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Fig. 1. Tumor nodules display altered tissue polarity and nuclear organization compared with acini formed by non-neoplastic cells. S1 and T4-2 cells were cultured in 3D in the presence of MatrigelTM for 10 days and labeled for differentiation markers collagen IV, 
6-integrin, 
-catenin, mucin-1 and Ki-67, and markers of nuclear organization H3K9m, H4K20m, NuMA, SC35, PML and lamin B. (A) Immunostaining for basal polarity markers collagen IV (green) and 6-integrin (green), lateroapical polarity marker -catenin (red), and apical polarity marker mucin-1 (red) in acinar S1 cells (top panel) and malignant T4-2 cells (bottom panel). Drawings show the organization of S1 and T4-2 cells after 10 days of 3D culture; nuclei are represented in black. (B) Immunostaining for cell cycle marker Ki67 (green). Nuclei are counterstained with DAPI (blue). Bar graph shows the percentage of cells in the cell cycle based on Ki67 expression. (C) Organization of heterochromatin markers H3K9m (red) and H4K20m (red) in the nuclei of acinar (S1) and malignant (T4-2) cells. Arrows indicate the concentration of H3K9m and H4K20m around the nucleolus (*) in differentiated cells. (D) Organization of coiled-coil protein NuMA (green), splicing factor speckle marker SC35 (green), PML (red, with DAPI-counterstained nuclei in blue) and lamin B (red) in the nuclei of acinar (S1) and malignant (T4-2) cells. The arrowhead indicates the formation of enlarged NuMA foci in mid-nucleus in S1 cells. SC35 is organized into large domains in the nucleus of S1 cells (dotted line delineates the cell nucleus). The arrow points to the twisted shape of the malignant nucleus as shown by lamin B staining. Bars, 5 µm.
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