Macrophages engulf apoptotic and primary necrotic thymocytes through similar phosphatidylserine‐dependent mechanisms

Abstract

One of the major roles of professional phagocytes is the removal of dead cells in the body. We know less about the clearance of necrotic cells than apoptotic cell phagocytosis, despite the fact that both types of dead cells need to be cleared together and necrotic cells appear often in pathological settings. In the present study, we examined phagocytosis of heat‐ or H 2 O 2 ‐killed necrotic and apoptotic thymocytes by mouse bone marrow‐derived macrophages ( BMDM s) in vitro and found that the two cell types are engulfed at equal efficiency and compete with each other when added together to BMDM s. Phagocytosis of both apoptotic and necrotic thymocytes was decreased by (a) blocking phosphatidylserine on the surface of dying cells; (b) inhibition of Mer tyrosine kinase, Tim‐4, integrin β3 receptor signaling, or Ras‐related C3 botulinum toxin substrate 1 activity; or (c) using BMDM s deficient for transglutaminase 2. Stimulation of liver X, retinoid X, retinoic acid or glucocorticoid nuclear receptors in BMDM s enhanced not only apoptotic, but also necrotic cell uptake. Electron microscopic analysis of the engulfment process revealed that the morphology of phagosomes and the phagocytic cup formed during the uptake of dying thymocytes is similar for apoptotic and necrotic cells. Our data indicate that apoptotic and necrotic cells are cleared via the same mechanisms, and removal of necrotic cells in vivo can be facilitated by molecules known to enhance the uptake of apoptotic cells.

Key differences between olfactory ensheathing cells and Schwann cells regarding phagocytosis of necrotic cells: implications for transplantation therapies

“…We verified that glial recognition of necrotic bodies occurred mainly via binding to phosphatidylserine (PS); competitive blocking of PS by Annexin V conjugate resulted in decreased NB uptake by 40-50% in all cell types, which is similar to previous reported data for macrophage internalization of cargo 30,37,39 . However, the molecular alterations at the plasma membrane by necrotic cells remain to be characterised.…”

“…In the current study, we investigated whether two types of peripheral glia, olfactory ensheathing cells (OECs) and Schwann cells (SCs), could recognize and engulf NBs. We compared this activity in the glia to that of macrophages, which are efficient at recognizing and phagocytosing NBs 30,[37][38][39] .…”

“…To create NBs, we induced necrosis in a fibroblast cell line (mouse McCoy B cells) using heat treatment 30,37,38 . To determine the optimal incubation time for necrosis induction, McCoy B cells were heated to 55 °C for varying time periods (5, 10, 15, 20, 25 and 30 min) followed by staining with Trypan blue and DRAQ7 (which both are membrane-impermeable nuclear stains, thus staining necrotic/dead cells).…”

“…We then investigated the mechanism by which glia recognize necrotic targets. Both "professional" and "non-professional" phagocytes have been shown to phagocytose necrotic cells by recognition of PS displayed on dying cells 29,30,37,39,49,50 . Hence, prior to exposing macrophages, OECs and SCs to NBs, we incubated the NBs with the PS binding partner Annexin V that binds to PS and blocks PS recognition.…”

Uptake of osteoblast-derived extracellular vesicles promotes the differentiation of osteoclasts in the zebrafish scale

“…Exosomes and MVs are shed from a variety of cell types and have been implicated in cell-cell communication 34 . ABs are formed during the apoptotic process and are then engulfed by macrophages and other immune cells 35,36 . Since exosomes are recognized as very tiny vesicles (<100 nm) 37,38 , it is likely that the OB-derived EVs isolated by FCM and visualized by confocal microscopy are mainly MVs and/or ABs rather than exosomes.…”