FcγRs join in the cascade

A growing body of evidence indicates that neutrophil FcγRs support adhesion to immune complexes (ICs) under shear flow conditions. In this issue of Blood, Florey and colleagues report that both FcγRIIA (CD32) and FcγRIIIB (CD16) on human neutrophils can mediate adhesion to human IgG–coated vascular endothelial cells under in vitro shear flow conditions and exhibit differential requirements for endothelial accessory molecules of the classical multistep adhesion cascade.

Neutrophil recruitment to sites of infection or injury is widely held to occur by a multistep adhesion cascade leading to stable leukocyte adhesion to endothelium. Treatment of cultured endothelium with inflammatory cytokines (eg, TNF-α, IL-1β, LPS) triggers expression of adhesion molecules and chemokines that promote significant leukocyte adhesion and transmigration. The first step in adhesion (initial attachment and rolling) is mediated by members of the selectin family of adhesion molecules (E-, P-, and L-selectin).¹  The second step, firm adhesion, requires activation of leukocyte β2 integrins and their engagement of endothelial-cell counterreceptors ICAM-1 and ICAM-2.²  The β2 integrins also contribute to the deceleration of leukocyte rolling, which may facilitate firm adhesion. Integrin activation is mediated by chemoattractants and chemokines presented on the endothelial-cell surface. Transmigration is the final step and involves multiple adhesion molecules including β2-integrin, ICAM-1, PECAM-1, and CD99.³ 

In systemic immunologic diseases such as systemic lupus erythematosus (SLE) or rheumatoid arthritis, high levels of circulating self-reacting antibodies form ICs, deposit in tissues and organs, and trigger sustained recruitment of neutrophils (reviewed in Firestein⁴ ). Accordingly, effort has been placed on elucidating the mechanisms underlying IC-mediated neutrophil recruitment in order to create effective therapeutics, which begs the question whether IC-dependent mechanism(s) share components of the multistep cascade. Neutrophil interaction with ICs is mediated by 2 low-affinity receptors, FcγRIIA and FcγRIIIB. Coxon et al were the first to demonstrate a primary role for FcγRIIIB in capture and adhesion of human neutrophils to immobilized rabbit IgG containing ICs (2.0 dynes/cm²) in the absence of adhesion molecules that mediate attachment (ie, selectins).⁵  They further found that β2 integrins mediated shear-resistant adhesion to ICs. Recently, Skilbeck et al reported that human neutrophil adhesion and subsequent spreading on immobilized human IgG required FcγRIIIB, with a lesser role by FcγRIIA for adhesion, and that stable adhesion and spreading was β2 integrin dependent⁶ ; however adhesion occurred only at very low shear stress (0.5 dynes/cm²). These authors also suggested that the role of FcγRIIIB versus FcγRIIA was due to species differences in the IgG because adhesion to human IgG was FcγRIIA and FcγRIIIB dependent, whereas adhesion to rabbit IgG was solely FcγRIIIB dependent. In vivo, IC deposition within the vasculature resulted in rapid FcγRIII-dependent neutrophil recruitment in mice^5,7 ; albeit the relative contribution of the human receptors in vivo is not clear because murine neutrophils express a transmembrane form of FcγRIIIB that requires a signaling gamma subunit (FcγRIIIB), while human neutrophils express a GPI-linked FcγRIIIB and the single subunit FcγRIIA. FcγRs were also required for slow rolling through P-selectin and enhanced adhesion in the context of P-selectin and intravascular IC deposition.⁷  On the other hand, IC formation primarily in tissues resulted in endothelial-cell activation, which led to neutrophil recruitment that was dependent on many of the traditional players in neutrophil trafficking, including endothelial selectins and VCAM-1.⁸ 

The current report by Florey and colleagues extends the field by introducing a new model that contains endothelial monolayers coated with SLE patient IgG or ICs in an in vitro flow model. The key observation was that SLE patient IgG-coated human microvascular endothelial cells (HMECs) supported neutrophil adhesion and that adhesion depended on (1) the HMEC activation status (ie, TNF-α activation), and (2) whether HMECs were coated with IgG or ICs. The authors report that the IgG-coated HMECs provoked enhanced neutrophil adhesion under shear flow only if HMECs were preactivated with TNF-α, and that this augmented adhesion was dependent solely on FcγRIIA and not FcγRIIIB, and required E-selectin, leukocyte β2 integrin, and CXCL1/2 (IL-8R). A role for FcγRIIIB-dependent neutrophil adhesion was detected for IC-coated unstimulated HMECs expressing E-selectin, suggesting a prerequisite for selectin-mediated capture. High-density immobilized ICs also supported neutrophil adhesion as previously described.⁵  The take-home message is that both FcγRIIA and FcγRIIIB can mediate adhesion to ICs under flow but that FcγRIIA has a specialized nonredundant ability to augment neutrophil adhesion to monomeric IgG-coated HMECs and to promote stable adhesion by β2 integrins under shear flow, while FcγRIIIB predominates in the context of ICs. The authors speculate that E-selectin and/or IL-8R ligation stimulates incorporation of key signaling cofactors into lipid rafts in which FcγRIIA is reported to reside in monocytes. The requirement for FcγRIIA in polymorphonuclear leukocyte (PMN) adhesion to monomeric IgG–coated HMECs is surprising because of this receptor's known low-affinity for monomeric IgG. Indeed, FcγRI has high affinity for monomeric IgG. One possibility is that the mAb contained aggregated IgG and/or that the surface Ag recognized by the antibody (endoglin) is clustered and results in a “patch” of ICs. Taken together, these findings suggest that in patients with circulating self-directed antibodies, augmented neutrophil adhesion to microvascular endothelium may be disproportionately higher in relation to the level of endothelial-cell activation, driving sustained neutrophil recruitment and perhaps dysregulation of an inflammatory response to infection or tissue injury.

In summary, the relative roles of FcγRIIIB versus FcγRIIA may be dictated by the species and/or isotype of the IgG/ICs present or the density of ICs. At low density of immobilized monomeric IgG, the activation state of the endothelium and its accessory molecules augment neutrophil recruitment, whereas at high-density ICs, FcγRs are sufficient to capture and mediate adhesion. Furthermore, the location of the ICs, primarily intravascular versus extravascular, may dictate the requirement for neutrophil FcγRs, endothelial activation, and associated molecules. The molecular requirements for IC-induced neutrophil recruitment and the circumstances under which different FcγRs are engaged are fruitful areas for future investigation.