Proteome analysis of cerebrospinal fluid in patients with Multiple System Atrophy (MSA)

Background: Multiple system atrophy (MSA) is a degenerative neurological disorder which belongs to the synucleinopathies. As a Parkinson plus syndrome, MSA can be sub-grouped into a cerebellar subtype (MSA-c), a parkinsonian subtype (MSA-p) and an autonomic subtype (MSA-a), dependent on the brain area most affected. Differential diagnosis of MSA to other parkinsonian syndromes frequently is difficult especially in early disease stages. In order to identify candidate biomarkers to differentiate between MSA subtypes we analysed the cerebrospinal fluid (CSF) proteome profile of patients with MSA and controls.

Methods: CSF samples from patients with MSA-c (n=9) and MSA-p (n=7) were compared to age matched normal controls (n=16). In the first step CSF samples were pooled and analysed by 2-D Fluorescence Difference In Gel Electrophoresis (2D DIGE) to screen for potential candidate proteins. Proteins, which showed a significant difference in its spot volume, were analysed with MALDI-TOF mass spectrometry and identified by searching with the non-redundant protein database National Centre for Biotechnology Information (NCBI). In the second step we used individual CSF samples to validate the candidate proteins using immunoblot as a different method.

Results: 2-D DIGE revealed altered expression of 9 interesting proteins, including Zinc-Alpha-Glycoprotein (up-regulated in MSA-p compared to the reference group), Nebulin (down-regulated in MSA-p and MSA-c), alpha-1-Antitrypsin precursor (up-regulated in MSA-p and down-regulated in MSA-c) and Gelsolin (up-regulated in MSA-p).

The immunoblot analysis confirmed Nebulin to be down-regulated in both, MSA-c and MSA-p patients by 21 and 61% compared to the reference group. Gelsolin was found to be up regulated in 3 of 4 cases (between 35% and +101%) and alpha-1-Antitrypsin differed between -3% and +19%. In MSA-p patients Zinc-Alpha-Glycoprotein was up-regulated between +55% and +110%.

Conclusions: We believe the results of proteomic research will give us a useful tool to distinguish between MSA and clinically similar diseases by means of their CSF proteome profile. Particularly Nebulin, Zinc-Alpha-Glycoprotein and Gelsolin are of interest, as they were not yet linked to MSA. Further validation of these results by quantitative immunoassays such as sandwich enzyme immunoassay (ELISA) technique is warranted.