Figure 6
CXCR2 inhibition reduces frequency and duration of direct contact of dHL-60 cells with PANC-1 tumor spheroids in the “contact” scenario. (a) A schematic showing the definition of contact between a migrating dHL-60 cell (track shown in blue) and a given tumor spheroid. r1 = radius of the tumor spheroid, r2 = radius of a dHL-60 cell. T = duration of contact. Black = boundary of the tumor spheroid, yellow = boundary of the circular “contact” region with a radius of r1 + 2 * r2, green = dHL-60 cells in contact, red = dHL-60 cells not in contact. (b) Representative images showing dHL-60 cells (yellow) treated with DMSO vehicle control or 1 μM AZD-5069 interacting with PANC-1 tumor spheroids (brightfield) at t = 0 h, 2 h, 4 h, 6 h, 8 h, 10 h, and 12 h of time-lapse imaging in the “contact” scenario, overlaid with dHL-60 cell tracks extracted by TrackMate (ImageJ). Any dHL-60 cells that entered the circular ROI (yellow) around the spheroid are considered to be in contact with the spheroid. Scale bar, 50 µm. (c) Bar plots showing the frequency (left panel) and mean duration (right panel) of contact with dHL-60 cells per tumor spheroid over 12 h in specified conditions. Each data point represents a spheroid and n = 107–119 spheroids per condition. Bars show mean ± SD with mean values written above the points. ****: p < 0.0001, unpaired t test with Welch’s correction. (d) Distributions of durations of contact per dHL-60 cell in DMSO vehicle control and AZD-5069 treated conditions. Only contacts shorter than 150 min are shown. (e) Contact between a dHL-60 cell and a tumor spheroid was classified into two categories: short (≤ 30 min) and long (> 30 min). Scatter plot showing the duration of contact per dHL-60 cell in specified conditions. Each data point represents a dHL-60 cell and n = 133–461 cells per condition in long contact. Bars show mean (red) ± SD (black) with mean values written above the points. At least three independent experiments were performed. ns: ≥ 0.05, ***: p < 0.001, unpaired t test with Welch’s correction.
