Selective T-cell depletion in CD70-Tg peripheral lymph nodes is also not caused by IFN-γ 30. Higher apoptosis percentages and LBH589 order up-regulated CD95 expression in CD70-Tg

NK cells indicate that the observed NK cell depletion is at least partly due to apoptotic events and that these are mediated via CD95. However, we performed an in vivo CD95 ligand blocking experiment in CD70-Tg mice and we did not observe rescue of NK cells. Therefore, we have no evidence that the increased expression of CD95 on NK cells from CD70-Tg mice leads to their death. Taken together, our results show that although CD27 cross-linking initially induces activation of lymphocytes, continuous stimulation results in severe homeostatic changes of the lymphocyte population, Nutlin3a including activation-induced cell death of

NK cells. Residual NK cells in CD70-Tg mice exhibited decreased, but not absent expression of CD11b and CD43. This demonstrates that continuous CD27 triggering does not induce a total blockade of NK cell differentiation. In addition, it has been evidenced that under some circumstances CD11blowCD43− NK cells express Ly49 receptors and are lytic, which demonstrates their acquired effector functions 37. This stresses that the CD11blowCD43− phenotype already might be an important checkpoint in the functional differentiation of NK cells. This hypothesis is supported by the findings that only CD11blowCD43− NK cells are present in mice deficient for several different transcription factors, such as GATA-3, IRF2 or T-bet, and in mice bearing constitutively active NFκB. In all these models, the CD11blowCD43− NK cells exhibit normal cytotoxic capacities 38–41. In our study, we found that splenic NK cells from CD70-Tg mice, whether stimulated through the IL-12/IL-18 receptor or through the NK1.1 receptor, produced less IFN-γ compared with WT NK cells, whereas in liver no differences were demonstrated. Regarding cytotoxicity, both liver and splenic NK cells from CD70-Tg mice showed increased activity. Hence, we evidenced opposite effects of CD70 triggering on the major NK cell effector functions. Different outcomes of those NK cell effector functions

upon the same triggering have been described before, for example in the GATA-3 deficient mouse model 38. On the other hand, as several hours are required for ifn-γ HAS1 transcription and translation, the NK cells were incubated for 6 h in the IFN-γ assay, during which the higher apoptosis in the mNK cell population of CD70-Tg mice can have an important impact. Conversely, the outcome of the cytotoxicity assay is probably less influenced by the increased apoptosis of the CD70-Tg NK cell effector population as the trigger to induce NK cell-mediated cytotoxicity of YAC-1 targets requires only 20 min 42. Since YAC-1 lysis is known to be NKG2D-mediated 33, NK cell expression of this receptor was measured in CD70-Tg mice. As expected, enhanced NKG2D expression confirmed the observed up-regulated cytotoxicity in CD70-Tg NK cells.