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. 2016 Mar 11;12(3):e1005894. doi: 10.1371/journal.pgen.1005894

Fig 5. Abnormal 3D structures in shNEK8 and shNEK8 re-expressing NEK8 mutant protein cells.

Fig 5

(A) 3D structures formed by shNEK8 and shNEK8 re-expressing NEK8 mutants. Cells were grown for 5 days in matrigel prior to phalloidin staining and six categories of structures were observed: (a) normal-sized spheres, (b) large-sized spheres, (c) malformed structures with lumen, (d) elongated structures, (e) branching structures and (f) aggregates without lumen. (A’-A”) Graphs (mean ± SEM of three independent experiments) showing the distribution of normal versus abnormal structures (A’) and quantification of the sphere areas (A”) for each cell line. Scale bar, 10 μm. Statistical analyses by Bonferroni post-hoc test following ANOVA, ns = non significant, **p < 0.01, ***p < 0.001 when samples are compared to shNEK8 + NEK8-GFP WT and ###p < 0.001 to shNEK8. (B). Three-dimensional (3D) spheroids derived from control pLKO and shNEK8 cells were fixed after 2, 3 and 5 days of culture, and were stained for Ki-67 (green), phalloidin (red, F-actin), Hoechst (blue, nuclei). Equatorial sections of representative spheres are shown. Scale bars, 10 μm. (B”) Quantification of Ki-67-positive spheres at 2, 3, and 5 days of culture.