Red blood cell (RBC) susceptibility to oxidative and osmotic stress in vitro was investigated in cells from trained and untrained men before and after submaximal exercise. Whilst no significant change in peroxidative haemolysis occurred immediately after 1 h of cycling at 60% of maximal aerobic capacity (VO2max), a 20% increase was found 6h later in both groups (P < 0.05). The RBC osmotic fragility decreased by 15% immediately after exercise (P < 0.001) and this was maintained for 6h (P < 0.001). There was an associated decrease in mean cell volume (P < 0.05). Training decreased RBC susceptibility to peroxidative haemolysis (P < 0.025) but it did not influence any other parameter. These exercise-induced changes were smaller in magnitude but qualitatively similar to those found in haemopathological states involving haem-iron incorporation into membrane lipids and the short-circuiting of antioxidant protection. To explore this similarity, a more strenuous and mechanically stressful exercise test was used. Running at 75% VO2max for 45 min reduced the induction time of O2 uptake (peroxidation), consistent with reduced antioxidation capacity, and increased the maximal rate of O2 uptake in RBC challenged with cumene hydroperoxide (P < 0.001). The proportion of high-density RBC increased by 10% immediately after running (P < 0.001) but no change in membrane-incorporated haem-iron occurred. In contrast, treatment of RBC with oxidants (20-50 mumol.l-1) in vitro increased cell density and membrane incorporation of haem-iron substantially. These results showed that single episodes of submaximal exercise caused significant changes in RBC susceptibility to oxidative and osmotic stress. Such responses may account for the increase in RBC turnover found in athletes undertaking strenuous endurance training.

We investigated whether uptake of lactate anions by red blood cells (RBC) during maximal exercise altered certain cellular physical properties including volume, density and osmotic fragility. Irrespective of whether individuals undertook 1 min of exercise at maximal effort or a progressive exercise test to exhaustion, whole-blood lactate concentration increased about tenfold relative to pre-exercise values (P < 0.001). The plasma and cellular concentrations of lactate increased uniformly, which maintained a constant plasma-to-cell gradient. Exercise-induced increases in RBC osmotic fragility (P < 0.001) and mean cell volume (MCV; P < 0.05), and decreases in mean cell haemoglobin concentration (MCHC; P < 0.05) and cell density (P < 0.005) also occurred. These exercise-induced cellular responses could be simulated in vitro by treating blood with L-lactic acid (15 mM), but not by low pH (7.1) or L-lactate alone. The uptake of lactate by RBCs during vigorous exercise alters certain cellular properties. Although the physiological implications of these changes are unclear, the traditional view that the lactate anion has no known harmful effects should be reconsidered.

Sendai virus (150 haemagglutinating units/10(6) cells) stimulates rat thymocytes incubated in medium containing 5 mM-glucose at 37 degrees C to produce luminol-dependent chemiluminescence and a simultaneous increase in O2 consumption of 40%. Stimulation of thymocytes with Sendai virus is accompanied by reduction of exogenous acetylated ferricytochrome c, which is inhibited by superoxide dismutase, and the quantitative conversion of ferricyanide to ferrocyanide, which is not. Replacement of air in the gas space with N2 inhibits the chemiluminescent response by 97% but does not prevent the virus-stimulated reduction of ferricyanide. The non-permeant ferricyanide anion (2 mM) also inhibits the chemiluminescent response to Sendai virus, its accompanying 'extra' O2 uptake and the reduction of acetylated ferricytochrome c without affecting the basal respiration of the cells. Thymocytes in which the basal O2 consumption has been stimulated maximally with dinitrophenol (10 microM) or inhibited completely with antimycin A (0.1 microM) respond to Sendai virus with an additional increment of ferricyanide-inhibitable O2 consumption. The chemiluminescent response to virus is not inhibited by concentrations of antimycin A that block the basal respiration completely. We suggest that a portion of the increased O2 uptake induced by Sendai virus is involved in the non-mitochondrial reduction of O2 to O2- at the cell surface where the non-permeant ferricyanide anion inhibits O2-. formation by acting as an alternative high-affinity electron acceptor to O2.

1. The luminol-dependent chemiluminescence of rat thymocytes responding to concanavalin A can be resolved into glucose-dependent and glucose-independent portions. 2. The glucose-dependent portion, supported by D-glucose and D-mannose oxidation, is inhibited by catalase (200 microgram/ml), amobarbital (1 mM) and hexose analogues that block D-glucose uptake. Thus concanavalin A may activate, transiently, an NAD(P)H oxidase that utilizes reducing equivalents derived from the oxidation of exogenous glucose to give dismutation products of O2- (including H2O2) as its major products. 3. The glucose-independent portion is inhibited by eicosa-5,8,11,14-tetraynoic acid but not by indomethacin. It may therefore be associated with the conversion of hydroperoxy intermediates of arachidonic acid metabolism to hydroxy products by the lipoxygenase pathway. 4. Preincubation of thymocytes for 18 h in serum-free medium enhances the subsequent chemiluminescent response to concanavalin A severalfold and evokes the response at a lower threshold concentration. The incorporation of [3H]thymidine by preincubated cells is similarly enhanced at low doses of concanavalin A, whereas the response to optimal doses is unaltered. 5. Catalase does not inhibit the enhanced incorporation of [3H]thymidine obtained in response to concanavalin A, but instead amplifies the response to low doses in the same manner as preincubation.

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