![]() When FBP is added to normoxic and hypoxic cultured astrocytes and CO 2 and lactate 14 14 production are measured from glucose, glu- 14 cose, and, BP has little effect on CO 2 production by glycolysis, but increases CO 2 production by the PPP. While the activity of the PPP and the concentrations of NADPH are decreased by hypoxia, FBP stimulates the PPP during hypoxic conditions. Activity of this enzyme, in turn, depends on the presence of NADPH, which is produced in the PPP (Fig. Glutathione reductase activity is essential for GSH maintenance (Fig. To understand how FBP preserves GSH, we first determined whether FBP affects GSH regeneration from its oxidized form. #Bso death proof download free#By itself, FBP does not scavenge free radicals, but it decreases production of superoxide and it modulates mitochondrial function, possibly by maintaining higher concentrations of ATP. In fact, studies on superoxide production in neutrophils stimulated by phorbol myris- tate acetate show a lower production of superoxide and higher intracellular ATP concentrations when FBP is present. Neurons are very sensitive to decrease in the concentration of intracellular ATP and the ability of FBP in maintaining higher ATP concentrations in different cell types and brain slices, and 2 1 FBP’s ability to reduce increases in intracellular Ca during hypoxia can both decrease oxidant production. Intracellular GSH concentrations depend on the rate of GSH utilization, its synthesis, and its regeneration from its oxidized form. FBP may improve post-hypoxic GSH maintenance in several ways. Not surpris- ingly, the protective effect of FBP does not appear to be fully dependent on the preservation of GSH levels. Further, modulation of cellular antioxidative metabolism affects toxicity of this compound, causing more injury in the presence of exogenous SOD and elevated hydrogen peroxide levels. In the case of SIN-1, neuronal injury is mediated ? 2 not only by NO and O 2 radicals themselves, but also by the product of the reaction between these two radicals, peroxynitrite. Although both hypoxia–reoxygenation and SIN-1 induce oxidative stress, mechanistic components and injury propagation following these treatments are different. ![]() Following SIN-1 treatment, FBP preserves GSH concentrations from decreasing for at least 6 h after treatment, but FBP fails to alter the almost complete depletion of GSH following combined BSO and SIN-1 treatments. Despite the durations of hypoxia, GSH concentrations are significantly higher in neurons when FBP is present in the medium. During hypoxia, the concentration of intracellular GSH decreases significantly after a short period of hypoxia and even further when the hypoxic period is prolonged. The latter findings are consistent with our previous observations that 8 h of hypoxia causes neuronal death when astrocytes are absent, and that the addition of FBP to the medium does not prevent cell death any more. However, FBP fails to rescue neurons if the hypoxic episode is prolonged to 6 h when additional decrease of neuronal viability occurs. FBP also protects against another type of oxidative stress, hypoxia–reoxygenation, when the hypoxic period is short. While FBP does not alter neuronal viability when a mild injury is produced by application of SIN-1 itself, it protects neurons against more severe injury caused by pre-depletion of GSH prior to SIN-1 application. Since in a co-culture of neurons and astrocytes FBP reduces neuronal death partly via an astrocyte-mediated increase in glutamate uptake, glutamine production, and increased glutamine synthetase activity, we utilized almost pure neuronal cultures to avoid astrocyte-dependent effects in the present study. The protective effect of FBP is associated with preservation of GSH metabolism in neurons that likely occurs in part via increased glutathione reductase activity with a consequent increased GSH regeneration from its oxidized form. The current study shows that FBP protects neurons against hypoxia–reoxygenation and reactive oxidants directly, not through effects mediated by astrocytes. Under these conditions, FBP did not preserve GSH from falling (3.2 6 0.8%). Exposure of cells with pre-depleted GSH to SIN-1 resulted in further decrease of GSH over the value post-BSO treatment alone (2.6 6 1.8 vs. ![]() The presence of FBP in the medium during the SIN-1 exposure lead to a significantly higher GSH level than in cells not treated with FBP (60.5 6 6.9%, P, 0.045). ![]() While GSH level was not significantly different from the control levels 1.5 h after SIN-1 exposure and was not changed by the presence of FBP during the SIN-1 exposure, GSH level declined by 6 h post-treatment (47.2 6 9.3%, P, 0.0001). Neuronal GSH level reduced over time following the SIN-1 treatment for 5 min ( Fig. 6 2.6% of control when FBP was present during hypoxic period ( P, 0.05). ![]()
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