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Integrative proteomic analysis of the NMDA NR1 knockdown mouse model reveals effects on central and peripheral pathways associated with schizophrenia and autism spectrum disorders.

Wesseling H, Guest PC, Lee CM, Wong EH, Rahmoune H, Bahn S - Mol Autism (2014)

Bottom Line: The highest magnitude changes were found for neurotrophic factors (VEGFA, EGF, IGF-1), apolipoprotein A1, and fibrinogen.We also found decreased levels of several chemokines.In contrast, increased levels of proteins involved in neurotransmitter metabolism and release were found only in the frontal cortex and abnormalities of proteins involved in the purinergic system were found exclusively in the hippocampus.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QT, UK.

ABSTRACT

Background: Over the last decade, the transgenic N-methyl-D-aspartate receptor (NMDAR) NR1-knockdown mouse (NR1(neo-/-)) has been investigated as a glutamate hypofunction model for schizophrenia. Recent research has now revealed that the model also recapitulates cognitive and negative symptoms in the continuum of other psychiatric diseases, particularly autism spectrum disorders (ASD). As previous studies have mostly focussed on behavioural readouts, a molecular characterisation of this model will help to identify novel biomarkers or potential drug targets.

Methods: Here, we have used multiplex immunoassay analyses to investigate peripheral analyte alterations in serum of NR1(neo-/-) mice, as well as a combination of shotgun label-free liquid chromatography mass spectrometry, bioinformatic pathway analyses, and a shotgun-based 40-plex selected reaction monitoring (SRM) assay to investigate altered molecular pathways in the frontal cortex and hippocampus. All findings were cross compared to identify translatable findings between the brain and periphery.

Results: Multiplex immunoassay profiling led to identification of 29 analytes that were significantly altered in sera of NR1(neo-/-) mice. The highest magnitude changes were found for neurotrophic factors (VEGFA, EGF, IGF-1), apolipoprotein A1, and fibrinogen. We also found decreased levels of several chemokines. Following this, LC-MS(E) profiling led to identification of 48 significantly changed proteins in the frontal cortex and 41 in the hippocampus. In particular, MARCS, the mitochondrial pyruvate kinase, and CamKII-alpha were affected. Based on the combination of protein set enrichment and bioinformatic pathway analysis, we designed orthogonal SRM-assays which validated the abnormalities of proteins involved in synaptic long-term potentiation, myelination, and the ERK-signalling pathway in both brain regions. In contrast, increased levels of proteins involved in neurotransmitter metabolism and release were found only in the frontal cortex and abnormalities of proteins involved in the purinergic system were found exclusively in the hippocampus.

Conclusions: Taken together, this multi-platform profiling study has identified peripheral changes which are potentially linked to central alterations in synaptic plasticity and neuronal function associated with NMDAR-NR1 hypofunction. Therefore, the reported proteomic changes may be useful as translational biomarkers in human and rodent model drug discovery efforts.

No MeSH data available.


Related in: MedlinePlus

Volcano plots of group comparisons (10 NR1neo−/− vs. 10 wildtype mice) showing the adjusted significance P value (log2) versus fold change (log2). The plots indicate the most robust protein changes in the NR1neo−/−. Horizontal grey lines indicate an adjusted P value threshold of 0.05, vertical grey dotted lines indicate a fold-change threshold of 10%. Uniprot-identifiers indicate altered proteins. Significant proteins are quantified by at least two peptides. Full information can be found in Additional file 3: Table S3.
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Figure 1: Volcano plots of group comparisons (10 NR1neo−/− vs. 10 wildtype mice) showing the adjusted significance P value (log2) versus fold change (log2). The plots indicate the most robust protein changes in the NR1neo−/−. Horizontal grey lines indicate an adjusted P value threshold of 0.05, vertical grey dotted lines indicate a fold-change threshold of 10%. Uniprot-identifiers indicate altered proteins. Significant proteins are quantified by at least two peptides. Full information can be found in Additional file 3: Table S3.

Mentions: LC-MSE analysis resulted in the identification of 11,345 distinct peptides (563 proteins) in the frontal cortex and 14,775 distinct peptides (883 proteins) in the hippocampus after filtering the data using the criteria described in the Materials and Methods section. In the frontal cortex, 48 proteins were found to be significantly altered by more than 10% (Figure 1, Additional file 3: Table S3). In the hippocampus, 41 proteins showed significant changes using the same criteria.


Integrative proteomic analysis of the NMDA NR1 knockdown mouse model reveals effects on central and peripheral pathways associated with schizophrenia and autism spectrum disorders.

Wesseling H, Guest PC, Lee CM, Wong EH, Rahmoune H, Bahn S - Mol Autism (2014)

Volcano plots of group comparisons (10 NR1neo−/− vs. 10 wildtype mice) showing the adjusted significance P value (log2) versus fold change (log2). The plots indicate the most robust protein changes in the NR1neo−/−. Horizontal grey lines indicate an adjusted P value threshold of 0.05, vertical grey dotted lines indicate a fold-change threshold of 10%. Uniprot-identifiers indicate altered proteins. Significant proteins are quantified by at least two peptides. Full information can be found in Additional file 3: Table S3.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4109791&req=5

Figure 1: Volcano plots of group comparisons (10 NR1neo−/− vs. 10 wildtype mice) showing the adjusted significance P value (log2) versus fold change (log2). The plots indicate the most robust protein changes in the NR1neo−/−. Horizontal grey lines indicate an adjusted P value threshold of 0.05, vertical grey dotted lines indicate a fold-change threshold of 10%. Uniprot-identifiers indicate altered proteins. Significant proteins are quantified by at least two peptides. Full information can be found in Additional file 3: Table S3.
Mentions: LC-MSE analysis resulted in the identification of 11,345 distinct peptides (563 proteins) in the frontal cortex and 14,775 distinct peptides (883 proteins) in the hippocampus after filtering the data using the criteria described in the Materials and Methods section. In the frontal cortex, 48 proteins were found to be significantly altered by more than 10% (Figure 1, Additional file 3: Table S3). In the hippocampus, 41 proteins showed significant changes using the same criteria.

Bottom Line: The highest magnitude changes were found for neurotrophic factors (VEGFA, EGF, IGF-1), apolipoprotein A1, and fibrinogen.We also found decreased levels of several chemokines.In contrast, increased levels of proteins involved in neurotransmitter metabolism and release were found only in the frontal cortex and abnormalities of proteins involved in the purinergic system were found exclusively in the hippocampus.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QT, UK.

ABSTRACT

Background: Over the last decade, the transgenic N-methyl-D-aspartate receptor (NMDAR) NR1-knockdown mouse (NR1(neo-/-)) has been investigated as a glutamate hypofunction model for schizophrenia. Recent research has now revealed that the model also recapitulates cognitive and negative symptoms in the continuum of other psychiatric diseases, particularly autism spectrum disorders (ASD). As previous studies have mostly focussed on behavioural readouts, a molecular characterisation of this model will help to identify novel biomarkers or potential drug targets.

Methods: Here, we have used multiplex immunoassay analyses to investigate peripheral analyte alterations in serum of NR1(neo-/-) mice, as well as a combination of shotgun label-free liquid chromatography mass spectrometry, bioinformatic pathway analyses, and a shotgun-based 40-plex selected reaction monitoring (SRM) assay to investigate altered molecular pathways in the frontal cortex and hippocampus. All findings were cross compared to identify translatable findings between the brain and periphery.

Results: Multiplex immunoassay profiling led to identification of 29 analytes that were significantly altered in sera of NR1(neo-/-) mice. The highest magnitude changes were found for neurotrophic factors (VEGFA, EGF, IGF-1), apolipoprotein A1, and fibrinogen. We also found decreased levels of several chemokines. Following this, LC-MS(E) profiling led to identification of 48 significantly changed proteins in the frontal cortex and 41 in the hippocampus. In particular, MARCS, the mitochondrial pyruvate kinase, and CamKII-alpha were affected. Based on the combination of protein set enrichment and bioinformatic pathway analysis, we designed orthogonal SRM-assays which validated the abnormalities of proteins involved in synaptic long-term potentiation, myelination, and the ERK-signalling pathway in both brain regions. In contrast, increased levels of proteins involved in neurotransmitter metabolism and release were found only in the frontal cortex and abnormalities of proteins involved in the purinergic system were found exclusively in the hippocampus.

Conclusions: Taken together, this multi-platform profiling study has identified peripheral changes which are potentially linked to central alterations in synaptic plasticity and neuronal function associated with NMDAR-NR1 hypofunction. Therefore, the reported proteomic changes may be useful as translational biomarkers in human and rodent model drug discovery efforts.

No MeSH data available.


Related in: MedlinePlus