EFFECTS OF H2O2 ON THE GROWTH, SECRETION, AND METABOLISM OF HYBRIDOMACELLS IN CULTURE

The effect of low concentrations of hydrogen peroxide (H2O2) (5 X 10(- 7)-9.5 X 10(-7) M) on cell growth and antibody production was investig ated with murine hybridoma cells (Mark 3 and anti-hPL) in culture. Cel l growth, measured by flow cytometry with morphological parameters, wa s significantly stimulated by H2O2 (8 X 10(-7) M) but H2O2 concentrati on of X 10(-6) M and above increased cell death. H2O2 stimulation of a ntibody production was nonsignificant. The metabolism of cells treated with 8 X 10(-7) or 1 X 10(-5) M H2O2 was similar to that of the contr ol in terms of glucose and glutamine consumption, lactate and ammonia production, and amino acid concentrations in the medium. The concentra tions of lactate dehydrogenase, a marker of cell death, in test and co ntrol cells were similar. However, concentrations of intracellular fre e radicals measured by flow cytometry with dihydrorhodamine 123 (DHR 1 23) and dichlorofluorescein diacetate (DCFH-DA) as fluorochromes were different. The reactive oxygen species content of cells in 8 X 10(-7) M H2O2 was similar to that of the controls, but there was a sudden, ma rked production of superoxide anions (detected with DHR 123) and H2O2 or peroxides (detected with DCFH-DA) by cells incubated with 1 X 10(-5 ) M H2O2 which increased with increasing H2O2 until cell death.