Figure 3 | Scientific Reports

Figure 3

From: Doublecortin-like kinase 1 promotes hepatocyte clonogenicity and oncogenic programming via non-canonical β-catenin-dependent mechanism

Figure 3

DCLK1-overexpressing hepatoma cells exhibit clonogenicity and produce dedifferentiated lineages. (a) Live cell imaging of spheroids derived from Huh7-RFP-DCLK1 (upper panel) and Huh7-RFP cells (lower panel) at × 4 magnification. Single cell suspension of these cells from monolayer cultures were added onto the matrigel layer in Lab-Tek-II 8-well chambered cover slides. The cells were cultured in Hepato-STIM hepatocyte defined media containing EGF. The photographs of developing spheroids (same areas, marked by arrows) were taken at day 4, 8 and 12 post-plating under bright field. (b) Quantitation of the number of spheroids formed by 1,000 Huh7-RFP-DCLK1 (black bar) or Huh7-RFP (gray bar) cells. The spheroids were manually counted for each culture. (c) Confocal microscopy of spheroids directly immunostained in the matrigel culture. The spheroids derived from Huh7-RFP-DCLK1 (upper panel) or Huh7-RFP (lower panel) were fixed in the matrigel culture and stained for active and total β-catenin, α-fetoprotein, and SOX9 (all shown in green). RFP-DCLK1 or RFP (red) was directly visualized without staining. Blue, nuclear stain with Dapi. Insets, highlighted for positive staining of each protein marker (green).

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