Human NK Cytotoxicity against Porcine Cells Is Triggered by NKp44 and NKG2D

Abstract

Pig-to-human xenotransplantation has been proposed as a means to alleviate the shortage of human organs for transplantation, but cellular rejection remains a hurdle for successful xenograft survival. NK cells have been implicated in xenograft rejection and are tightly regulated by activating and inhibitory receptors recognizing ligands on potential target cells. The aim of the present study was to analyze the role of activating NK receptors including NKp30, NKp44, NKp46, and NKG2D in human xenogeneic NK cytotoxicity against porcine endothelial cells (pEC). 51Cr release and Ab blocking assays were performed using freshly isolated, IL-2-activated polyclonal NK cell populations as well as a panel of NK clones. Freshly isolated NK cells are NKp44 negative and lysed pEC exclusively in an NKG2D-dependent fashion. In contrast, the lysis of pEC mediated by activated human NK cells depended on both NKp44 and NKG2D, since a complete protection of pEC was achieved only by simultaneous blocking of these activating NK receptors. Using a panel of NK clones, a highly significant correlation between anti-pig NK cytotoxicity and NKp44 expression levels was revealed. Other triggering receptors such as NKp30 and NKp46 were not involved in xenogeneic NK cytotoxicity. Finally, Ab-dependent cell-mediated cytotoxicity of pEC mediated by human NK cells in the presence of xenoreactive Ab was not affected by blocking of activating NK receptors. In conclusion, strategies aimed to inhibit interactions between NKp44 and NKG2D on human NK cells and so far unknown ligands on pEC may prevent direct NK responses against xenografts but not xenogeneic Ab-dependent cell-mediated cytotoxicity.

Human tumor‐induced and naturally occurring Treg cells differentially affect NK cells activated by either IL‐2 or target cells

“…In agreement with previous reports, we found that IL-2 induces activation of primary human peripheral blood NK cells resulting in upregulation of activating receptors, NKG2D and NKp44, as well as increased degranulation and IFN-g secretion [21,22]. These effects were significantly impaired in the presence of tumor iTreg cells, nTreg cells or TGF-b.…”

Porcine UL16-Binding Protein 1 Expressed on the Surface of Endothelial Cells Triggers Human NK Cytotoxicity through NKG2D

“…The relative inhibition was stronger when freshly isolated NK cells were used as compared with IL-2-activated NK cells. This finding was expected, because only the latter express NKp44, which is also involved in xenogeneic NK cytotoxicity (21,23). However, the incomplete blocking of xenogeneic NK cytotoxicity observed using freshly isolated NK effectors indicated the presence of yet unidentified interactions between porcine ligands with human activating NK receptors.…”

“…There is a direct correlation between the surface density of NCR and the ability of NK cells to lyse various tumor cells (22), but the cellular ligands recognized by NCR remain unidentified. As recently reported, human NK cytotoxicity against pEC is mediated primarily through NKG2D and NKp44, whereas NKp30 and NKp46 are not involved (23).…”

“…Besides the lack of MHC class I inhibition, the recognition of putative porcine ligands by human activating NK receptors plays an important role in xenogeneic NK cytotoxicity. Recently, we demonstrated that lysis of pEC mediated by freshly isolated human NK cells is mainly triggered by NKG2D (23). In contrast, lysis mediated by IL-2-activated human NK cells depends on both NKG2D and NKp44.…”

“…Blocking of human NK cytotoxicity against porcine cells with anti-NKG2D mAb provided indirect evidence for cross-species interactions between human NKG2D and one or several different porcine ligands (23). However, little is known about the nature and function of these putative ligands.…”