Functional Proteome analysis of the molecular mechanism of (myo)fibroblast differentiation

Abstract: Tissue remodeling is observed in several physiological conditions such as embryogenesis, senescence, wound healing and pregnancy, as well as in several pathological conditions such as fibrosis and tumorgenesis. Fibroblasts play a key role in this tissue remodeling due to their capability of altering the turnover of the extracellular matrix. During the remodeling process, the fibroblasts are believed to differentiate into fibroblast-smooth muscle-like cells called myofibroblasts. This differentiation process is mediated by a well-orchestrated interplay between the extracellular matrix and growth factors such as platelet-derived growth factor (PDGF) and transforming growth factor-beta (TGF-beta ). In this thesis, the overall aim was to investigate the mechanism for fibroblast activation by employing various proteomic technologies. We present a mechanistic explanation as to how TGF-beta induces alternative splicing of a number of extracellular matrix components that have been found to be essential for the differentiation process. This thesis also shows how specific carbohydrate moieties can modulate the PDGF-induced cellular response. By isolating fibroblast populations from different bronchial locations, the in vitro results could be compared with the bronchial-derived fibroblast populations by proteome profiling. To achieve these goals, application of a number of proteomic tools was necessary such as two-dimensional gel electrophoresis, multidimensional chromatography and mass spectrometers utilizing both laser desorption and electrospray ionization. A database was also constructed to store and analyze the data derived from the studies. In summary, the results showed that myofibroblast differentiation was dependent upon a coordinate interplay between the extracellular matrix components and growth factors. These growth factors act locally on the fibroblasts to induce a tissue remodeling similar to that observed in diseases such as asthma.