Gene and protein expression profiling of human cerebral endothelial cells activated with tumor necrosis factor-alpha

An increase in permeability of the blood-brain barrier is a critical event in the pathophysiological process of multiple sclerosis and other neurodegenerative diseases. Tumor necrosis factor alpha (TNFalpha) is known to play a crucial role in this process and is a powerful activator of endothelial cell inflammatory responses. Although many reports describe effects of TNFalpha activation in endothelial cells, the molecular mechanisms specific for activation of cerebral endothelial cells remains unclear. The objective of this study was to identify potential pharmaceutical targets for the treatment of multiple sclerosis using molecular profiling techniques. Gene expression measurements (Affymetrix Hu6800 oligonucleotide arrays) and proteomics (two-dimensional gel electrophoresis and mass spectrometry) were applied to analyze early alterations in human cerebral endothelial cells (HCEC) activated by TNFalpha. Human umbilical vein endothelial cells (HUVEC) were used as the reference system. The results presented show that HCEC and HUVEC respond similarly with respect to cell adhesion molecules, chemotaxis, apoptosis and oxidative stress molecules. However, nuclear factors NFkB1 and NFkB2, plasminogen activator inhibitor 1 and cofilin 1 are examples of cerebral specific responses. Our results indicate involvements of the urokinase plasminogen activator system and cytoskeletal rearrangements unique to TNFalpha activation of cerebral endothelial cells

Effects of natalizumab treatment on the cerebrospinal fluid proteome of multiple sclerosis patients

Natalizumab is a very effective, relatively new drug for the treatment of relapsing remitting multiple sclerosis. Inflammatory and neurodegenerative processes in the central nervous system are presumed to cause adverse effects during the course of this disease. To monitor the effects of natalizumab treatment on the cerebrospinal fluid (CSF) proteome of patients, CSF samples were taken from patients before commencing treatment as well as after 1 year of treatment. Profiling proteomics experiments using electrospray Orbitrap mass spectrometry and pair wise comparison of patients before and after 1 year of natalizumab treatment revealed a number of candidate biomarkers that were significantly differentially abundant between the before and after treatment groups. Three proteins were subsequently validated using selected reaction monitoring (SRM) in a new, independent sample set. All three proteins, Ig mu chain C region and haptoglobin, both known inflammation-related proteins, as well as Chitinase-3-like protein 1, were confirmed by SRM to be significantly lower abundant in CSF of multiple sclerosis patients after 1 year of natalizumab treatment. The findings for Chitinase-3-like protein 1, a presumed biomarker for more rapid progression from a first clinically isolated syndrome to clinically definite multiple sclerosis, was further confirmed by ELISA measurements