Mechanism of growth promotion of mouse lymphoma L1210 cells in vitro by feeder layer or 2‐mercaptoethanol

Mouse lymphoma L1210 cells require some thiol compounds (such as 2-mercaptoethanol) or feeder layer cells for their growth in normal culture media in vitro. We found that feeder layer cells (human diploid fibroblast IMR-90) constantly produce thiol compounds; cysteine was the major thiol compound accumulating in the culture medium. In the culture medium of L1210 cells, added cysteine was rapidly oxidized and was toxic to the cells at high concentrations. However, cysteine promoted growth of L1210 cells when it was added repeatedly to the medium at low concentrations. These results show that the major role of feeder layer cells is to provide cysteine continuously. The glutathione content of L1210 cells depended largely on the cysteine concentration in the medium. In normal culture media containing cystine but not cysteine, the cellular glutathione content decreased notably within a few hours. Cysteine had to be supplied repeatedly to keep the content high. Cystine promoted the cellular glutathione content at unphysiologically high concentrations. These results were attributable to the extremely low uptake rate of cystine by the cells as compared with that of cysteine. In the presence of 2-mercaptoethanol, the uptake of radioactive cystine by the cells was increased and a high cellular glutathione level was maintained. A thiol-independent variant of L1210 took up cystine far more rapidly than L1210. From these results we concluded that the deficiency of the cystine uptake limits the growth of L1210 cells in normal culture media.