Fig. 4: GASC1 promotes aberrant nuclear structure and cell-cycle arrest of leukemia cells.
From: Chromatin-modifying enzymes as modulators of nuclear size during lineage differentiation
A Representative images of DAPI staining for K562 cells treated with DMSO as control and 5-Aza, IOX1, GSK-LSD1, and JIB04. It was shown that the bigger size and twisted shape of nuclei are in inhibitors treatment. The images were taken at 40× on a confocal microscope. White scale bars = 25 μm. B Measurements of the nuclear size using DAPI staining under fluorescence microscopy after K562 cells treated with inhibitors of chromatin-modifying enzymes. Two-tailed t-test, mean ± SD. **P < 0.01, ***P < 0.001. C Brightfield and fluorescent images show that K562 leukemia cells differentiate to iN cells with Ngn2 overexpression after 14 days differentiation and stain with the neuronal marker Tuj1 (red), compared to healthy and as cultured K562 cells without any treatment. The yellow arrows indicate the cells of morphological changes. D, E Brightfield images of K562 leukemia cells with induced expression of GASC1 with or without doxycycline (dox). The no dox-induced GASC1 overexpressed K562 cells show spherical morphology, same as normal cells. The dox-induced GASC1 overexpressed K562 cells show morphology change from spherical to twisted shape and cell blebs. The red arrow indicates the cells with twisted shapes and blebs. White scale bars = 25 μm. F, G Fluorescent images by DAPI staining demonstrate that the dox-induced GASC1 overexpressed K562 cells show the twisted nuclear shape and bigger nuclei compared with the no dox-induced GASC1 overexpressed K562 cells. The red arrow indicates the enlarged and twisted nucleus. White scale bars = 25 μm. H, I Flow cytometric analysis of K562 cells were double-stained with BrdU and PI. The dox-induced GASC1 overexpressed K562 cells proliferate significantly slower than the no dox-induced GASC1 overexpressed K562 cells in the BrdU assay. J Cell number counting after induction by dox for 7 days. The cell number is decreased in dox-induced GASC1 overexpressed K562 cells than no dox-induced GASC1 overexpressed K562 cells. Values represent mean ± SD (n = 3). ***P < 0.001. K The genes play a role in cell-cycle arrest detected by qPCR in the dox-induced GASC1 overexpressed K562 cells and no dox-induced GASC1 overexpressed K562 cells. Unpaired two-tailed Student t-test. Data are mean ± SD. ***P < 0.001.
