Abstract
The identification of differentially regulated proteins in animal models of psychiatric diseases is essential for a comprehensive analysis of associated psychopathological processes. Mass spectrometry is the most relevant method for analyzing differences in protein expression of tissue and body fluid proteomes. However, standardization of sample handling and sample-to-sample variability are problematic. Stable isotope metabolic labeling of a proteome represents the gold standard for quantitative mass spectrometry analysis. The simultaneous processing of a mixture of labeled and unlabeled samples allows a sensitive and accurate comparative analysis between the respective proteomes. Here, we describe a cost-effective feeding protocol based on a newly developed 15N bacteria diet based on Ralstonia eutropha protein, which was applied to a mouse model for trait anxiety. Tissue from 15N-labeled vs. 14N-unlabeled mice was examined by mass spectrometry and differences in the expression of glyoxalase-1 (GLO1) and histidine triad nucleotide binding protein 2 (Hint2) proteins were correlated with the animals' psychopathological behaviors for methodological validation and proof of concept, respectively. Additionally, phenotyping unraveled an antidepressant-like effect of the incorporation of the stable isotope 15N into the proteome of highly anxious mice. This novel phenomenon is of considerable relevance to the metabolic labeling method and could provide an opportunity for the discovery of candidate proteins involved in depression-like behavior. The newly developed 15N bacteria diet provides researchers a novel tool to discover disease-relevant protein expression differences in mouse models using quantitative mass spectrometry.
Highlights
The identification of candidate biomarkers as novel diagnostic tools and drug targets for psychopathologies is of great importance in neuropsychiatric research
In preliminary studies we found that the offspring of CD1 dams fed exclusively with a blue-green algae diet had severe developmental problems that led to death by malnutrition at about 10 days after birth
We demonstrate that 15N labeling of mice via a bacteria diet is a feasible and cost-effective method to study the proteomic differences related to psychopathologies in the HAB/NAB/LAB animal model using Mass spectrometry (MS)
Summary
The identification of candidate biomarkers as novel diagnostic tools and drug targets for psychopathologies is of great importance in neuropsychiatric research. While genes fundamentally shape physiology and pathophysiology, proteins are the final executive force of all cellular processes that drive physiology and behavior in health and pathology. Whole proteome approaches are scarce due to the complexity of differential protein analysis. Mass spectrometry (MS) is currently the most comprehensive method to analyze differences in protein expression and formation of the proteome. For an accurate and sensitive comparative proteome analysis metabolic labeling of one sample with a stable isotope is the preferred approach. This method results in an enrichment of the stable isotope in every protein in vivo, which can be compared with an unlabeled proteome by combining the two samples prior to MS analysis
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