Effect of IL-18 receptor deficiency on cytotoxic T-cell migration patterns and interaction behavior in a pancreatic carcinoma model

Introduction Infiltration of CD8+cytotoxic T-cells (CTL) in pancreatic ductal adenocarcinoma (PDAC) is an important factor in determining prognosis. However, tumor-infiltrating lymphocytes (TIL) reveal reduced effector functions, a dysfunctional state called “exhaustion”. NLRP3-dependent proinflammatory cytokines IL-1β and IL-18 play a prominent role for CTL induction and differentiation. CTL effector functions are mediated by antigen-dependent contacts with target cells. Migration pattern and interaction behavior of intratumoral CTL is pivotal for tumor rejection.

Aims Here, we investigate effects of T-cellular IL-18R signalling for intratumoral migration and interaction in the context of T cellular exhaustion phenomena.

Methods Murine adenocarcinoma cell line Panc02 was stably transfected with ovalbumin (OVA) as well as fluorophore H2B-cerulean to generate PancOVAH2B-cerulean tumor cells. Dorsal skinfold chambers (DSFC) were installed on wildtype mice and PancOVAH2B-cerulean tumor cells were implanted into the chambers. PancOVA spheroids were formed using Corning®​ Matrigel®​-based 3D cell culture technique. CTL were generated from OT-1 mice, Il1r-/-OT-1 mice or Il18r-/- OT-1 mice and were marked by CellTrackerTM Orange or CellTrackerTM Deep Red dye. This was followed by adoptive transfer of CTL into tumor-bearing mice and CTL were applied into formed spheroids, respectively. Microscopy was performed with a multiphoton microscope (MPM), and Imaris software (©Bitplane AG) was used for visualizing T-cell tracking.

Results Both in the adoptive T-cell transfer model and in the spheroid model, motility of CTL could be visualized by MPM. Imaris analysis indicated significantly higher accumulation ofIl18r-/- CTL in PancOVA tumors and significant reduction of tumor volume compared to wildtype CTL. Additionally, Il18r-/- CTL covered a longer distance (track displacement length) in comparison to wildtype CTL, and, had a higher average speed (track speed mean). Analysis of instantaneous velocity suggested higher percentage of arrested tracks (arrests:<4 μm/min) for Il18r-/- CTL.

Conclusions Previously we showed that IL-18R signal transduction results in ineffective tumor rejection and in induction of exhaustion phenomena. Here, NLRP3-dependent proinflammatory cytokine IL-18 influenced migration pattern and interaction behavior of intratumoral CTL.

 

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Artikel online veröffentlicht:
19. August 2022