Liver DX5- Natural Killer Cells Play a Crucial Role On Instant Blood-Mediated Inflammatory Reaction After Islet Transplantation.

Abstract

Introduction: Destruction of islet grafts by the instant blood-mediated inflammatory reaction (IBMIR) and the immune response after islet transplantation remain a major obstacle in islet transplantation. We have previously shown that natural killer (NK) cells in the liver play a key role in early inflammatory events that adversely affect islet function and engraftment in the liver. Here, we investigated the relationship between the IBMIR and activation of liver NK cells after islet transplantation by analyzing phenotypic alterations in liver NK cells under proinflammatory stress-mimicking IBMIR conditions in vitro. Methods: Liver NK cells from C57BL6 mice were cultured for 24 h in the presence of IFNγ, TNFα and IL-1β, which were predominantly and transiently produced on the IBMIR conditions. The expression of activation marker CD69 and TNF-related apoptosis-inducing ligand (TRAIL) on liver NK cells was analyzed by flow cytometry. Results: Liver NK cells were classified into subsets of DX5-NK1.1+ or conventional DX5+ NK1.1+ NK cells. Liver NK cells contained approximately seven times as many DX5- NK cells as splenic NK cells. In naive conditions, the proportions of CD69+ and TRAIL+ NK cells in the DX5- NK cell were significantly higher than in DX5+ NK cells (CD69: 84.1 ± 7.5% vs 12.8 ± 4.2%, TRAIL: 70.8 ± 9.3% vs 4.9 ± 2.2%, respectively), suggesting that DX5- NK cells are in an activation stage even without any stimulation. The expression of these markers on splenic NK cells was similar to that on liver DX5+ NK cells. The proportion of CD69+ NK cells was not increased when cultured with IFNγ, TNFα, or IL-1β alone (24h/fresh ratio; IFNγ 0.79, p=0.29; TNFα 1.24, p=0.31; IL-1β 0.63, p=0.14). However, the proportion of CD69+ NK cells was significantly increased only when all three cytokines were simultaneously added (24h/fresh ratio; 2.00, p<0.01). Furthermore, the population of DX5- CD69+ NK cells expanded under co-culture with all three cytokines (27.5%±5.8% vs. 65.7±13.8%, p<0.01), whereas DX5+ CD69+ NK cells did not. In addition, IFNγ production was promoted in DX5- NK cells. Conclusion: Liver-resident DX5-NK cells play a crucial role in the liver NK cells, and activation of these NK cells was promptly induced by IBMIR circumstance. Thus, targeting the elimination or inactivation of liver-resident DX5- NK cells may improve the engraftment of transplanted islets without inhibiting the innate immune system.