Phorbol myristate acetate induces differentiation of THP-1 cells in a nitric oxide-dependent manner.

Abstract

INTRODUCTION\nTHP-1 cells, a human leukemia monocytic cell line, differentiated by phorbol myristate acetate (PMA) are widely used as surrogate of human macrophages. Differentiated THP-1 cells acquire macrophage-like characteristics including more adherence and altered cell function. Nitric oxide (NO), an intracellular messenger, is critical in regulating cell differentiation. Here we elucidated whether NO relates to PMA-induced monocyte-to-macrophage differentiation of THP-1 cells. The mutual regulation of calcium and NO was also investigated.\n\n\nMATERIAL & METHODS\nTHP-1 cells were incubated with PMA for 24 hours, followed by assay of adherence, morphological change, migration or IL-1β release. L-NG-Nitroarginine methyl ester (L-NAME, a nitric oxide synthase inhibitor) or BAPTA-AM (a calcium chelator) was added before PMA stimulation, and levels of calcium and NO were measured. Furthermore, a selective inhibitor of inducible nitric oxide synthase (iNOS) activity was employed to study the role of iNOS.\n\n\nRESULTS\nand Discussion: Effects of PMA on upregulation of adherence, lipopolysaccharide-triggered IL-1β, and migration ability of THP-1 cells were consistent with NO concentrations. Both L-NAME and BAPTA-AM mitigated effects of PMA on THP-1 cells differentiation. BAPTA-AM decreased levels of NO, while L-NAME had no effect on calcium levels. Of note, inhibition of iNOS activity decreased PMA-triggered upregulation of NO.\n\n\nCONCLUSION\nPMA induced differentiation of THP-1 cells partially in a NO-dependent manner. The calcium signaling may mediate PMA-triggered upregulation of NO.