Two-stage carcinogenesis with rat embryo cells in tissue culture

Abstract

Summary.-Transformation of rat embryo fibroblasts in vitro has been investigated using initiation with either benzo(a)pyrene (BaP), 7,12-dimethylbenz(a)anthracene (DMBA) or benzo(e)pyrene (BeP) and promotion with either phorbol ester (TPA) or croton oil (Cr.Oil). The criteria used to assess in vitro transformation were (a) the efficiency of cloning in liquid medium, (b) abnormal cellular morphology and (c) the development of malignant tumours following s.c. inoculation of newborn rats.The results show that the cloning efficiency, which remained low in the control cells, was increased to a variable extent in the treated groups. Transformation occurred in all groups, but occurred earliest in cells that were initiated and promoted. Initiation with DMBA or BaP and promotion with TPA or Cr.Oil led to the earliest acquisition of malignancy. Correlations were found between the transformation of cells in vitro and the acquisition of malignant potential, and between the carcinogenic action of the compounds in vitro and their action in vivo, but cloning efficiency was not a reliable indicator of in vitro transformation or of malignancy. In most cases in vitro transformation appeared to precede the acquisition of malignancy, but in two cases it occurred later. The studies also show that BeP, which is a tumour initiator in vivo, also acts in this way in vitro. The conclusion drawn from a discussion of these results and of two -stage carcinogenesis in vivo is that two -stage carcinogenesis can be reproduced in tissue culture; this model may be useful in studies of those mechanisms of chemical carcinogenesis that involve the processes of initiation and promotion.

Two-stage malignant transformation in hamster embryo cells

“…In contrast to our observations, others have shown that when TPA was applied 3-5 days after carcinogen treatment in mouse cells or 7 days after carcinogen treatment in rat cells (Lasne et al, 1977) or rat cells (Lasne et al, 1977 Morphological changes appeared to reflect the initial events in transformation that are followed later by tumorigenesis and growth in soft agar. The studies reported here indicate that cells become tumorigenic before they acquire the ability to grow in soft agar (Tables I and II).…”

“…In vitro effects of 12-0-tetradecanoyl-phorbol-13-acetate (TPA) on cellular multiplication, RNA synthesis and DNA repair synthesis have been studied in mouse fibroblasts (Sivak & Van Duuren, 1970;Sivak et al, 1969Sivak et al, , 1972Trosko et al, 1975). More recently, TPA has been shown to enhance the focal transformation rate of mouse C3H/1OT1/2 cells and to enhance transformation in rat fibroblasts in culture (Lasne et al, 1974(Lasne et al, , 1977. The rate of focal transformation in mouse cells could be increased or decreased, depending * To whom requests for reprints should be sent. on the time at which TPA was administered relative to the carcinogen .…”

DNA synthesis inhibition in response to benzo[a]pyrene dihydrodiol epoxide is associated with attenuation of p34cdc2: Role of p53

“…Although BP carcinogenicity involves initial DNA damage, tumorigenicity is greatly enhanced by tumor promoters [11, 12], which interfere with these protective events [42, 43]. Cell cycle arrest gives cells time to repair DNA damage; apoptosis (programmed cell death) eliminates severely damaged cells.…”

Chronic Application of a Tumor Promoter to Confluent Nontransformed Rat Cells Induces the Irreversible Expression of Transformed Phenotype

“…This acquisition of'immortality' might be the only indication, over quite a long period of time in tissue culture, that some early events toward neoplas tic development had occurred. After extensive propagation in tissue culture, karyotypes may become unstable and cells may irreversibly acquire some or all transformation characteristics, including even that of tumor formation [15,16]. Thus, from an operational point of view, events leading to "immortalisation' might be considered as one type of "initiation" [ 17.…”