Combined growth-inhibitory responses and ultrastructural alterations produced by 1,3-bis(2-chloroethyl)-1-nitrosourea and dexamethasone in rat glioma cell cultures.

The effect of 0.0001 to 10 muM 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and 1 muM dexamethasone on cell proliferation was studied by measuring cell densities in control and drug-treated rat glioma (strain C6) monolayer cultures. When C6 cultures were exposed to 0.01 to 10 muM BCNU, the growth rates decreased for 2 days as control cell populations continued to proliferate at log phase rates. These growth-inhibitory responses were dose dependent and ranged from 20 to 80%, relative to control growth. Subsequently, the growth rates increased and the inhibitory responses ranged from 0 to 12% 4 days later. Cell densities in C6 cultures exposed to 1 muM dexamethasone for 1 day did not differ significantly from controls. Then cell proliferation ceased and the inhibitory response remained at 50% relative to controls in stationary phase. When 0.03 muM BCNU and 1 muM dexamethasone were supplied simultaneously to C6 cultures, a 35% inhibitory response occurred after 1 day. This response did not differ significantly from that observed with 0.03 muM BCNU alone. After 4 days, the inhibitory response did not decrease in cultures containing both drugs, but did decrease to 13% in the 0.03 muM BCNU-treated cultures. In 1 muM BCNU-treated cultures, the response was 66% after 1 day, which decreased to 21% 5 days later. When 1 muM BCNU was supplied to C6 cultures that were pretreated for 1 day with 1 muM dexamethasone, the response was 91% the following day, and this decreased to only 54% 5 days later. Dose-response curves showed that the inhibitory responses after 1 day in these pretreated cultures exposed to 0.001 to 10 muM BCNU increased up to 22% relative to the responses produced by either drug alone. After 5 days, the responses in the pretreated cultures exposed to 0.001 to 1 muM BCNU was 50%, which was similar to the response produced by 1 muM dexamethasone alone. Ultrastructural studies revealed that control and 1 muM BCNU-treated C6 cells contained 18 mitochondria, but the treated cells were 10% smaller after 1 day. Cells exposed to 1 muM dexamethasone for 1 day conount of granular endoplasmic reticulum increased greater than 80% in cells treated with BCNU for 1 day or dexamethasone for 2 days. C6 cells pretreated with dexamethasone and exposed to BCNU for an additional day (a) contained 23 mitochondria, (b) did not decrease in size, and (c) exhibited a greater than 250% increase in the amount of granular endoplasmic reticulum. These results demonstrate that combined growth-inhibitory responses and ultrastructural alterations occur when C6 cells are treated sequentially with 1 muM dexamethasone and BCNU.