Figure_6.tif (3.6 MB)

Characterization of primary pancreatic cancer cell lines from iKrasp53 mice.

figure

posted on 2013-02-19, 19:22 authored by Meredith A. Collins, Jean-Christophe Brisset, Yaqing Zhang, Filip Bednar, Josette Pierre, Kevin A. Heist, Craig J. Galbán, Stefanie Galbán, Marina Pasca di Magliano

(A) Primary cell line iKras*p53*-1 exhibits epithelial morphology and demonstrates doxycycline dependent Kras* expression (* p<0.05), and pERK1/2 levels at passage 5. (B) The same cell line at passage 12; Kras* expression is still dependent on doxy (*** p<0.001), but pERK1/2 levels do not depend on Kras* expression. (C) A second primary cell line, iKras*p53*-2, has mesenchymal morphology. At passage 6, Kras* expression is dependent on doxy (** p<0.01), and pERK1/2 levels depend on Kras* expression. (D) Analysis at passage 11: Kras* is still regulated by doxy (* p<0.05, ** p<0.01), but pERK1/2 levels remain elevated. (E) Western blot analysis of apoptosis, indicated by cleaved caspase-3 (CC3), and proliferation, measured by proliferating cell nuclear antigen (PCNA), in both iKras*p53*-1 and iKras*p53*-2 cell lines. (F) Immunofluorescence of apoptosis, indicated by cleaved caspase-3 (CC3), in iKras*p53*-2 cells either in the presence of (+48 h) or absence (−48 h) of doxy in the media. DAPI staining marks the nuclei. Scale bar 100 um. (G) Ras pull-down assay demonstrates that Ras activity levels are comparable between iKras*p53* cell lines and cells from KPC tumors.