Intravital characterization of tumor cell migration in pancreatic cancer

Abstract

Curing pancreatic cancer is difficult as metastases often determine the poor clinical outcome. To gain more insight into the metastatic behavior of pancreatic cancer cells, we characterized migratory cells in primary pancreatic tumors using intravital microscopy. We visualized the migratory behavior of primary tumor cells of a genetically engineered pancreatic cancer mouse model and found that pancreatic tumor cells migrate with a mesenchymal morphology as single individual cells or collectively as a stream of non-cohesive single motile cells. These findings may improve our ability to conceive treatments that block metastatic behavior.

Keywords: collective migration; intravital microscopy; migration; pancreatic tumors.

Figure 1.

(A)genetic fluorescent mouse model of pancreatic cancer. (A) Schematic representation of the fluorescent mouse model in which all tumor cells express YFP upon recombination by Cre recombinase under the Pdx-promotor. In these cells, Cre-regulated mutated forms of Kras (Kras^LSL-G12D) and p53 (p53^LSL-R172H), combined with a floxed p53 give rise to pancreatic tumors. (B) Representative intravital microscopy images showing different types of tumor morphology in yellow and surrounding collagen I in magenta. Scale bar, 50 µm. (C) The number of migrating cells per imaging field (n = 26 fields in 5 animals). The 5 different animals are shown in symbols with different shapes/colors. Mean ± SEM.

Figure 2.

Tumor cells of pancreatic carcinoma migrate as mesenchymal cells. Time-lapse intravital imaging of a fluorescent mouse model of pancreatic cancer with tumor cells shown in yellow, collagen I in magenta, intravenously injected 70 k_D Dextran in red and nuclei in blue. (A) Example of an imaging field (showing 4 time points (120 minute interval)), in which polar and elongated single cell migration was observed (boxed area is enlarged in B). Scale bar, 50 µm. (B) Boxed area in (A)showing an example of an imaging field (7 timepoints (60 minute interval) with migrating polar and elongated single tumor cells (example in circle) with a track (last timepoint, white line). Separate channels show the nuclei (Hoechst) and tumor cells (YFP), indicating that the particles that migrate contain nuclei. Scale bar, 50 µm.

Figure 3.

Single tumor cell migration in pancreatic carcinoma. Time-lapse intravital imaging of a genetic fluorescent mouse model of pancreatic cancer with tumor cells shown in yellow and collagen I in magenta. (A) Example of an imaging field (showing 4 time points (30 minute interval)), in which polar and elongated single cell migration was observed (boxed area is enlarged in B). Scale bar, 50 µm. (B) Boxed area in (A)showing migrating polar and elongated single tumor cells (in dashed circles) with a track (last time point, white line). Scale bar, 20 µm.

Figure 4.

Collective streaming of tumor cells in pancreatic carcinoma. Time-lapse intravital imaging of a genetic fluorescent mouse model of pancreatic cancer with tumor cells shown in yellow and collagen I in magenta. (A) Example of an imaging field (showing 4 time points (30 minute interval)), in which a collective stream of polar and elongated single tumor cells was observed (boxed area is enlarged in B). Scale bar, 50 µm. (B) Boxed area in a showing a collective stream of polar and elongated single tumor cells with a track (last timepoint, white line). Scale bar, 20 µm. (C) The persistency (left graph) and migration speed (right graph) are plotted for every migrating tumor cell. As a control the speed of non-migratory cells are shown (right graph). (D) The circularity (left graph) and aspect ratio (right graph) are plotted for every migrating tumor cell.