Efficient Killing of Murine Pluripotent Stem Cells by Natural Killer (NK) Cells Requires Activation by Cytokines and Partly Depends on the Activating NK Receptor NKG2D

Abstract

Natural killer (NK) cells play an important role as cytotoxic effector cells, which scan the organism for infected or tumorigenic cells. Conflicting data have been published whether NK cells can also kill allogeneic or even autologous pluripotent stem cells (PSCs) and which receptors are involved. A clarification of this question is relevant since an activity of NK cells against PSCs could reduce the risk of teratoma growth after transplantation of PSC-derived grafts. Therefore, the hypothesis has been tested that the activity of NK cells against PSCs depends on cytokine activation and specifically on the activating NK receptor NKG2D. It is shown that a subcutaneous injection of autologous iPSCs failed to activate NK cells against these iPSCs and can give rise to teratomas. In agreement with this result, several PSC lines, including two iPSC, two embryonic stem cell (ESC), and two so-called multipotent adult germline stem cell (maGSC) lines, were largely resistant against resting NK cells although differences in killing were found at low level. All PSC lines were killed by interleukin (IL)-2-activated NK cells, and maGSCs were better killed than the other PSC types. The PSCs expressed ligands of the activating NK receptor NKG2D and NKG2D-deficient NK cells from Klrk1−/− mice were impaired in their cytotoxic activity against PSCs. The low-cytotoxic activity of resting NK cells was almost completely dependent on NKG2D. The cytotoxic activity of IL-2-activated NKG2D-deficient NK cells against PSCs was reduced, indicating that also other activating receptors on cytokine-activated NK cells must be engaged by ligands on PSCs. Thus, NKG2D is an important activating receptor involved in killing of murine PSCs. However, NK cells need to be activated by cytokines before they efficiently target PSCs and then also other NK receptors become relevant. These features of NK cells might be relevant for transplantation of PSC-derived grafts since NK cells have the capability to kill undifferentiated cells, which might be present in grafts in trace amounts.

NK cells prevent the formation of teratomas derived from human induced pluripotent stem cells

“…Here, we provide evidences that hiPSCs are the target of NK cells in vitro and in humanized mice. In opposition to what was found using murine iPSCs, human NK cells could prevent the formation of teratomas without prior activation [5]. Other than the species differences, such disparity may be explained by the use of a different injection site in our study (renal capsule versus subcutaneous) and perhaps by the high expression level of NK activating ligands (i.e.…”

“…However, the need for pluripotent stem cells to evade immune responses may be required for the growth of teratomas in vivo . Pioneer work by Dressel and colleagues showed that mouse iPSCs can occasionally give rise to teratomas in autologous/syngeneic mice in absence of the activation of the host natural killer (NK) cells [4; 5]. Likewise, human iPSCs were shown to be the target of human NK cells in vitro [6; 7].…”

Immunological Properties of Murine Parthenogenetic Stem Cells and Their Differentiation Products

“…We have previously reported that mouse ESC, maGSC, and iPSC lines are highly susceptible targets for LAK cells as well as purified IL-2-activated NK cells ( 32 , 36 , 37 ). These pluripotent stem cell types were all killed by allogeneic and syngeneic LAK cells to a similar extent or even better than YAC-1 cells that are the typical target cell line for murine NK cells.…”

“…However, the expression patterns of these NK receptor ligands cannot explain the higher susceptibility of PSC A3 than PSC A6 cells to LAK and NK cells that we observed in our experiments. The functional relevance of the NKG2D receptor for the killing of ESCs ( 32 , 37 , 47 ) and other pluripotent stem cell types ( 36 , 37 ) has been demonstrated in inhibition and gene knockout experiments. Initially, some studies reported resistance of ESCs to NK cells ( 48 , 49 ), while we and others found them to be highly susceptible ( 32 , 36 , 47 , 50 ).…”

“…So far, the analysis of the immunological properties of PSCs is lacking behind the other pluripotent stem cell types. We have shown previously that mouse ESCs, iPSCs, and maGSCs are targets for IL-2-activated allogeneic as well as syngeneic NK cells ( 32 , 36 , 37 ) and that NK cells can impair the growth of teratomas upon transplantation of the stem cells ( 20 , 36 , 38 ). Since in vitro differentiation increased the resistance of these pluripotent stem cell types ( 36 ), NK cells might increase the safety of transplantations because they can target undifferentiated cells while ignoring differentiated cells.…”