Minocycline Effects on the Cerebrospinal Fluid Proteome of Experimental Autoimmune Encephalomyelitis Rats

To identify response biomarkers for pharmaceutical treatment
of
multiple sclerosis, we induced experimental autoimmune encephalomyelitis
(EAE) in rats and treated symptomatic animals with minocycline. Cerebrospinal
fluid (CSF) samples were collected 14 days after EAE induction at
the peak of neurological symptoms, and proteomics analysis was performed
using nano-LC-Orbitrap mass spectrometry. Additionally, the minocycline
concentration in CSF was determined using quantitative matrix-assisted
laser desorption/ionization-triple-quadrupole tandem mass spectrometry
(MALDI-MS/MS) in the selected reaction monitoring (SRM) mode. Fifty
percent of the minocycline-treated EAE animals did not show neurological
symptoms on day 14 (“responders”), while the other half
displayed neurological symptoms (“nonresponders”), indicating
that minocycline delayed disease onset and attenuated disease severity
in some, but not all, animals. Neither CSF nor plasma minocycline
concentrations correlated with the onset of symptoms or disease severity.
Analysis of the proteomics data resulted in a list of 20 differentially
abundant proteins between the untreated animals and the responder
group of animals. Two of these proteins, complement C3 and carboxypeptidase
B2, were validated by quantitative LC-MS/MS in the SRM mode. Differences
in the CSF proteome between untreated EAE animals and minocycline-treated
responders were similar to the differences between minocycline-treated
responders and nonresponders (70% overlap). Six proteins that remained
unchanged in the minocycline-treated animals but were elevated in
untreated EAE animals may be related to the mechanism of action of
minocycline.