NKR-P1A stimulation of arachidonate-generating enzymes in rat NK cells is associated with granule release and cytotoxic activity

NKR-P1A protein has been implicated in the triggering of NK-mediated natural killing contributing to target cell recognition by NK cells, The aim of the present work was to assess whether NKR-P1A receptor triggering also induced arachidonic acid (AA) generation and to determine the possible role of this event on granule release and cytotoxicity, We demonstrated that activation of fresh peripheral blood rat NK cells by cross-linking with the anti-NKR-P1A 3.2.3 mAb induced calcium-dependent AA release, which is due to the activation of cytosolic phospholipase A(2) (cPLA(2)), secretory phospholipase A(2) (sPLA(2)), and diacylglycerol/monoacylglycerol lipase, We also documented the presence of a type II sPLA, activity in the supernatant fluids from NKR-P1A-activated rat NK cells, suggesting that AA and lysophospholipids could be mobilized from the outside of the cell, The involvement of AA-generating enzymes in NKR-P1A-induced cytotoxic functions was also investigated, Treatment of effector cells with arachidonyl trifluoromethylketone, a cPLA(2) inhibitor; p-bromophenacylbromide, a sPLA(2) inhibitor; or RHC80267, a diacylglycerol lipase inhibitor, led to a partial inhibition of the redirected lysis against P815 target cells as well the granule content release induced by NKR-P1A cross-linking, A complete abolishment of these events was observed when the cells were simultaneously incubated with all three inhibitors, Taken together, our results support a crucial role for the arachidonate-generating enzymes in the induction of lytic activity of NK cells directly or by leading to generation of additional mediators that can play a role in the context of NK cell activation and cytotoxic functions.