CD14 Mediates Phagocytic Activity during the Granulocytic Differentiation Process in Acute Promyelocytic Leukemia Cells

All-trans retinoic acid (ATRA) can induce acute promyelocytic leukemia (APL) cells differentiation into mature granulocytes. CD14 and Toll-like receptor 4 (TLR-4) play an important role in the phagocytic activity of macrophage, however, their role during granulopoiesis is still unclear. In this study, we determined the role of CD14/TLR-4 in the development of phagocytic activity in NB4 APL cells after induction into the process of granulocytic differentiation by ATRA. Flow cytometry analysis demonstrate that, during ATRA treatment for 6 days, the phagocytic activity of NB4 cells in engulfing either fluorescein-latex beads or idarubicin-induced apoptotic cells increased in a time-dependent manner, and the level of CD14 expression on NB4 cells was also significantly increased in a time dependent manner, though its level was only minimally expressed in ATRA-untreated NB4 cells. However, TLR-4 was constitutionally expressed in ATRA-untreated cells and its level did not changed significantly during the first 5 days of ATRA treatment. Further study demonstrates that the phagocytic activity of ATRA-NB4 cells was significantly inhibited by pre-treating cells with antibodies specific to either CD14 or TLR-4 before phagocytosis assay. In exploring the role of CD14/TLR4 associated signal transduction mediators, NF-κB and IRF-3, we further demonstrate that the phagocytic activity of ATRA-NB4 cells in engulfing beads was significantly inhibited when cells were pretreated with either a NF-κB inhibitor (BAY 11-7082) or an IRF-3 inhibitor (SP600125). However, this activity in engulfing apoptotic cells was only significantly inhibited by pretreatment with BAY11-7082, but not by pre-treatment with SP600125. Finally, our results indicate that the level of CD14(+) microparticles (MPs) released by ATRA-NB4 cells was significantly enhanced when those cells were induced into the process of apoptosis by pre-treatment with idarubicin. Moreover, by incubation with MPs derived from apoptotic ATRA-NB4 cells, the phagocytic activity of living ATRA-NB4 cells in engulfing apoptotic cells was significantly enhanced, and this phagocytic activity was also significantly inhibited by pre-treating MPs with antibody specific to CD14 before phagocytic assay. We conclude that CD14 contributes to the phagocytic activity of APL cells during the process of granulocytic differentiation.