Skip to main content
Fig. 3 | Journal of Biomedical Science

Fig. 3

From: A chemical probe inhibitor targeting STAT1 restricts cancer stem cell traits and angiogenesis in colorectal cancer

Fig. 3

In vitro and in vivo binding of THIF to STAT1. A THIF blocks the STAT1-STAT1 interaction in CRC cells. HCT116 cells were treated with 50 μM THIF or vehicle (control) for 6 h, and proteins were isolated and immunoprecipitated with the indicated antibodies. B Schematic diagram of the STAT1 constructs (upper panel). The indicated plasmids (3 μg) were transfected into 293 T cells. Twenty-four hours later, the cells were treated with 50 μM THIF for 6 h. After incubation, cells were collected and used for IP experiments. Western blot analysis was performed using the indicated antibodies (lower panel). C Bright-field images and fluorescence images of BODIPY-THIF (5 mM) were acquired with an IVIS imaging system (left panel). Chemical structure of fluorescently labeled THIF (BODIPY-THIF) (right panel). The core structures of the BODIPY fluorophore are highlighted in red. D The cellular distribution of BODIPY-THIF in CRC cells. HCT116 cells were treated with BODIPY-THIF (50 μM) for 6 or 12 h and were then subjected to fluorescence microscopy to detect BODIPY-THIF (red fluorescence) and DAPI (blue fluorescence). Representative fluorescence images of the cells; the inset shows higher-magnification images of the areas in the yellow boxes, as indicated. The scale bar is 100 μm. E Fluorescence images of BODIPY-THIF in HCT116 cells were quantitatively analyzed with ImageJ software. F Colocalization of STAT1 and BODIPY-THIF in CRC cells. HCT116 cells were either treated with BODIPY-THIF (50 μM) alone for 12 h or pretreated with fludarabine (10 μM) for 30 min prior to BODIPY-THIF treatment and subjected to immunofluorescence staining for BODIPY-THIF (red fluorescence), STAT1 (Alexa Fluor 488, green fluorescence) and DAPI (blue fluorescence). Representative fluorescence images of the cells; the inset shows higher-magnification images of the areas in the yellow boxes, as indicated. Merged images are shown. The colocalization signal is colored yellow. Scale bar: 50 μm. G The colocalization of BODIPY-THIF and STAT1 in HCT116 cells was quantitatively analyzed with ImageJ software. ***p < 0.001

Back to article page