Extremely high production of thymidine by TNF-susceptible L929 cells

We confirmed with the use of crystal violet bioassay the high susceptibility of mouse L929 cells to the cytotoxic action of recombinant tumor necrosis factor-alpha (TNF). However, when a [3H]thymidine release assay was used for the same purpose, we found that [3H]thymidine uptake by the L929 cells, in contrast to low-malignant TNF-resistant spontaneously transformed Syrian hamster embryo cells of the STHE strain, was significantly reduced (P < 0.001). To investigate the mechanism of the low incorporation of [3H]thymidine in L929 cells the culture media from intact L929 cells was used for competition experiments with [3H]thymidine incorporation in the STHE target cells. The undiluted supernatant from L929 cells significantly (up to 83-97%) reduced [3H]thymidine uptake by the STHE cells. Fifty percent inhibition of [3H]thymidine uptake was achieved at L929 supernatant dilutions up to 1:8 (in 4 h incubation), up to 1:16 (in 20-42 h incubation) and even up to 1:32 (in 42 h incubation). The same high level of inhibition of [3H]thymidine uptake by STHE cells was seen with a commercial specimen of a thymidine (Sigma) at a concentration near 500 ng/ml. Thus, we conclude that a resistance of L929 cells to [3H]thymidine uptake is related to their unusually high production of cold thymidine.