Mass Spectrometry-based Label-free Quantitative Proteomics To Study the Effect of 3PO Drug at Cellular Level

Human
endothelial cells (ECs) have been employed to monitor the
protein changes induced by [3-(3-pyridinyl)-1-(4-pyridinyl)-2-propen-1-one]
(3PO), a compound able to inhibit the glycolytic flux partially and
transiently and to reduce pathological angiogenesis in a variety of
disease models. Normal and TNFα induced inflamed ECs were incubated
with and without 3PO at a concentration (20 μM) able to inhibit
cell proliferation without cell death. At the end of the incubation
period, samples were submitted to the following steps: (a) whole protein
extraction, reduction, alkylation, and digestion by trypsin; (b) peptide
separation by nano-LC–MS/MS analysis using a high-resolution
mass spectrometer; (c) data analysis including protein identification,
quantification, and statistical analysis. An altered protein expression
profiling in combination with protein network analysis was employed
by using a mass spectrometry-based label-free quantification approach
to explore the underlying mechanisms of 3PO at cellular level.