Role of lipoxin A4 in the cell‐to‐cell interaction between all‐trans retinoic acid‐treated acute promyelocytic leukemic cells and alveolar macrophages

Abstract

Differentiation therapy with all-trans retinoic acid (ATRA) has been used successfully to treat acute promyelocytic leukemia (APL), but such treatment also causes differentiation syndrome (DS) by inducing APL cell infiltration into alveolar spaces. The mechanism underlying the clearance of infiltrated APL cells has not been investigated in detail. Lipoxin A(4) (LXA(4)) is an important anti-inflammatory mediator during the resolution of inflammation. In this study, the role of LXA(4) in the cell-cell interaction between alveolar macrophages (AMφ; NR8383 cells) and APL NB4 cells was investigated and found that conditioned medium (CM) harvested from ATRA-treated NR8383 (ATRA-NR8383) cells was able to induce the transmigration of ATRA-NB4 cells. However, the pro-migratory activity of CM was attenuated progressively when ATRA-NR8383 cells were co-cultured with increased cell dosages of apoptotic NB4 cells. A significantly higher amount of LXA(4) was released into the CM by ATRA-NR8383 cells when they were co-cultured with apoptotic ATRA-NB4 cells. Expression of a receptor for LXA(4) (ALX/FPR2) was enhanced in both ATRA-NB4 cells and ATRA-NR8383 cells. Exogenous LXA(4) treatment was able to inhibit the transmigration of ATRA-NB4 cells and induce the phagocytic clearance of apoptotic cells by ATRA-NR8383 cells. The anti-migratory activity of exogenous LXA(4) was attenuated by pre-treating ATRA-NB4 cells with an ALX/FPR2 inhibitor. We conclude that AMφ-derived LXA(4) plays an important role in the interaction between AMφ and APL cells and that this contributes to clearance of apoptotic APL cells.

CX3CL1(+) Microparticles Mediate the Chemoattraction of Alveolar Macrophages toward Apoptotic Acute Promyelocytic Leukemic Cells

“…The non-permeablizied ATRA-NB4 cells and NR8383 cells were stained with either rabbit CX3CL1 polyclonal antibody (MAB365; R&D System, Minneapolis, USA) and rabbit CX3CR1 polyclonal antibody (AF5825; R&D System, Minneapolis, USA), respectively, before analysis by FAC Scan as reported previously [4,15]. …”

“…Non-ingested cells were removed after washing with PBS. After myeloperoxidase staining, the number of NR8383 cells containing ingested NB4 cells was counted by light microscopy [4]. The results were expressed as phagocytosis index indicating a fold increase relative to the percentage of NR8383 cells engulfing UV-untreated ATRA-NB4 cells as control.…”

“…Mouse polyclonal antibodies against human CX3CL1 (MAB3561; R&D Systems, Minneapolis, USA) and monoclonal antibody against Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (Sigma, St. Louis, Missouri, USA) were used for the Western blotting. The results were first normalized against the GAPDH present in the same cells and then expressed as a fold increase relative to the UV un-irradiated ATRA-NB4 cells as control [4,15]. …”

“…NR8383 cells (CRL-2192; ATTC, Manassas, VA) are a well-characterized, semi-adherent AMϕ cell line that is derived from normal Sprague-Dawley (SD) rat lung and can be extrapolated to freshly derived AMϕ of various species[11]. Both NB4 cells and NR8383 cells were cultured in RPMI-1640 medium (GIBCO, Grand Island, NY) and in Ham's F12K medium, respectively, as described previously [4]. ATRA treatment can induce NB4 cells into the process of granulocytic differentiation [1].…”

“…Several chemokines are involved in the pathogenesis of DS, such as interleukin-8 and monocyte chemotactic protein -1 [1,2,3]. Recently, we reported that the alveolar macrophage (AMϕ) is able to engulf apoptotic ATRA-APL cells; this may contribute to the clearance of apoptotic APL cells in the alveolar space during the resolution phase of DS [4]. However, the mechanism underlying the chemoattraction of AMϕ toward apoptotic APL cells has not been investigated in detail.…”