0-TESTO COMPLETO.pdf - Fondazione Santa Lucia

FISM.9 – Proteomics and metabolomics investigations for the identification of multiple…
experimental model of the disease, Experimental Autoimmune Encephalomyelitis
(EAE), disease can be induced by transferring T cells specific for
myelin antigens; moreover, the histopathological hallmark of MS consists of
the demyelinating lesion, characterized by a perivenular cellular infiltrate,
composed mainly of activated T lymphocytes and reactive macrophages, thus
underlining the substantial role of autoreactive T cells in the inflammatory
process that leads to damage of the myelin sheath.
Autoreactive T cells are also present in the periphery of healthy individuals,
but their destructive activity seems to be kept in check by regulatory T
cells. It is well known that CD4+, CD8+ and gd + T cells can be functionally
divided in naïve and memory cells, and each subpopulation shows unique
autoimmune properties. As naïve cells migrate preferentially into lymphoid
organs, we will focus our attention on memory cells. Memory cells have been
divided in two subsets: “ central ” memory cells that migrate in lymphoid
organs and in sites of inflammation, and “ effector ” memory cells that migrate
exclusively in sites of inflammation.
The interactions between the different components of the immune system
are indeed very complex, and new technologies have revealed that cellular
populations which apparently seem homogeneous are in fact composed of
several subpopulations with different phenotypes and functions.
The large genomic projects are opening the route to the development of proteomics
investigation to possibly highlight new molecular markers diagnostic
and/or prognostic of multiple sclerosis. Proteomics is a discipline which allows
an open, holistic, investigation of the polypeptides contained in a sample. This is
achieved by combining high resolution protein separations, mass spectrometry
investigation and bioinformatics analysis of the collected evidences.
Direct mass spectrometry approach of complex mixtures, is today, the last
challenge for gel-free analysis of disease biomarkers. In particular, Linear
Matrix-Assisted Laser Desorption-Ionization Time-of-Flight mass spectrometry
(MALDI-TOF-MS), is a promising technique for clinical applications given
its high sensitivity, accuracy and resolution in discriminating the low molecular
weight protein profiles [D’Aguanno et al. 2007; Del Boccio et al. 2007].
Such an approach consists in comparative analysis of protein expression profiles
in clinical groups in order to identify differentially expressed proteins
that may represent new molecular markers.
Serum patterns that distinguish between disease and normal subject with
remarkable accuracy have been reported for several type of neurodegenerative
disorders and other diseases [Biroccio et al. 2006]. This direct mass spectrometry
approach yield to a comprehensive profiles of peptides and proteins in
biological fluids without the need to separate the complex mixture, thus
allowing reduced amount of sample for high throughput studies. However,
the presently available commercial solutions to this technique (SELDI-TOF-
MS) have shown several problems mainly due to the high cost of activated
chips and of the poor analytical performances, which are currently strongly
limiting the significance of the acquired data. Our group has been actively
involved in establishing protocols to ensure precision and accuracy within the
IFCC (International Federation in Clinical Chemistry) and Clinical and Labo-
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