The effects of Phorbol 12-myristate 13-acetate concentration on the expression of miR-155 and miR-125b and their macrophage function-related genes in the U937 cell line.

Abstract

The phorbol 12-myristate 13-acetate (PMA)-induced U937 cell line has been widely used as an in vitro model for studying the functions of human macrophages. However, there are several concentrations of PMA commonly used to drive the differentiation of monocytic cell line to macrophage. Also, the expression of microRNA-155 (miR-155) and miR-125b in PMA-treated human monocytic cell line has not yet been reported. The five usual concentrations of PMA for stimulating macrophage differentiation are 10, 25, 50, 100, and 200 nM. In this study we compared the expression levels of miR-155, miR-125b and their related genes involved in macrophage functions in U937-derived cells after treatment with those five concentrations. The morphological study results showed that the five concentrations of PMA could induce macrophage differentiation in a similar manner. Moreover, cell proliferation and viability were not significantly different among these five conditions excepted the lower cell viability at 200 nM of PMA treatment. The five concentrations of PMA could upregulate the expression of miR-155 and miR-125b and increase the phagocytic activity of U937-derived cells in dose-reversal manner. The upregulation of miR-155 was correlated with increased expression levels of TNFα and decreased expression levels of BACH1 and CEBPβ, while the reduction of IRF4 was correlated with increased expression levels of miR-125b. Our study found that PMA could stimulate macrophage differentiation in a broad range of concentrations, however, the lower concentration could upregulate the higher expression of both miR-155 and miR-125b, and that correlated with the phagocytic functional activity of U937-derived macrophages.