Parameters Distinguishing Herpes Simplex Virus Type 2-transformed Tumorigenic and Nontumorigenic Rat Cells¹

A newly developed experimental model system was used to determine in vitro transformation-specific parameters which correlate with tumorigenicity. The data suggested that clonal herpes simplex virus type 2-transformed syngeneic rat embryo cells with intermediate, transformed rat embryo fibroblasts (t-REF-G-1) or high, rat fibrosarcoma tumorigenic potential in syngeneic rats could be differentiated from clonal transformed nontumorigenic (t-REF-G-2) and nontransformed rat embryo fibrobiast cells by their growth to increased saturation density and cloning efficiency in soft-agar medium. All clonal herpes simplex virus type 2-transformed cells, regardless of tumorigenic potential, possessed an increased rate of hexose transport and plasminogen activator activity and were less fibroblastoid in morphology compared to nontransformed rat embryo fibroblast cells. There was no significant difference in cell-doubling time or total phospholipid composition between clonal transformed tumorigenic, nontumorigenic, and nontransformed cells. However, all clonal herpes simplex virus type 2-transformed cell lines possessed a decreased percentage of arachidonic acid and an increased percentage of monounsaturates compared to percentages in nontransformed rat embryo fibroblast cells. Saturation density, cloning efficiency, and tumorigenicity in newborn and adult rats of one nontumorigenic clonal cell line (t-REF-G-2) increased with cell passage in tissue culture. Other transformation-specific biochemical parameters (hexose transport, plasminogen activator activity, and fatty acid composition) remained stable with in vitro passage of the clonal cell lines. Five subclonal cell lines established from 12-O-tetradecanoylphorbol-13-acetate-treated clonal t-REF-G-2 (low passage) cells showed increased cloning efficiencies compared to 2 subclonal cell lines from mock-treated cells; also, all 5 subclonal cell lines from 12-O-tetradecanoylphorbol-13-acetate-treated clonal cells were tumorigenic in newborn rats. Subclonal cell lines from mock-treated clonal cells were non-tumorigenic. It was clear from the results that the absolute cloning efficiency did not correlate with tumorigenicity. This is an important conclusion, since it suggests that quantitation of transformation-specific characteristics at a given point in time will not necessarily be indicative of the tumorigenic potential of the cells.