Actin cytoskeletal organization loss in the benign-to-malignant tumor transition in cultured human colonic epithelial cells

The colonic epithelium in vivo is a highly indented sheet one cell thick. Culture methods have been developed to allow the normal cellular migration of the cells comprising this sheet to flatten it into a patch on the surface of a Petri dish [Friedman, E. A., Higgins, P.J., Lipkin , M., Shinya , H., and Gelb , A.M., In Vitro (Rockville), 17: 632-644, 1981]. Actin cytoskeletal organization was analyzed in such epithelial "patches" derived from several human colonic adenocarcinomas and their precursors, adenomas (benign tumors). The actin cytoskeleton was visualized by fluorescence microscopy after the fixed, permeabilized cells were stained with rhodamine-conjugated phalloidin. This drug has a very high affinity for actin filaments and a much lower affinity for monomeric actin. Actin organization was scored from 0 (no cables) to 5 points (extensive intercellular cable network). The phalloidin-stained actin found in seven adenocarcinomas had a predominantly granular fluorescence pattern with very little cable organization, scoring an average of 0.9 +/- 0.8 (S. D.). Three established cell lines derived from human colon carcinomas contained no cables by this analysis, scoring 0.0 +/- 0.0. In marked contrast, all 12 of the cultured adenomas had extensive actin cable networks, scoring an average of 4.3 +/- 0.4. There was no statistical difference between adenomas of differing histopathology class and malignant potential. However, cytoskeletons of plasminogen-activator-secreting "late-stage" preneoplastic cells from adenomas became disorganized by exposure to 12-O-tetradecanoyl-phorbol-13-acetate or another tumor promoter, teleocidin B. They scored, respectively, average actin organization values of 0.0 +/- 0.0 and 0.4 +/- 0.6. In contrast, nonplasminogen -activator- secreting "early-stage" preneoplastic cells from less advanced benign tumors were unaffected by 12-O-tetradecanoyl-phorbol-13- acetate or teleocidin B and retained extensive actin organization. Most, if not all, adenocarcinomas arise from preexisting preneoplastic adenomatous cells. Thus, loss of actin organization appears to mark the transition of noninvasive benign colonic tumors to invasive malignant tumors in humans. This transition is mimicked in vitro by exposure of certain "late-stage" preneoplastic cells to a tumor promoter which induces secretion of a plasminogen activator.