Neuropilin 1 Mediates Keratinocyte Growth Factor Signaling in Adipose-Derived Stem Cells: Potential Involvement in Adipogenesis

Figure 1

Phenotypical characterization of ASCs. (a) Representative phase-contrast photomicrograph of ASCs, showing homogeneous, spindle-shaped morphology (A) and immunofluorescence analysis showing positive staining for the mesenchymal antigens CD29 (B) and CD166 (C) and negative staining for the hematopoietic CD34 antigen (D). Nuclei (blue) were visualized with 4,6-diamidino-2-phenylindole (DAPI). Scale bars: 100 μm. (b) Flow cytometric analysis of ASCs. Cells were stained with monoclonal antibodies directed against CD29, C44, CD90, CD166, CD73, CD105, CD34, and CD45. Dark grey areas represent patterns obtained with antibodies against the indicated markers, whereas light grey lines represent the isotype-matched monoclonal antibody that served as a control. Each panel reported the percentage of positive cells for the corresponding marker. (c) Multilineage differentiation of ASCs. Adipogenic differentiation was assessed by positive staining with oil red O (A) and by positive immunofluorescence analysis for the adipocyte-specific fatty acid binding protein 4 (FABP4) (B). Osteogenic differentiation was demonstrated by positive staining for alizarin red (C) and positive immunofluorescence reactivity to osteocalcin (D). Chondrogenic differentiation was assessed by positive staining for Alcian Blue (E) and by positive immunofluorescence with anti-aggrecan antibodies (F). (d) Immunofluorescence analysis of ASCs showing positive staining for the mesenchymal marker vimentin (A) and negative staining for the epithelial marker cytokeratin 14 (K14) (B). Scale bars: 100 μm. (e) Expression of vimentin and K14 assessed by Western blot analysis on ASC whole cell lysates. HFs and MCF-7 cells were used as controls for mesenchymal and epithelial lineage, respectively. Western blot with anti-tubulin antibody served as loading control. The images are representative of at least three independent experiments.

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