Miguel D. Ferrer

Acute Administration of Inorganic Nitrate Reduces V˙o2peak in Endurance Athletes

PURPOSE Humans can reduce inorganic nitrate (NO(3)(-)) to nitrite (NO(2)(-)), nitric oxide (NO), and other bioactive nitrogen oxides. The purpose of this study was to test the hypothesis that a single dose of inorganic nitrate before exercise might enhance the tolerance of endurance athletes to high intensity exercise. METHODS Eleven cyclists (age = 34.3 ± 4.8 yr, VO(2peak) = 65.1 ± 6.2 mL·kg(-1)·min(-1)) participated in this randomized, double-blind, crossover study. Subjects received dietary supplementation with nitrate (NaNO(3) 10 mg·kg(-1) of body mass) or a placebo (NaCl) 3 h before exercise. They then performed a cycle ergometer test that consisted of four 6-min submaximal workloads, corresponding to 2.0, 2.5, 3.0, and 3.5 W·kg(-1) of body mass, interspersed with 3 min of passive recovery. After a 5-min recovery period, subjects performed one incremental exercise test until exhaustion. RESULTS Plasma nitrate and nitrite were significantly higher (P < 0.05) 3 h after supplementation (nitrate = 250 ± 80 μM, nitrite = 2313 ± 157 nM) than after the placebo (nitrate = 29 ± 8 μM, nitrite = 1998 ± 206 nM) at resting conditions. Nitrate supplementation significantly reduced VO(2peak)(nitrate = 4.64 ± 0.35 L·min(-1), placebo = 4.82 ± 0.33 L·min(-1), P = 0.010) and the ratio between VO(2) and power at maximal intensity (nitrate = 11.2 ± 1.1 mL·min(-1)·W(-1), placebo = 11.8 ± 1.1 mL·min(-1)·W(-1), P = 0.031). This reduction of VO(2) occurred without changes in the time to exhaustion (nitrate = 416 ± 32 s, placebo = 409 ± 27 s) or in the maximal power (nitrate = 416 ± 29 W, placebo = 410 ± 28 W). CONCLUSIONS A single oral dose of inorganic nitrate acutely reduces VO(2peak)without compromising the maximal exercise performance.

Antioxidant regulatory mechanisms in neutrophils and lymphocytes after intense exercise

Abstract The aims of this study were to assess the effects of a swimming session on the peripheral blood neutrophil and lymphocyte pro- and antioxidant system, identify any differences between the sexes and the regulatory mechanisms that might induce the immune cell adaptive response to exercise. Twenty-four swimmers (15 males, 9 females) participated in a one-hour swimming session at 75–80% of their maximal capacity. The session induced neutrophilia and decreased antioxidant enzyme activities and ascorbate levels in neutrophils. Malondialdehyde rose in neutrophils in males and females, whereas the carbonyl index only increased in males. Lymphocyte glutathione peroxidase activity was higher in males at baseline and rose as a consequence of exercise. The exercise decreased uncoupling protein-3 and Bcl-2 gene expression. The expression of PPARγ coactivator-1 alpha (PGC-1α) correlated positively with that of sirtuin 3 (SIRT3) and catalase. In summary, a swimming session of one hour at 75–80% of maximal capacity produced oxidative damage in neutrophils and induced the antioxidant defences in lymphocytes. PGC-1α and SIRT3 appear to be key effectors of this adaptive response in lymphocytes. Both the neutrophil and lymphocyte response to exercise were slightly weaker in females than males.

Effects of exercise intensity on lymphocyte H2O2 production and antioxidant defences in soccer players

Objective: Physical exercise is capable of enhancing or suppressing the immune response depending on the intensity and duration of exercise. This study investigated how exercise intensity influences the lymphocyte antioxidant response and the induction of cellular oxidative damage. Design: Eighteen voluntary male pre-professional soccer players participated in this study. Sportsmen played a 60 min training match, and were divided into three groups depending on the intensity degree during the match: low, medium and high intensities. Measurements: Malondialdehyde (MDA), vitamins C and E and haem oxygenase-1 (HO-1) gene expression were measured in lymphocytes. Reactive oxygen species (ROS) production was determined in lymphocytes and neutrophils. Results: Lymphocyte MDA levels and H2O2 production were significantly increased in the group which performed the most intense exercise. Neutrophil counts and ROS production increased progressively with the exercise intensity. Vitamin C significantly decreased after exercise in the highest-intensity group in comparison with initial values, whereas vitamin E levels significantly increased in the medium and high-intensity groups. HO-1 gene expression significantly increased in the medium and high-intensity groups. Conclusions: Exercise intensity affects the lymphocyte and neutrophil oxidant/antioxidant balance, but only exercise of high intensity induces lymphocyte oxidative damage.

Scuba diving enhances endogenous antioxidant defenses in lymphocytes and neutrophils.

The aim was to study the effects of a scuba diving session on the lymphocyte antioxidant system, NO synthesis, the capability to produce reactive oxygen species and the antioxidant response in neutrophils. For that purpose seven male divers performed an immersion at a depth of 40 m for 25 min. The same parameters were measured after an hyperbaric oxygen (HBO) treatment at resting conditions in a hyperbaric chamber. Lymphocyte H2O2 production rose after diving and after HBO treatment. Glutathione peroxidase (GPx) and catalase activities increased after diving in lymphocytes, while after HBO exposure only increased GPx activity. Lymphocyte HO-1 mRNA expression increased after diving and after HBO exposure, while iNOS levels and nitrite levels significantly increased after diving. The hyperoxia associated to scuba diving leads to a condition of oxidative stress with increased lymphocyte H2O2 production, HO-1 expression, NO synthesis and antioxidant enzyme adaptations in order to avoid oxidative damage.

Effects of L-citrulline oral supplementation on polymorphonuclear neutrophils oxidative burst and nitric oxide production after exercise

Seventeen volunteer male professional cyclists were randomly assigned to control or supplemented (6 g L-citrulline-malate) groups and participated in a cycling stage. Blood samples were taken in basal conditions, after the race and 3 h post-race. Citrulline supplementation significantly increased plasma concentration of both arginine and citrulline after the stage only in the supplemented group. Polymorphonuclear neutrophils (PMNs) from controls responded to exercise with a progressive decrease in ROS production. Supplemented PMNs significantly increased ROS production after exercise compared to basal values and diminished to values lower than basal at recovery. PMN nitrite concentration was significantly higher after exercise and recovery only in the supplemented group. Markers of oxidative damage—CK, LDH, malondialdehyde—and DNA damage remained unchanged in both groups. In conclusion, oral L-citrulline administration previous to a cycling stage increases plasma arginine availability for NO synthesis and PMNs priming for oxidative burst without oxidative damage.

Body temperature modulates the antioxidant and acute immune responses to exercise

Abstract The aim of this study was to determine the effects of whole body heat in combination with exercise on the oxidative stress and acute phase immune response. Nine male endurance-trained athletes voluntarily performed two running bouts of 45 minutes at 75–80% of VO2max in a climatic chamber in two conditions: cold and hot humid environment. Leukocyte, neutrophil and basophil counts significantly rose after exercise in both environments; it was significantly greater in the hot environment. Lymphocyte and neutrophil antioxidant enzyme activities and carbonyl index significantly increased or decreased after exercise only in the hot environment, respectively. The lymphocytes expression of catalase, Hsp72 and CuZn-superoxide dismutase was increased in the hot environment and Sirt3 in the cold environment, mainly during recovery. In conclusion, the increased core body temperature results in the acute phase immune response associated to intense exercise and in the immune cell adaptations to counteract the oxidative stress situation.

Antioxidant response and oxidative damage induced by a swimming session: Influence of gender

Abstract In this study, we examined oxidative stress after a swimming session, the responses of the antioxidant defences, and the influence of gender on these responses. Fifteen boys and eight girls participated voluntarily in the study. Plasma concentrations of 17-β-estradiol, vitamin E, retinol, carotenes, ascorbate, malondialdehyde, and the carbonyl index were determined. Creatine kinase, gamma glutamyl transpeptidase, and lactate dehydrogenase activities, as well as glucose, urea, urate, cholesterol, and triglycerides, were determined in serum. Plasma concentrations of 17-β-estradiol were higher in girls than in boys. Exercise increased plasma ascorbate both in boys and in girls. Malondialdehyde increased in boys but was maintained in girls after exercise. Creatine kinase values corrected for lean body mass were similar in boys and girls at baseline, but the post-exercise values in boys were higher than in girls. A positive correlation was observed in boys, but not in girls, between plasma malondialdehyde and creatine kinase corrected concentrations. Furthermore, a negative correlation was observed between the increase in circulating neutrophils and in creatine kinase activity in girls but not in boys. In conclusion, a swimming session induced higher muscular and oxidative damage in boys than in girls.

Intense physical activity enhances neutrophil antioxidant enzyme gene expression. Immunocytochemistry evidence for catalase secretion

We studied the effects of intense exercise on the neutrophil antioxidant enzyme activities and gene expression. Blood samples were taken from seven cyclists in basal conditions and 3 h after two competition stages of 165 km. Serum creatine kinase (CK) activity, plasma carbonyl derivatives and uric acid levels increased after exercise. The cycling stage induced neutrophilia and increased myeloperoxidase (MPO) activity and reactive oxygen species (ROS) production. Antioxidant enzyme activities (catalase, glutathione peroxidase and superoxide dismutase) decreased after exercise, although gene expression increased. Immunocytochemistry showed catalase (CAT) enzyme equally distributed between the cytoplasm and organelles before exercise, and after exercise the cytoplasmic CAT levels were reduced and were absent in the compartments. After in vitro stimulation with opsonized zymosan (OZ) the extracellular CAT levels increased. This suggests a CAT secretion in order to avoid neutrophil-induced oxidative damage at a local level or to regulate the function of ROS as extracellular signalling molecules.

Lymphocyte antioxidant response and H2O2 production after a swimming session: Gender differences

This study evaluated the gender differences in response to intense exercise on lymphocyte hydrogen peroxide production, nitric oxide handling and mitochondrial superoxide dismutase (MnSOD) activity and gene expression. Fifteen males and nine females participated voluntarily in the study and performed a swimming session at 75–80% of the maximal capacity. In basal conditions females presented higher lymphocyte MnSOD activity compared to males (p<0.05). Exercise increased MnSOD activity in males (p<0.05) reaching similar values to females. MnSOD gene expression was also increased in males after exercise (p<0.05) but not in females. Nitrite concentration and iNOS gene expression significantly increased only in males after swimming (p<0.01). The exercise decreased UCP-3 gene expression in both genders (p<0.05). Lymphocyte H2O2 production significantly increased in males after exercise in non-stimulated and in PMA-stimulated cells (p<0.01). In conclusion, females seem to be more protected against oxidative stress induced by a swimming session. Hydrogen peroxide is mainly produced in males and this subsequently leads to increases in MnSOD gene expression and activity.

The Double Edge of Reactive Oxygen Species as Damaging and Signaling Molecules in HL60 Cell Culture

Aims: Our aim was to establish the conditions in which reactive oxygen species produce pathological or hormetic effects on HL60 cells. Methods: HL60 cells were treated with either single bouts (1, 10 and 100 µM) or a sustained production (0.1, 1.0 and 10.0 nM/s) of H₂O₂. Results: Exposure to 10 and 100 µM H₂O₂ activated catalase, glutathione peroxidase and glutathione reductase through post-transcriptional mechanisms and induced oxidative modification of proteins. When cells where exposed to sustained H₂O₂ production, a clear dose-response effect was detected in the activity of the antioxidant enzymes catalase, glutathione peroxidase and Mn-SOD, with higher concentrations of H₂O₂ inducing greater enzyme activities. Catalase, HO-1, UCP-3, iNOS and PGC-1α expressions were activated after sustained exposure to 1 and 10 nM H₂O₂/s. Although the antioxidant defenses were activated, oxidative damage appeared in DNA and proteins in cells treated with 1 and 10 nM/s. Conclusions: HL60 cells respond to exposure to sustained levels of H₂O₂ in a dose-response manner to H₂O₂ concentration by activating the expression and activity of the antioxidant machinery, although the activation of the antioxidant defenses is not enough to avoid the appearance of oxidative damage. Of the two designs proposed, continuous exposure seems to be more appropriate to study the antioxidant response of HL60 cells to H₂O₂.