Effect of tunicamycin, an inhibitor of protein glycosylation, on division of tumour cells in vitro

We determined the effect tunicamycin (TM), an inhibitor of protein glycosylation, had on cells in vitro that were derived from solid and ascites variants of a chemically induced rat hepatoma. Using flow microfluorometry (FMF), labelling index (LI), and population-doubling time assays, we monitored the progression of cells through the cell cycle after treatment with TM. Cells in monolayer culture were first incubated in 0.05 or 0.10 micrograms TM/ml medium for 24 h then analysed or given fresh medium without TM and allowed to recover for 6–24 h. Exposing cells to 0.05-0.50 micrograms TM/ml medium did not affect the percentage of viable cells as determined using the Trypan Blue exclusion procedure. However, continuous exposure to 0.05 micrograms TM/ml medium did affect the population-doubling times of both the ascites and solid variants, and the ascites tumour cells were more sensitive than the solid tumour cells. TM reversibly inhibited hepatoma cells from entering S phase of the cell cycle. After exposure to TM for 24 h, the percentage of solid tumour cells in vitro in S phase decreased to 19%, as determined by autoradiography of tritiated-thymidine-labelled cells, and to 21% as determined by FMF; 49% of untreated solid tumour cells were in S phase. The percentage of ascites tumour cells in vitro decreased to 12% after exposure to TM for 24 h; 37% of untreated cells were in S phase. We concluded that TM can inhibit division of rat hepatoma cells in vitro by blocking them in G1 phase of the cell cycle.