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. 2019 Sep 13;18(12):2373–2387. doi: 10.1074/mcp.RA119.001455

Fig. 5.

Fig. 5.

Proposed mechanism of DDW action and its validation. A, Interaction network of proteins most involved in DDW action mechanism according to summary of proteomics analysis. B-D, Proposed mechanism: B, In normal conditions, the deuterium concentration in mitochondrial matrix water is lower than in the intermembrane space water. C, Lowering deuterium concentration in ambient water reverses the deuterium gradient across the inner membrane. To restore the H/D equilibrium, mitochondria increase export of protons from the matrix to intermembrane space, which causes an increase in the mitochondrial membrane potential (↑+). This in turn promotes production of ROS as energy-carrying electrons are passed by ETC from NADH and FADH2 to oxygen, the final electron acceptor. Mitochondrial and cellular oxidative stress results, with slower growth and eventual apoptosis. D, Further lowering of deuterium concentration in ambient water causes too great an increase in ROS production to be controlled by intrinsic enzymatic antioxidant mechanisms. An additional antioxidant mechanism is triggered, in which activation by superoxide (O2●−) of UCPs combats enhanced ROS production. In this feedback model, the activated UCPs in the inner mitochondrial membrane open greater proton influx into the matrix, with concomitant reduction in the mitochondrial membrane potential (↓+), which leads to diminished O2●− production and near restoration of the mitochondrial and cellular redox equilibrium, with reduced cellular oxidative damage. E-H, Validation of the DDW-induced redox disbalance mechanism. E, Combined effect of 80 ppm DDW with addition of 3 μm auranofin on ROS level compared with 80 ppm DDW or 3 μm auranofin alone. F, Time course analysis of relative ROS production in the cells treated with 3 μm auranofin, grown in vDDW and DDW (80 ppm) compared with untreated cells grown in NW. G, Effect on cell count of NAC addition to DDW- or auranofin-treating cells. H, Effect on cell count of simultaneous DDW and auranofin application compared with auranfin only. G-H show the mean ± SEM of nine independent experiments with four replicates, * p < 0.05, ** p < 0.01, *** p < 0.005, **** p < 0.0001 in two-tailed unparied t test.