TRANSFORMING-GROWTH-FACTOR BETA(1) INDUCES SQUAMOUS CARCINOMA CELL VARIANTS WITH INCREASED METASTATIC ABILITIES AND A DISORGANIZED CYTOSKELETON

Previous studies indicated that mouse transformed keratinocytes underg o an epithelial-fibro-blastic conversion when cultured in the presence of TGF-beta(1). This conversion is associated in vivo with a squamous -spindle carcinoma transition. We derived epithelioid (A6, FPA6) and s pindle (B5) clonal cell variants from a squamous carcinoma cell line ( PDV) after treatment with TGF-beta(1). FPA6 cells were isolated from t he ascites fluid of an AG-tumor-bearing mouse. FPA6 and A6 cell lines produced in nude mice mixed carcinomas with a squamous and poorly diff erentiated component. Both cell lines coexpressed keratins and vimenti n and synthesized E-cadherin protein, although FPA6 cells cultured at early passages (FPA6-ep) had reduced levels of E-cadherin mRNA and inc reased synthesis of keratin K8, a marker of malignant progression. Imm unofluorescence analysis revealed that FPA6-ep cells exhibited a disor ganized cytoskeleton with keratins forming focal juxtanuclear aggregat es and loss of F-actin stress fibers and cortical bundles, and E-cadhe rin was localized in the cytoplasm out of cell-cell contact areas. Spo radic cells in A6 and PDV cultures also presented those anomalous kera tin structures, suggesting that FPA6 cells originated from a subpopula tion of A6 tumor cells that metastasized into the peritoneal cavity. T he analysis of the spontaneous and experimental metastatic potentials of the cell lines showed that epithelioid and fibroblastic cell varian ts had acquired metastatic abilities compared to PDV which was nonmeta static. The FPA6-ep cell line exhibited a highly aggressive behavior, Billing the animals at about 17 days after intravenous injection of th e cells into athymic mice. The phenotype of FPA6-ep cells was unstable and reverted at later passages in which the normal organization of ke ratin and F-actin in filaments and the localization of E-cadherin at c ell-cell contacts were restored. This phenotypic reversion occurred co ncomitantly with a reduction of the experimental metastatic potential of FPA6 cells. (C) 1998 Academic Press.