RESUMO
Phorbol-12-myristate-13-acetate (PMA) and 1,25-dihydroxyvitamin D3 (VD3) are stimuli commonly used to induce macrophage differentiation in monocytic cell lines, but the extent of differentiation in comparison to primary tissue macrophages is unclear. Here, we examine the morphological/phenotypic markers associated with differentiation of U937 cells into monocytes/macrophages, in response to PMA or VD3 treatment. PMA stimulus but not with VD3, induced changes in cell morphology indicative of differentiation, but did not show differentiation comparable to monocyte-derive macrophage (MDM). The cells treated with PMA+VD3 for 2 days (d) acquired morphological/phenotypic features similar to those acquired by monocytes. In contrast, U937 cells treated for 2d with PMA and VD3 followed by 6d of resting in culture without PMA but in the presence of VD3 acquired morphological and phenotypic markers similar to those of MDM; i.e. reduced nucleus/cytoplasmic ratio, high auto-fluorescence and cytoplasmic complexity. Furthermore, low expression of CD14/TLR2 and high expression of CD68/CD86 were observed. In conclusion, our results indicate a synergistic effect between PMA and VD3 in U937 cells differentiation into both monocytes or macrophages and we propose a modified PMA differentiation protocol to enhance monocyte/macrophage differentiation of U937 cells.
