CX3CL1(+) Microparticles-Induced MFG-E8 Enhances Apoptotic Cell Clearance by Alveolar Macrophages

Abstract

During the resolution phase of acute lung injury, apoptotic cells release CX3CL1 as a “find-me” signal to attract alveolar macrophage transmigration toward apoptotic cells for phagocytosis. However, it is still not clear whether CX3CL1 has pro-phagocytic activity on alveolar macrophage. In this study, we investigated the role of apoptotic NB4 cells-derived CX3CL1(+) microparticles (apo-MP) on the phagocytic activity of NR8383 cells. We demonstrate that exogenous CX3CL1 and apo-MP enhanced the phagocytic activity of NR8383 cells in a CX3 CR1-dependent manner. The apo-MP-enhanced phagocytic activity on NR8383 was attenuated when apo-MP and NR8383 cells were pre-treated with anti-CX3CL1 antibodies and anti-CX3CR1 antibody, respectively, before incubating both for phagocytic assay. Further studies demonstrate that exogenous CX3CL1 and apo-MP also enhanced NR8383 cells in their surface expression and release of MFG-E8 in a CX3CR1 dependent manner. The enhanced phagocytic activity of CX3CL1-treated NR8383 cells was attenuated when NR8383 cells were pre-treated with an anti-MFG-E8 antibody before CX3CL1 treatment. We conclude that apoptotic cell-derived CX3CL1(+) microparticles enhance the phagocytic activity of NR8383 cells by up-regulating their MFG-E8 as a bridge molecule, and these contribute to the formation of phagocytic synapses between apoptotic cells and alveolar macrophages for the subsequent phagocytic clearance of apoptotic cells.

Apoptotic Cell-Derived CD14(+) Microparticles Promote the Phagocytic Activity of Neutrophilic Precursor Cells in the Phagocytosis of Apoptotic Cells

“…Regarding this, we demonstrate that (a) CD14(+) apo-MP was released by apoptotic ATRA-NB4 cells in an idarubicin-dosage dependent manner, (b) membranous CD14 expression of viable ATRA-NB4 cells was enhanced after incubation with apo-MP harvested from apoptotic ATRA-NB4 cells, and (c) the apo-MP-enhanced phago-cytic activity of viable ATRA-NB4 cells was significantly attenuated when that apo-MP was pre-treated with an anti-CD14 antibody before incubation with viable cells for phagocytic assay (Figure 5D). Our data imply that membranous CD14 on apo-MP, derived from apoptotic ATRA-NB4 cells, was transferred to the membrane of recipient viable ATRA-NB4 cells via adhesion and/or a fusing mechanism [12]. Besides CD14, apoptotic ATRA-NB4 cell-derived apo-MP also enhance MFG-E8 expression on recipient viable ATRA-NB4 cells via CX3CL1-CX3CR1 axis to promote the binding between MFG-E8 and integrin ανβ3/5, resulting in the induction of STAT3-mediated SOCS3 activation to inhibit both NF-κB signaling pathway and pro-inflammatory cytokine production, and this contributes to the phagocytosis of apoptotic cells without inducing inflammation [49].…”

“…Thereafter, we address the crucial role of CD14(+)apo-MP in the phagocytosis of apoptotic cells by viable ATRA-NB4 cells. We have reported recently that apoptotic ATRA-NB4 cells release CX3CL1(+)apo-MP to (a) induce both viable ATRA-NB4 cells and alveolar macrophages transmigration toward apoptotic cells and (b) upregulate the MFG-E8 expression on alveolar macrophages as a bridge molecule to promote the binding between phosphatidylserine and phagocytic receptors (integrin αvβ3/5) on apoptotic cells and macrophages, respectively, for subsequent phagocytosis [11][12][13]. In this study, we provide the novel finding that apoptotic ATRA-NB4 cells release CD14(+) apo-MP to promote the recipient viable ATRA-NB4 cells in the early recognition and tethering of apoptotic cells for subsequent phagocytosis.…”

“…The surface of MPs carries antigens and adhesion molecules from their original cell membrane, and these are able to mediate intercellular cross-talk by transferring receptors, antigens, and cytokines from donor to recipient cells [9]. Recently, we have reported that MPs play a crucial role in the cell-cell interaction in the phagocytosis of apoptotic cells via macrophages during the resolution phase of inflammation [11][12][13].…”

“…However, the role of CD14 in the phagocytic activity of NPC has been rarely studied. Recently, we and others reported that CX3CL1 on microparticles (MPs) derived from apoptotic cells (apo-MP) acts as a "find-me" signal to attract macrophage transmigration toward apoptotic cells and to promote the phagocytic activity of macrophages, indicating the crucial role of apo-MP in the cell-cell interaction during the process of the efferocytic clearance of apoptotic cells [11,12,18]. However, it is still not clear whether CD14(+)apo-MP, derived from apoptotic NPC, plays any role in the phagocytosis of apoptotic cells.…”

Chemokines and phosphatidylserine: New binding partners for apoptotic cell clearance

“…For instance, S1P has been reported to promote apoptotic cell clearance indirectly in vivo by activating erythropoietin signaling, which upregulates the expression of both scavenger receptors and bridging molecules by macrophages ( Luo et al, 2016 ). Furthermore, CX3CL1 is known to upregulate MFG-E8, a PS-binding bridging molecule important for phagocytosis, in microglia and macrophages ( Hanayama et al, 2004 ; Miksa et al, 2007 ; Tsai et al, 2021 ). Thus, the precise find-me signals that promote phagocyte migration for efferocytosis in vivo require further investigation and definition.…”