Fig. 3

Loss of CAMKKβ reduces EOC spheroid viability and has cell line-dependent effects on spheroid invasion and cell migration. a Representative images and viable cell number of OVCAR8 parental, OVCAR8 CAMKK2 KO, HeyA8 parental, and HeyA8 CAMKK2 KO spheroids at time points of 3, 5, and 7 days after seeding to spheroid culture conditions, as determined via trypan blue exclusion cell counting. b Representative images of OVCAR8 parental, OVCAR8 CAMKK2 KO, HeyA8 parental, and HeyA8 CAMKK2 KO spheroids prior to reattachment (starting size), ZT-GFP mesothelial cell monolayers 48 h after spheroid reattachment was initiated (cleared area), and quantification of normalized cleared area. c Representative images and quantification of scratch closure over time in OVCAR8 parental, OVCAR8 CAMKK2 KO, HeyA8 parental, and HeyA8 CAMKK2 KO monolayers. For a, data represent mean cell count after 3, 5, or 7 days in spheroid culture ± SEM for n = 4 independent experiments. Data were analyzed via unpaired two-tailed student’s t-tests and the Holm-Šídák method was used to control for multiple comparisons (*p < 0.05, ***p < 0.001, ****p < 0.0001). For b, data represent mean normalized cleared area ± SEM for 3 independent experiments, each comprising 8 individual spheroids (24 spheroids in total). Cleared area was normalized on a per-spheroid basis to the cross-sectional area of the spheroid prior to initiating reattachment. Data were analyzed via two-tailed student’s t-tests. For c, data represent mean scratch closure as a percentage of the initial scratch area ± SEM for 3 independent experiments, each comprising 3 fields of view from each of 2 wells (18 fields of view in total). Data were analyzed via two-way repeated measures ANOVA (⁺⁺p < 0.01). Scale bars: a, c 1 mm, b 500 μm (equal scale for starting size and cleared area).