Fig. 3.

Three-dimensional (3D) matrix culture of HK-2 cells. A: HK-2 cells plated at a density of 100,000 cells/cm² were overlaid with collagen matrix and grown up to 14 days after overlay. Phase-contrast images are presented on the left (A, C, E, G, and I). Composite extended focus images of fluorescent image set labeled for actin and nuclei are on the right (B, D, F, and H). A single confocal fluorescent image plane is shown in J. A and B: phase-contrast image of the tubule structures and phalloidin staining of F-actin (red) with nuclei staining (Hoescht, blue), to show tubule structures and cell organization within tubules in control untreated cells. C and D: 3D culture at 48 h after adenovirus microinjection with adenovirus expressing the tet transactivator only. Rare knoblike extension similar to that seen in control 3D cultures were seen. E and F: adenovirus expressing cadherin 8 was microinjected and the cell culture was fixed 48 h later. No cystic structures were observed, but again some rare knoblike structures were found. G and H: adenovirus expressing cadherin 8 and tet transactivator were coinjected at a multiplicity of infection (MOI) of 1E6. Free cysts and stalk structures were observed in the phase-contrast image (arrow and arrowhead, respectively). I and J: phase-contrast image and a single image plane from the data set. The arrow points to a cyst with a small lumen. Actin staining with phalloidin is shown using gray scale instead of a red scale. Bar, 135 μm.