CD31 delays phagocyte membrane repolarization to promote efficient binding of apoptotic cells.

Homophilic ligation of CD31, a member of the Ig superfamily of adhesion receptors, promotes macrophage clearance of apoptotic leukocytes by a mechanism hitherto not described. In studying CD31-dependent regulation of beta1-integrin binding of fibronectin-coated Latex beads, we discovered a role for the voltage-gated potassium channel ether-à-go-go-related gene (ERG) as a downstream effector of CD31 signaling. ERG was identified by tandem mass spectrometry as a 140-kDa protein, which was selectively modified with biotin following the targeted delivery of a biotin-transfer reagent to CD31 using Fab fragments of an anti-CD31 mAb. Similar results were obtained with macrophages but not K562 cells, expressing a truncated cytoplasmic tail of CD31, which failed to regulate bead binding. Colocalization of CD31 with ERG was confirmed by immunofluorescence for K562 cells and macrophages. We now demonstrate that the resting membrane potential of macrophages is depolarized on contact with apoptotic cells and that CD31 inhibits the ERG current, which would otherwise function to repolarize. Sustained depolarization favored the firm binding of phagocytic targets, a prerequisite for efficient engulfment. Our results identify ERG as a downstream effector of CD31 in the regulation of integrin-dependent binding of apoptotic cells by macrophages.

CD31 promotes beta1 integrin-dependent engulfment of apoptotic Jurkat T lymphocytes opsonized for phagocytosis by fibronectin.

Phagocyte integrins, by binding "bridging" molecules, mediate the ingestion of late apoptotic cells and apoptotic bodies by mechanisms that remain obscure. We recently reported that human monocyte-derived macrophages capture viable and apoptotic human leukocytes through homophilic interactions involving CD31 and that CD31 then promotes the engulfment of apoptotic cells or the detachment of viable cells. We now report that CD31 homophilic interactions between phagocyte and target cells lead to activation of phagocyte alpha5beta1 integrin and the engulfment of apoptotic Jurkat T lymphocytes via a fibronectin (Fn) "bridge." Although Fn and serum served as an opsonin for beta1 integrin-dependent phagocytosis of apoptotic leukemic T cells, they failed to do so for neutrophils. Given the complexities and inherent variability of working with primary cells, we have refined our model to show that ligation of CD31 on THP-1 macrophages also regulates beta1 integrin-dependent phagocytosis of Fn-coated Latex beads. Thus, selective "tethering" of apoptotic leukocytes by phagocyte CD31 not only discriminates dying from viable cells but also selectively activates phagocyte integrins for the engulfment of apoptotic cells.

Promoting apoptosis in disease management: a panacea or Trojan horse?

In our enthusiasm to advocate apoptosis as a therapeutic strategy for the management of disease we need to be mindful that the clearance of apoptotic cells is itself immunomodulatory and that it may not always be as benign or beneficial as we think. Indeed, the existence of free apoptotic cells in situ may potentially be pathological, and not necessarily physiological, and any attempt to promote apoptosis in the absence of an appropriate phagocytic response for the treatment of, for example, inflammation or cancer might exacerbate or initiate an autoimmune pathology.