time-saving and sensitive alternative to existing methods for the analysis of FAEs in biological samples

Anandamide levels in cerebrospinal fluid of first-episode schizophrenic patients: impact of cannabis use.

BackgroundPrevious studies have shown that cerebrospinal fluid (CSF) from schizophrenic patients contains significantly higher levels of the endogenous cannabinoid anandamide than does CSF from healthy volunteers. Moreover, CSF anandamide levels correlated inversely with psychotic symptoms, suggesting that anandamide release in the central nervous system (CNS) may serve as an adaptive mechanism countering neurotransmitter abnormalities in acute psychoses. In the present study we examined whether cannabis use may alter such a mechanism.MethodsWe used liquid chromatography/mass spectrometry (LC/MS) to measure anandamide levels in serum and CSF from first-episode, antipsychotic-naïve schizophrenics (n=47) and healthy volunteers (n=81). Based on reported patterns of cannabis use and urine delta9-tetrahydrocannabinol (delta9-THC) tests, each subject group was further divided into two subgroups: 'low-frequency' and 'high-frequency' cannabis users (lifetime use < or = 5 times and > 20 times, respectively). Serum delta9-THC was investigated to determine acute use and three patients were excluded from the analysis due to detectable delta9-THC levels in serum.ResultsSchizophrenic low-frequency cannabis users (n=25) exhibited > 10-fold higher CSF anandamide levels than did schizophrenic high-frequency users (n=19, p=0.008), healthy low-frequency (n=55, p<0.001) or high-frequency users (n=26, p<0.001). In contrast, no significant differences in serum anandamide levels were found among the four subgroups. CSF anandamide levels and disease symptoms were negatively correlated in both user groups.ConclusionsThe results indicate that frequent cannabis exposure may down-regulate anandamide signaling in the CNS of schizophrenic patients, but not of healthy individuals. Thus, our findings suggest that alterations in endocannabinoid signaling might be an important component of the mechanism through which cannabis impacts mental health

Validation and Prediction of Schizophrenia Group Membership Using a PLS Model

<p>A PLS model was constructed using the OSC-filtered data from 37 drug-naïve patients with first-onset schizophrenia from the first cohort (red points) and 50 randomly selected healthy volunteers (blue points) (the “training set”). The scores plot (A) and the loadings plot (B) indicate the key resonances contributing to the separation: lactate, glucose, glutamine, and citrate. This model was then used to predict “group membership” (i.e., schizophrenia or control) in a test set of 17 drug-naïve patients (second cohort) with first-onset schizophrenia and the remaining 20 healthy volunteers which had not been used in the construction of the model. Predictions are made using a Y-predicted scatter plot with an a priori cut-off of 0.5 for class membership (C).</p

Metabonomic Analysis of CSF Samples from Drug-Naïve Patients with Schizophrenia

<div><p>(A) Partial ¹H NMR spectrum of a CSF sample from a representative drug-naïve patient with first-onset schizophrenia (red) and a matched control (black) illustrate a characteristic pH-dependent shift in the β-CH₂ and γ-CH₂ resonances of glutamine. The prominent signals at ~3.7 and 1.2 ppm correspond to ethanol, a contaminant from skin disinfection prior to lumbar puncture. These signals were removed from statistical analysis.</p> <p>(B) PLS-DA scores plot showing a differentiation of drug-naïve patients with schizophrenia from demographically matched controls as determined by the ¹H NMR CSF spectra.</p> <p>(C) PLS-DA loadings plot showing major contributing variables towards the separation in the PLS-DA scores plots.</p></div

Effects of “Typical” and “Atypical” Medication on CSF Metabolic Profiles in Patients with First-Onset Schizophrenia

<div><p>(A) Spectra from 28 CSF samples from patients with first-onset schizophrenia minimally treated (<9 d, see text for details) with either typical (<i>n</i> = 6, blue diamonds) or atypical (<i>n</i> = 22, green circles) antipsychotic medications were compared to first onset, drug naïve patients (red triangles) and healthy volunteers (black squares) using PLS-DA. The PLS-DA scores plots show that atypical antipsychotic drug treatment resulted in a shift of approximately 50% of patients with schizophrenia towards the cluster of healthy controls.</p> <p>(B) The same PLS-DA scores plot as (A) except that only minimally treated patients (from both drug groups) with more than one psychotic episode prior to antipsychotic treatment are shown. Note that none of these patients shifted towards the healthy control cluster.</p></div

Mapping and Identification of the Biomarker Peptide Derived from the Native VGF Protein

<div><p>The 3,959-Da “schizophrenia” peptide was mapped to amino acids 23–62 of the native VGF protein (in bold; underlined), immediately next to a predicted secretory signal peptide (using InterProScan: <a href="http://www.ebi.ac.uk/cgi-bin/iprscan" target="_blank">http://www.ebi.ac.uk/cgi-bin/iprscan</a>).</p> <p>Another native VGF-derived peptide did not show differential expression between healthy volunteers and patients with schizophrenia (as shown in <a href="http://www.plosmedicine.org/article/info:doi/10.1371/journal.pmed.0030428#pmed-0030428-g002" target="_blank">Figure 2</a>A). This peptide has an identical sequence except that it is three amino acids shorter at the N-terminus compared to the “disease-specific” VGF peptide.</p></div