Intracellular levels of H2O2 in BHK-21 cells are not static but decline progressively with cell growth. Exposure of cells to inhibitors of catalase, or glutathione peroxidase, not only diminishes this decline but also depresses rates of cell proliferation, suggesting important growth regulatory roles for those antioxidant enzymes. Other agents which also diminish the growth-associated decline in intracellular levels of H2O2, such as the superoxide dismutase mimic, copper II-(3,5-diisopropylsalicylate)2, or docosahexaenoic acid, also reduced cell proliferation. In contrast, proliferation can be stimulated by the addition of 1 microM exogenous H2O2 to the culture medium. Under these conditions, however, intracellular levels of H2O2 are unaffected, whereas there is a reduction in intracellular levels of glutathione. It is argued that critical balances between intracellular levels of both H2O2 and glutathione are of significance in relation both to growth stimulation and inhibition. In addition growth stimulatory concentrations of H2O2, whilst initially leading to increased intracellular levels of lipid peroxidation breakdown products, appear to "trigger" their metabolism, possibly through aldehyde dehydrogenase, whose activity is also stimulated by H2O2.

Addition of H2O2 at 100 microM, or 1 mM, to the culture medium of BHK-21 fibroblasts results in increased intracellular levels of H2O2. Whilst exposure of BHK-21 cells to lower levels of H2O2 (1 microM) actually stimulates proliferation, these higher oxidant concentrations not only depress proliferation rates but also lead to an increase in the appearance of apoptotic-like cells in the cultures. Other agents such as inhibitors of glutathione peroxidase and catalase, or mimics of superoxide dismutase, which also bring about elevated cellular levels of H2O2 in BHK-21 cells, similarly lead to decreased proliferation and an apparent increase in cells with apoptopic features. Thus intracellular conditions which are considered more prooxidant than normal, appear to favour apoptosis over proliferation in BHK-21 fibroblasts. Additionally these abnormal cellular conditions also appear to favour excessive DNA replication, in remaining non-apoptotic cells.

In BHK-21 cells (baby hamster kidney fibroblasts) cellularly generated active oxygen species such as hydrogen peroxide and superoxide appear to be important growth regulatory signals as judged from the growth inhibitory effects of catalase, superoxide dismutase and superoxide dismutase mimics. These active oxygen species may contribute a novel redox system of regulatory control superimposed upon established growth signal pathways. This may be achieved by direct oxidative modification of cell regulatory proteins such as transcription factors or protein kinases or indirectly through, for example alterations in levels of glutathione (GSH). This latter possibility is suggested from observations that catalase, or superoxide dismutase treatment of BHK-21 cells brings about increased cellular levels of GSH. However during the normal growth phase cellular levels of GSH actually decline although this effect can be partly reversed by N-acetylcysteine and by mercaptosuccinate which also impair proliferation of these cells.