path4189-sup-0001-AppendixS1.docxWord document13KAppendix S1 Supplementary methods
path4189-sup-0002-figureS1.tifTIFF image1382KCo-staining of CK19 and PCNA confirms that cholangiocyte-like cells are proliferating within biliary tract lesions. Double immunofluorescence against the biliary epithelial marker cytokeratin 19 (CK19, red) and proliferating cell nuclear antigen (PCNA, green) allows ready identification of cells that are positive for both antigens. This confirms that cholangiocyte-like cells within lesions found in PtenF/F mice were undergoing proliferation and contributing to the lesion. Inset shows higher magnification
path4189-sup-0003-figureS2.tifTIFF image7348KFigure S1. BNF/TAM-treated KrasV12/+ and Ptenf/f; KrasV12/+ animals, but not Ptenf/f or WT animals, are susceptible to squamous hyperplasia of the forestomach, driven by AhCreERT. At dissection, no WT or Ptenf/f animals were found to bear abnormalities of the forestomach. In contrast, gross and abnormal enlargement of the forestomach was apparent in 55% of KrasV12/+ animals (six of 11 individuals) and 82% of Ptenf/f;KrasV12/+ animals (nine of 11 individuals) (A). The Rosa26R LacZ reporter was used to confirm that AhCreERT drives recombination in the squamous forestomach in response to BNF/TAM treatment. X-Gal wholemount staining of the forestomach revealed patches of blue, recombined cells within the squamous forestomach epithelium (B). Histologically, lesions arising in both KrasV12/+ and Ptenf/f;KrasV12/+ animals were found to be similar, with both genotypes showing diffuse papillomatosis of the forestomach with marked papillary hyperplasia of the squamous epithelium and florid luminal hyperkeratosis. Scale bars = 200 µm (C)

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