Journal: Neoplasia (New York, N.Y.)

Article Title: Type III TGF-β receptor enhances colon cancer cell migration and anchorage-independent growth.

doi: 10.1593/neo.11528

Figure Lengend Snippet: Figure 3. TβRIII expression increases colon cancer in vitro tumorigenicity. (A) HT29-Neo and TβRIII cells were treated with 20 or 40 nM BMP-2 for 24 hours. Proliferation was analyzed by a 3H incorporation assay. The percent proliferation was determined by normalizing the counts to those of untreated samples. (B) HT29-Neo and TβRIII cells were treated with 50 or 100 pM TGF-β1 for 24 hours. Proliferation was analyzed by a 3H incorporation assay. (C) HT29-Neo and TβRIII cells were treated with 2 or 10 nM BMP-2 and 50 or 100 pM TGF-β1 for 24 hours. HT29-Neo and TβRIII cells were treated with 200 μM FTS or DMSO for 3 days. Western blot analyses were performed to analyze protein levels of p21, p27, p15, cyclin D, and Ras with β-actin as a total protein control. (D) HT29-Neo and TβRIII cells were plated in a soft agar assay untreated or treated with 20 nM BMP-2, 40 nM BMP-2, 50 pM TGF-β, or 100 pM TGF-β for 21 days. The mean percent colony formation ± SEM is shown normalized to the untreated Neo or TβRIII. Average colony number is shown above the bar graph. (E) HT29-Neo and TβRIII cells were treated with 40 nM BMP-2 or 100 pM TGF-β for 48 hours and examined for apoptosis by Western blot analysis of caspase 9 levels. Densitometric analysis is shown normalized to β-actin. (F) HT29-Neo and TβRIII cells were treated with 50 μM 5-fluorouracil for 48 hours. Cells were concurrently treated with 20 nM BMP-2 or 100 pM TGF-β and examined for apoptosis induction by Western blot analysis of PARP cleavage. Densitometric analysis is shown normalized to β-actin.

Article Snippet: Western blot analyses were performed to analyze protein levels of p21 (no. 2946; Cell Signaling Technology) and p27 (no. 2552; Cell Signaling Technology) protein level with β-actin as a total protein control.

Techniques: Expressing, In Vitro, Western Blot, Control, Soft Agar Assay

Journal: Cell Death & Disease

Article Title: Lysosomes, caspase-mediated apoptosis, and cytoplasmic activation of P21, but not cell senescence, participate in a redundant fashion in embryonic morphogenetic cell death

doi: 10.1038/s41419-023-06326-6

Figure Lengend Snippet: In all graphs, the percentage of dead cells is represented by red columns, and the percentage of SA-β-Gal-positive cells is represented by blue columns. Treatments were administered for 12 h after 30 h of culture. The values in control micromasses (treated only with drug vehicle) were considered 100%. Cell death was measured by flow cytometry using propidium iodide. SA-β-Gal-positive cells were quantified by histological examination. A Bars from left to right respectively represent micromasses treated with control conditions; 20 µM Q-VD-OPh; 5 µM Pepstatin A; B combination of 20 µM Q-VD-OPh plus 5 µM Pepstatin A. C Cultures treated with 5 µM UC2288. D Cultures treated with 5 µM UC2288 plus 20 µM Q-VD-OPh (left bar), and 5 µM UC2288 plus 5 µM Pepstatin A (right bar). E Representative flow cytometry plots (propidium iodide) of control and 0.25 µM Navitoclax treated cultures showing differences in the intensity of death cells (left images). Bars show the fraction of apoptotic cells with fragmented DNA in the treated and control cultures. The image on the right is a graphic representation of the incidence of dead cells (red columns) and the number of SA-β-Gal-positive cells in cultures treated with 0.10 µM, 0.15 µM, and 0.25 µM Navitoclax. The dotted line represents values in control conditions. Statistical significance: *** p < 0.001; ** p < 0.01; * p < 0.05.

Article Snippet: UC2288 (Merk) was employed as a p21 inhibitor that attenuates p21 expression at both at the transcriptional and protein levels and lacks effects on AKT and ERK signalling pathways [ ].

Techniques: Control, Flow Cytometry