Abstract
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. DisclosuresNo relevant conflicts of interest to declare.
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