Critical and differential roles of NKp46- and NKp30-activating receptors expressed by uterine NK cells in early pregnancy.

In early human pregnancy, uterine decidual NK cells (dNK) are abundant and considered as cytokine producers but poorly cytotoxic despite their cytolytic granule content, suggesting a negative control of this latter effector function. To investigate the basis of this control, we examined the relative contribution to the cytotoxic function of different activating receptors expressed by dNK. Using a multicolor flow cytometry analysis, we found that freshly isolated dNK exhibit a unique repertoire of activating and inhibitory receptors, identical among all the donors tested. We then demonstrated that in fresh dNK, mAb-specific engagement of NKp46-, and to a lesser extent NKG2C-, but not NKp30-activating receptors induced intracellular calcium mobilization, perforin polarization, granule exocytosis and efficient target cell lysis. NKp46-mediated cytotoxicity is coactivated by CD2 but dramatically blocked by NKG2A coengagement, indicating that the dNK cytotoxic potential could be tightly controlled in vivo. We finally found that in dNK, mAb-specific engagement of NKp30, but not NKp46, triggered the production of IFN-gamma, TNF-alpha, MIP-1alpha, MIP-1beta, and GM-CSF proinflammatory molecules. These data demonstrate a differential, controlled role of NKp46- and NKp30-activating receptors expressed by dNK that could be critical for the outcome of pregnancy and the killing of uterine cells infected by pathogens.

CD160-activating NK cell effector functions depend on the phosphatidylinositol 3-kinase recruitment.

CD160 NK cell-activating receptor is a glycosyl-phosphatidylinositol-anchored molecule that, upon specific engagement, triggers both cytotoxicity and a unique cytokine production [IFN-gamma, tumor necrosis factor-alpha (TNF-alpha) and IL-6] through an undefined signaling pathway. In the current study, we have identified several signaling molecules recruited after mAb-specific CD160 engagement in freshly isolated human circulating NK cells. Using confocal microscopy, we found that CD160 engagement induces the recruitment and co-localization of phosphorylated molecules with redistributed, capped CD160 at the cell surface. We then demonstrated that phosphatidylinositol 3-kinase (PI3K) signaling molecule is required for CD160-mediated cytotoxicity and cytokine release. First, we observed by confocal microscopy that engagement of CD160 induces its polarization and co-localization with PI3K. Second, we showed that pharmacological inhibitors of PI3K abrogate both CD160-mediated cytotoxicity and IFN-gamma, TNF-alpha and IL-6 cytokine release. We further found that CD160 engagement induced marked phosphorylation of Akt, as evidenced by western blotting. We identified additional CD160-mediated signaling molecules recruited downstream and upstream of PI3K. Both induction of phosphorylated ERK molecules after CD160-specific engagement and prevention of CD160-induced cytokine release by MEK pharmacological inhibitor indicate that ERK downstream pathway is implicated. Similarly, we identified that Syk molecule upstream of PI3K is involved in the signaling cascade mediated by CD160 engagement. Two different Syk-specific inhibitors blocked CD160-mediated cytokine release, and CD160-specific engagement induced the enhancement of phosphorylated Syk proteins. These data demonstrate that PI3K is a crucial signaling element for both effector functions of the CD160 NK cell-activating receptor.

Engagement of the CD160 activating NK cell receptor leads to its association with CD2 in circulating human NK cells.

CD160 activating NK cell receptor is a glycosylphosphatidylinositol-anchored molecule. Upon specific engagement by its HLA class I physiological ligand, CD160 NK cell receptor triggers cytotoxicity and release of different cytokines, including IL-6. How CD160 intracellular signaling is mediated following its specific ligation is unknown. One key element of this signaling may be through the co-activation of other receptors associated to CD160. Using confocal microscopy and freshly isolated human peripheral blood NK cells, we observed a co-polarization of CD160 and CD2 after their respective specific engagement. These results demonstrate that CD160 activating NK cell receptor, upon its specific engagement, physically associates to the CD2 co-activating receptor present in the lipid raft that might play a role as signaling molecule.

HLA class I/NK cell receptor interaction in early human decidua basalis: possible functional consequences.

Human decidual NK (dNK) cells differ from their peripheral blood (PB)-NK counterparts. The major subset of PB-NK is CD56dim, CD16+, CD160+ (highly cytolytic), whereas the major subpopulation of dNK is CD56bright, CD16- and CD160- (high cytokine producer). Extravillous cytotrophoblast invading the decidua basalis in early pregnancy expresses the polymorphic HLA-C, and nonpolymorphic HLA-E and HLA-G molecules that can interact with specific HLA class I-dependent dNK receptors, including the immunoglobulin-like KIRs, the lectin-like CD94/NKG2 and the CD160 receptors. There is no clear evidence thus far that dNK cells kill trophoblast cells. Instead they are able to secrete cytokines which are likely to be beneficial for the placental development, maternal uterine spiral arteries remodeling, and the antiviral immune response.

Cutting edge: engagement of CD160 by its HLA-C physiological ligand triggers a unique cytokine profile secretion in the cytotoxic peripheral blood NK cell subset.

CD160 is an Ig-like activating NK cell receptor expressed on the majority of circulating NK cells. This population corresponds to the nonproliferating, highly cytolytic, CD56dimCD16+ subset. CD160 engagement by HLA-C molecules mediates cytotoxic function. In this study, we report that upon specific activation by the physiological ligand HLA-C, or Ab cross-linking, CD160+ peripheral blood NK cells produce IFN-gamma, TNF-alpha, and IL-6. This unique CD160-mediated cytokine production differs from the one observed after CD16 engagement whose expression is also restricted to the CD56dim cytotoxic NK cell subset. As already reported for the CD160-mediated cytotoxic effector function, CD160-mediated cytokine production by peripheral blood-NK cells is negatively controlled by the killer Ig-like receptor CD158b. Thus, the CD160 receptor represents a unique triggering surface molecule expressed by cytotoxic NK cells that participates in the inflammatory response and determines the type of subsequent specific immunity.

Engagement of CD160 receptor by HLA-C is a triggering mechanism used by circulating natural killer (NK) cells to mediate cytotoxicity.

Circulating human natural killer (NK) lymphocytes have been functionally defined by their ability to exert cytotoxic activity against MHC class I-negative target cell lines, including K562. Therefore, it was proposed that NK cells recognized the "missing self." We show here that the Ig-like CD160 receptor expressed by circulating CD56(dim+) NK cells or IL-2-deprived NK cell lines is mainly involved in their cytotoxic activity against K562 target cells. Further, we report that HLA-C molecules that are constitutively expressed by K562 trigger NK cell lysis through CD160 receptor engagement. In addition, we demonstrate, with recombinant soluble HLA-Cw3 and CD160 proteins, direct interaction of these molecules. We also find that CD158b inhibitory receptors partially interfere with CD160-mediated cytotoxicity, whereas CD94CD159a and CD85j have no effect on engagement with their respective ligands. Thus, CD160HLA-C interaction constitutes a unique pathway to trigger NK cell cytotoxic activity.