The Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disease characterized by segmental premature aging. Applying a two-dimensional chromatographic proteomic approach, the 20 Protein Fractionation System (PF2D), we identified 30 differentially expressed proteins in cultured HGPS fibroblasts. We categorized them into five groups: methylation, calcium ion binding, cytoskeleton, duplication, and regulation of apoptosis. Among these 30 proteins, 23 were down-regulated, while seven were up-regulated in HGPS fibroblasts as compared to normal fibroblasts. Three differentially expressed cytoskeleton proteins, vimentin, actin, and tubulin, were validated via Western blotting and characterized by immunostaining that revealed densely thickened bundles and irregular structures. Furthermore in the HGPS cells, the cell cycle G1 phase was elongated and the concentration of free cytosolic calcium was increased, suggesting intracellular retention of calcium. The results that we obtained have implications for understanding the aging process. (C) 2011 Elsevier Inc. All rights reserved.