Molecular assessment of the phenotypic changes associated with smooth muscle cells using two-dimension...

Phenotypic modulation of arterial smooth muscle cells (SMC) is essential for the evolution of atheromatosis and restenosis after angioplasty (1). During these pathological phenomena, SMC express numerous genes such as those responsible for cell migration and proliferation (2). The identification of genes differentially expressed is obviously important for the understanding of the mechanisms leading to restenosis and atheromatous plaque development and may be a key step in the planning of new therapeutic approaches. Among the different techniques available for the identification of differentially expressed genes, in this chapter we describe the two-dimensional analytical gel electrophoresis (2-D PAGE) protein expression analysis (3). This method offers the advantage of producing protein maps of defined SMC phenotypes, which can be used as references for such applications as SMC phenotype comparison and drug effect analysis (4,5). In the last decade, this classical biochemical technique has benefited from modifications improving the reproducibility and the quality of 2-D gels. One of the most important contributions was the introduction of immobilized pH gradients (IPGs) as the first dimension (3). In comparison to the conventional carrier ampholyte, they ensure reproducible gradients that are insensitive to disturbances from sample components. Moreover, recent developments concerning protein databases as well as computer systems facilitate spot detection and protein identification (6).