Intravital Imaging Reveals Distinct Dynamics for Natural Killer and CD8+ T Cells during Tumor Regression

Abstract

Recognition of NKG2D ligands by natural killer (NK) cells plays an important role during antitumoral responses. To address how NKG2D engagement affects intratumoral NK cell dynamics, we performed intravital microscopy in a Rae-1β-expressing solid tumor. This NKG2D ligand drove NK cell accumulation, activation, and motility within the tumor. NK cells established mainly dynamic contacts with their targets during tumor regression. In sharp contrast, cytotoxic T lymphocytes (CTLs) formed stable contacts in tumors expressing their cognate antigen. Similar behaviors were observed during effector functions in lymph nodes. In vitro, contacts between NK cells and their targets were cytotoxic but did not elicit sustained calcium influx nor adhesion, whereas CTL contact stability was critically dependent on extracellular calcium entry. Altogether, our results offer mechanistic insight into how NK cells and CTLs can exert cytotoxic activity with remarkably different contact dynamics.

Rapid and Unidirectional Perforin Pore Delivery at the Cytotoxic Immune Synapse

“…The slower degranulation kinetics and multiple calcium flux events of mouse CTLs compared with mouse NK cells are consistent with the longer-lived target contact times and sustained calcium signaling profiles observed previously with mouse CTLs (15). Interestingly, NK cells appear to degranulate more quickly upon target cell engagement, irrespective of species.…”

“…Importantly, the mode of target cell recognition, engagement, and intracellular signaling events largely underpin their specificity (14). Recent in vivo imaging studies showed that NK cells tend to make transient contacts with their tumor targets, whereas CTLs form longer, more stable contacts (15). The authors suggested these differences in contact time could be explained by differences in calcium responses, with NK cells showing weak calcium influxes, whereas CTLs showed high, sustained calcium influxes.…”

Cutting Edge: Tumor-Targeting Antibodies Enhance NKG2D-Mediated NK Cell Cytotoxicity by Stabilizing NK Cell–Tumor Cell Interactions

“…In vitro experiments revealed that, although NKG2D signaling enhanced degranulation, it had little influence on adhesion to target cells, whereas FcR engagment improved adhesion but not degranulation. Using two-photon microscopy, we showed that recognition through FcR affected intratumoral NK cell behavior and promoted long-lasting interactions with targets in vivo, whereas NKG2D signals alone failed to promote NK cell arrest, as we described previously (14). Altogether, our results uncover a novel aspect of cooperation between activating receptors in NK cells and suggest that cell dynamics represent a critical regulator of cell-mediated cytotoxicity.…”

“…Imaging of tumors was performed as described before (14,15) using an upright microscope (DM6000B; Leica) with a water-dipping 253/1.05 NA objective (Olympus). Excitation was provided with a Ti:sapphire laser (Coherent) tuned at 950 nm.…”

Visualization of Myelomonocytic Cells in Tumours

“…Although it is known that both CTLs and NK cells mediate similar cytotoxic activity (Russell and Ley 2002), this study showed that NK cells had a distinct intratumoral behaviour compared to CTLs. Tumor infiltrating NK cells only formed transient contacts with target cells after initial interaction, whereas CTLs normally establish long lasting contacts that can last for more than 20 minutes (Deguine et al 2011). …”