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. 2016 Mar 15;25(9):661–673. doi: 10.1089/scd.2015.0344

FIG. 3.

FIG. 3.

Pop#8 exclusive in vitro generation of MPCs. After 5 days in MPC-selective culture conditions (DMEM/10%PhABS) Pop#8 generated MPC-like cells characterized by the peculiar fried egg-shaped morphology (A). CD3/20/14/CD66+ cells were sorted in parallel, as control population (B). Immunofluorescence analysis of Pop#8-derived MPCs, revealed podosome-like actin organization (red in C) and intense positivity to nestin (green in C). The MPC phenotype was confirmed by flow cytometry (D). After 7 days of culture under mesengenic-differentiating conditions (MesenPRO® RS medium), Pop#8-derived MPCs gave rise to confluent cells showing MSC phenotype (E) and morphology (F). To confirm the mesenchymal nature of Pop#8 MPC-derived MSCs, cultures were further exposed to adipogenic or osteogenic stimuli. Lipid droplet (red in G) accumulation and calcium deposition (green in G) were detected only in terminally differentiated cultures (G, upper panels). No fluorescence signals were detected in cultures maintained in MesenPRO RS medium (G) (lower panels). DMEM, Dulbecco's modified Eagle's medium; PhABS, pooled human AB-type serum; RS, reduced serum.