Research Article

Micro- and Macrostructured PLGA/Gelatin Scaffolds Promote Early Cardiogenic Commitment of Human Mesenchymal Stem Cells In Vitro

Figure 7

Stem cell colonization, growth, and elongation on monolayer and bilayer microstructured PLGA/gelatin scaffolds. (a) Stem cells stained with Calcein-AM and imaged using TPEF microscopy (green) visualized on a monolayer microstructured PLGA/gelatin scaffold imaged using CARS microscopy (blue) after 8 hours from seeding. Red arrows indicate a few hMSCs aligned in parallel inside the scaffold lanes. (b) Representative confocal microscopy images of living hMSCs cultured for either 1 or 15 days on bilayer scaffolds. Superposition of Calcein-AM (green) and bright field microscopy is shown. Magnification 20x, scale bar 50 μm. (c) Representative confocal microscopy image showing cytoskeletal organization of hMSCs cultured for 15 days on a bilayer scaffold. Actin filaments are stained with phalloidin in red, cytoplasm with Cell Mask Green, and nuclei with Hoechst in blue. Magnification 63x, scale bar 20 μm. (d) 3D reconstruction of a representative image showing hMSCs cultured for 24 h on a monolayer scaffold. Cells are stained with Calcein-AM. Magnification 20x, scale bar 100 μm. (e) Cell proliferation rates after 4, 8, 12, and 15 days of culture on bilayer scaffolds. Values were normalized on cell adhesion quantified at 24 hr. (f) Mean cell lengths of hMSCs cultured on monolayer and bilayer scaffolds. Elongation measurements were performed on living cells stained with Calcein-AM at 2, 4, and 8 days of culture. Lengths were not statistically different from the 2D control.