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Differences in Anchorage-dependent Growth and Tumorigenicities between Transformed C3H/10T Cells with Morphologies That Are or Are Not Reverted to a Normal Phenotype by Ascorbic Acid¹

Division of Hematology-Oncology, Department of Medicine, Childrens Hospital of Los Angeles, Los Angeles 90054 [W. F. B., W. L. W., P. A. J.], and Departments of Pediatrics [W. F. B., P. A. J.] and Biochemistry [P. A. J.], University of Southern California School of Medicine, Los Angeles, California 90027

C3H/10T mouse embryo cells were transformed with 3-methylcholanthrene. Several type III morphologically transformed cell lines were selected with morphologies that either could or could not be reverted back to normal at passage 1 by daily addition of ascorbic acid (1 or 5 µg/ml). Those transformed cell lines with morphologies that could be caused to revert to normal did not produce colonies in agarose or tumors in nude mice at early passages. Such transformed cell lines at later passages all formed colonies in agarose, but only 2 of 8 lines produced tumors at any passage tested. Subsequently, clones of transformed cells from each cell line have been isolated which are tumorigenic. In contrast, the transformed cell lines which were unresponsive to ascorbic acid at passage 1 were able to form colonies and to produce tumors in early passages. The reversion of the transformed morphology by ascorbic acid is apparently not caused by cytotoxicity since no cell kill was observed following exposure to ascorbate in any newly transformed cell lines at the concentrations used. Thus, the use of ascorbic acid allowed morphologically transformed cell lines to be isolated which appeared to be at different stages in the progression of an initiated cell from a morphologically transformed cell to a highly tumorigenic one. These studies also suggest that low concentrations of ascorbic acid in C3H/10T/CL8 cells can be effective in suppressing oncogenic progression only prior to a stage where an initiated cell achieves the capacity to grow in semisolid medium and to produce tumors in immunosuppressed animals. The importance of these cell lines for elucidating key changes required for the promotion and/or progression of cells to a tumorigenic phenotype is also presented.

¹ This work was supported by Grant CA-14226 from the National Cancer Institute.

Received 7/24/81. Accepted 12/ 1/81.

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