Self inhibition of phagocytosis: The affinity of 'marker of self' CD47 for SIRPα dictates potency of inhibition but only at low expression levels

Phagocytes engulf foreign cells but not 'self' in part because self cells express CD47 as a ligand for signal regulatory protein SIRP alpha, which inhibits phagocytosis. Motivated by reports of upregulation of CD47 on both normal and cancerous stem cells [1: Jaiswal et al., 2009] and also by polymorphism in SIRP alpha [2: Takenaka at al., 20071, we show here that inhibition of engulfment correlates with affinity of CD47 for SIRP alpha - but only at low levels of CD47. One common human polymorph of SIRP alpha is studied and binds more strongly to human-CD47 than to mouse-CD47 (K(d)approximate to 0.12 mu M and 6.9 mu M, respectively) and does not bind sheep red blood cells (RBCs) - which are well-established targets of human macrophages; in comparison, a common mouse polymorph of SIRP alpha binds with similar affinity to human and mouse CD47 (K(d)approximate to 0.22 mu M). Using immunoglobulin (IgG)-opsonized particles with varying levels of either human- or mouse-CD47, the effective inhibition constants K(i) for blocking phagocytosis are then determined with both human- and mouse-derived macrophages. Only human phagocytes show significant differences in man versus mouse K(i)'s and only at CD47 levels below normal densities for RBCs. While phospho-signaling through human-SIRP alpha shows similar trends, consistent again with the affinity differences, saturating levels of CD47 (>K(i)) can signal and inhibit phagocytosis regardless of man versus mouse. Quantitative analyses here prompt more complete characterizations of both CD47 levels and SIRP alpha polymorphism when attempting to study in vivo effects of these key proteins in innate immunity. (C) 2010 Elsevier Inc. All rights reserved.