Human neutrophils interact with both 6-sulfo LacNAc+ DC and NK cells to amplify NK-derived IFNγ: role of CD18, ICAM-1, and ICAM-3

Abstract

The role of neutrophils as key players in the regulation of innate and adaptive immune responses is increasingly being recognized. We report that human neutrophils establish a network with both natural killer (NK) cells and 6-sulfo LacNAc ؉ dendritic cells (slanDCs), which ultimately serves to up-regulate NK-derived interferon␥ (IFN␥). This network involves direct reciprocal interactions and positive amplification loops mediated by cellderived cytokines. Accordingly, we show that after lipopolysaccharide ؉ interleu- IntroductionPolymorphonuclear neutrophil leukocytes are known as "professional" phagocytic cells of the innate immune system. However, numerous observations regarding their capacity to respond to and produce a wide variety of cytokines and chemotactic molecules and to change phenotype under specific circumstances also position them as key regulators of various cross talks. 1,2 The outcome(s) of potential interactions between neutrophils and other leukocytes, including macrophages, monocyte-derived dendritic cells (DCs), and T lymphocytes, have been addressed in recent studies [3][4][5][6] and reveal that neutrophils may participate in shaping not only the innate but also the adaptive immune response. 7 For example, activated neutrophils were shown to recruit T-helper 17 (Th17) cells 8 and to trigger T-cell proliferation and strong Th1-cell responses by inducing the maturation of monocyte-derived DCs in in vitro cocultures. 9 Natural killer (NK) cells are effector lymphocytes of the innate immune system endowed with cytotoxic and cytokine-producing capacities, typically recognized for their role in cancer immunosurveillance and pathogen clearance. 10 Although NK cells can directly recognize some pathogens, it is becoming increasingly clear that NK-cell activation requires the presence of accessory cells 11 such as monocytes, 12-14 macrophages, 15,16 and DCs. 17,18 While recent evidence for a potential role of neutrophils in NK-cell activation has been found in mice, 19 it remains unknown whether human neutrophils exert similar effects. Early investigations have focused on a putative inhibitory function of human neutrophils directed toward the cytotoxic activity of cocultured NK cells, 20,21 whereas most subsequent research has delineated the suppressive role of neutrophil-derived reactive oxygen species on NK-cell survival, expression of selected activating receptors, and cytotoxic activity. 22,23 It should also be emphasized that the neutrophil preparations used in most of these studies were not pure (containing approximately 5% of contaminant cells), which might well explain some of the controversial results. 20,21 Based on these premises, we examined whether human neutrophils can activate NK cells either by functioning as and/or modulating the activity of accessory cells. We report that neutrophils potently enhance the release of interferon␥ (IFN␥) by NK cells when cocultured with 6-sulfo LacNAc ϩ dendritic cells (slanDCs), but not CD1c ϩ DCs or plasmacytoid DCs (pDCs), in the presence of lipopoly...

Changing world of neutrophils

“…Some of these interactions are direct, like the effect of B-cell activating factor (BAFF) on survival, maturation and differentiation of B-cells [98], or activation of monocyte-derived dendritic cells (DC) via the integrin Mac1 and DC-SIGN (DC-specific ICAM3-grabbing non-integrin) [113]. In case of NK cells both a direct and an indirect activation by neutrophils has been described, the latter involving a PMN-DC-NK triangle including both contact and humoral effects [24].…”

On the potential involvement of CD11d in co-stimulating the production of interferon- by natural killer cells upon interaction with neutrophils via intercellular adhesion molecule-3

“…4 Recently, we have discovered and have started characterizing a novel network within the innate immune response in which human neutrophils can directly cooperate with both NK cells and one of the major DC subsets present in the blood and tissues, known as 6-sulfo LacNAc + DC (slanDC), to ultimately amplify the production of IFNγ by NK cells upon activation with LPS plus the IL-15/IL-18 combination. 5 Accordingly, we showed that neutrophils promote the release of IL-12p70 by slanDC via a CD18/ICAM-1 interaction, and that such IL-12p70 stimulates NK cells to produce IFNγ: the latter, in turn, has been shown to further potentiate the interaction between neutrophils and slanDC and the release of slanDC-derived IL-12p70, thus creating a positive feedback loop. On the other hand, we also showed that neutrophils could directly co-stimulate the production of IFNγ by NK cells via ICAM-3, 5 but its counter-receptor has not been identified.…”

“…To identify the specific counter-receptors of ICAM-3 that mediate the production of IFNγ by activated neutrophil/NK cell/slanDC co-cultures 5 we initially focused on CD11a that binds both ICAM-1 and ICAM-3, 17 since it was found to be constitutively expressed at high levels in neutrophils, NK cells and slanDC ( Figure 1A). αCD11a mAbs did not affect the release of IL-12p70 by NK cell/slanDC co-cultures, but significantly inhibited the production of IFNγ ( Figure 1B).…”

“…5 The Abs specifically used in this study were: αDC-SIGN [clone 120507, mouse IgG2b from Abcam], αCD11a (clone HI111, mouse IgG1 from BD BioSciences), and αCD11d (rabbit IgG) Abs.…”

“…Highly purified neutrophils, NK cells and slanDC were isolated and cultured for 18 h as previously described, 5 with or without 100 ng/mL LPS plus the IL-15/IL-18 combination (both at 10 ng/mL), in the presence of 10 mg/mL αCD11a/CD18 efalizumab, a humanized IgG1 antibody derived from the murine MHM24 clone 10 which, in preliminary experiments, confirmed its ability to prevent the binding of CD11a to either ICAM-1 11 or ICAM-3 12 in a specific T-cell adhesion assay 13 , 10 mg/mL αCD18 or αCD18 F(ab')2 fragments [clone IB4 (mouse IgG2a)], 10 mg/mL αDC-SIGN [clone 120507 (mouse IgG2b) 14 or clone MR-1 (mouse IgG1) 15 ], as well as corresponding isotype control antibodies. Monocytederived dendritic cells (moDC) were generated as previously described.…”