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In-water pre-breathing oxygen at various depths reduces decompression-induced bubble formation and platelet activation, but it could induce side effects such as oxidative stress. The aim of this study was to investigate the effect of in-water pre-breathing oxygen, at different depths, on the oxidative status and intracellular calcium ([Ca2+]i) of peripheral blood lymphocytes isolated from six divers. They participated in a 4-diving protocol. Two week recovery time was allowed between successive dives. Before diving, all divers, for 20 min, breathed normally at sea level (dive 1), 100% oxygen at sea level (dive 2), 100% oxygen at 6 msw (dive 3), 100% oxygen at 12 msw (dive 4). Then they dived to 30 msw for 20 min with air tank.Blood samples were collected before and after each dive. Hydrogen peroxide (H2O2) levels, catalase (CAT) activity, mRNA expression of CAT, glutathione peroxidase (GPx) and superoxide dismutase (SOD), and the [Ca2+]i in lymphocytes were measured.The dives slightly decreased lymphocyte number and significantly reduced lymphocyte H2O2 levels. CAT activity was higher after scuba diving and, dive 3 enhanced mRNA gene expression of CAT, GPx and SOD. The [Ca2+]i was higher after dive 1 and 2 than pre-diving, while was maintained at pre-diving value after dive 3 and 4.Our results suggest that pre-breathing oxygen, in particular at 12 msw, may enhance lymphocyte antioxidant activity and reduce reactive oxygen species levels. Pre-breathing oxygen in water may also preserve calcium homeostasis, suggesting a protective role in the physiological lymphocyte cell functions.