NK-cell Chemotaxis Towards Senescent Cells

NK-cell chemotaxis experiments were performed and analyzed as described elsewhere (Tasdemir et al., 2016 (link)) with the following variations: NK cells were stained with the CellTracker Red CMTPX Dye (Thermo Fisher Scientific) (according to the manufacturer’s instructions) and then seeded into 6-well plates containing proliferating or senescent mVenus-expressing IMR90 cells also expressing the indicated shRNAs, at 60% confluency. Cocultures were imaged over time using an Incucyte-HD or Incucyte-Zoom device (Essen Bioscience) in a 5% CO₂ atmosphere, using a 20x objective and the 488 nm and 561 nm laser excitations. Images were captured every 45 minutes, starting 30 minutes after NK-cell seeding onto the IMR90 cultures. Cell proliferation was determined through repeated-measures of confluency on phase or epifluorescent imaging.