Immune cell response to strenuous resistive breathing: comparison with whole body exercise and the effects of antioxidants.

Background/hypothesis: Whole body exercise (WBE) changes lymphocyte subset percentages in peripheral blood. Resistive breathing, a hallmark of diseases of airway obstruction, is a form of exercise for the inspiratory muscles. Strenuous muscle contractions induce oxidative stress that may mediate immune alterations following exercise. We hypothesized that inspiratory resistive breathing (IRB) alters peripheral blood lymphocyte subsets and that oxidative stress mediates lymphocyte subpopulation alterations following both WBE and IRB. Patients and methods: Six healthy nonathletes performed two WBE and two IRB sessions for 45 minutes at 70% of VO2 maximum and 70% of maximum inspiratory pressure (Pimax), respectively, before and after the administration of antioxidants (vitamins E, A, and C for 75 days, allopurinol for 30 days, and N-acetylcysteine for 3 days). Blood was drawn at baseline, at the end of each session, and 2 hours into recovery. Lymphocyte subsets were determined by flow cytometry. Results: Before antioxidant supplementation at both WBE end and IRB end, the natural killer cell percentage increased, the T helper cell (CD3+ CD4+) percentage was reduced, and the CD4/CD8 ratio was depressed, a response which was abolished by antioxidants only after IRB. Furthermore, at IRB end, antioxidants promoted CD8+ CD38+ and blunted cytotoxic T-cell percentage increase. CD8+ CD45RA+ cell percentage changes were blunted after antioxidant supplementation in both WBE and IRB. Conclusion: We conclude that IRB produces (as WBE) changes in peripheral blood lymphocyte subsets and that oxidative stress is a major stimulus predominantly for IRB-induced lymphocyte subset alterations.

Ex Vivo Induction of Multiple Myeloma-specific Immune Responses by Monocyte-derived Dendritic Cells Following Stimulation by Whole-tumor Antigen of Autologous Myeloma Cells.

The introduction of novel agents has significantly expanded treatment options for multiple myeloma (MM), albeit long-term disease control cannot be achieved in the majority of patients. Vaccination with MM antigen-loaded dendritic cells (DCs) represents an alternative strategy that is currently being explored. The aim of this study was to assess the immunogenic potential of ex vivo-generated monocyte-derived DCs (moDCs), following stimulation with the whole-antigen array of autologous myeloma cells (AMC). MoDCs were loaded with antigens of myeloma cells by 2 different methods: phagocytosis of apoptotic bodies from γ-irradiated AMC, or transfection with AMC total RNA by square-wave electroporation. Twenty patients with MM were enrolled in the study. Following stimulation and maturation, moDCs were tested for their capacity to induce T-helper 1 and cytotoxic T lymphocyte responses in vitro. Both strategies were effective in the induction of myeloma-specific cytotoxic T lymphocyte and T-helper 1 cells, as demonstrated by cytotoxicity and ELISpot assays. On the whole, T-cell responses were observed in 18 cases by either method of DC pulsing. We conclude that both whole-tumor antigen approaches are efficient in priming autologous antimyeloma T-cell responses and warrant further study aiming at the development of individualized DC vaccines for MM patients.

Antioxidant supplementation alters cytokine production from monocytes.

We studied in 10 healthy subjects the effect of chronic enteral supplementation of antioxidants (vitamins E, C, A, allopurinol, and N-acetylcysteine) on cytokine production by monocytes at rest, end exercise (60-min cycling at 60% of maximum oxygen consumption), and 60 min post-exercise (recovery). The percentage and the mean fluorescent intensity (MFI) of both unstimulated and lipopolysaccharide (LPS)-stimulated tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, and IL-6-producing monocytes were detected using flow cytometry. Antioxidants decreased the percentage of unstimulated IL-6-producing monocytes following exercise, while their MFI increased at rest. The percentage of LPS-stimulated monocytes increased after exercise and they produced more IL-6 both at rest and following exercise. The percentage of unstimulated and LPS-stimulated IL-1beta-producing monocytes was not affected by antioxidants. The MFI of IL-1beta-produced unstimulated monocytes was increased after antioxidants both at rest and following exercise. After antioxidants, LPS-stimulated monocytes produced more IL-1beta following exercise. Antioxidants decreased the percentage of TNF-alpha spontaneously-produced monocytes following exercise, which produced more TNF-alpha at recovery. Antioxidants did not affect the percentage of LPS-stimulated monocytes producing TNF-alpha, while LPS-stimulated production of TNF-alpha increased both at rest and following exercise. Antioxidants differentially affect TNF-alpha, IL-1beta, and IL-6 production by monocytes, with a general tendency of augmenting cytokine production.

Antioxidants increase the ventilatory response to hyperoxic hypercapnia.

RATIONALE: The mechanisms by which chemoreceptors process carbon dioxide stimuli are poorly understood. Recent in vitro studies suggest a role of reactive oxygen species in central carbon dioxide chemoreception. OBJECTIVES: We tested the hypothesis that antioxidant treatment modulates the ventilatory response to carbon dioxide in healthy humans, either during unloaded breathing or after strenuous resistive breathing. METHODS: In the first experiment of this randomized, double-blind, placebo-controlled study, 14 healthy males completed hyperoxic carbon dioxide rebreathing, received either antioxidants (vitamins E, A, and C for 2 mo, allopurinol for 15 d, and N-acetylcysteine for 3 d) (n = 7) or placebo (n = 7), and repeated rebreathing 3 mo later. In the second experiment, 18 healthy males completed a series of rebreathing tests before and after strenuous resistive breathing. Subjects repeated the same protocol 3 mo later, after they had received antioxidants (n = 9) or placebo (n = 9). MAIN RESULTS: After antioxidants, the sensitivity of the ventilatory (minute ventilation) response to carbon dioxide increased (mean [+/- SEM], 3.2 +/- 0.5 vs. 1.7 +/- 0.4 L/min/mm Hg; p < 0.001). Antioxidants also increased the sensitivity to carbon dioxide before and at 5, 30, and 120 min after resistive breathing (p = 0.01). This effect was entirely due to increased tidal volume. Antioxidants did not influence the breathing pattern during resting breathing or the rapid shallow breathing response to carbon dioxide at 5 min after resistive breathing. CONCLUSIONS: Antioxidants, by augmenting the tidal volume, increase the sensitivity of the ventilatory response to carbon dioxide, either during unloaded breathing or after resistive breathing.

Antioxidants attenuate the plasma cytokine response to exercise in humans.

Exercise increases plasma TNF-alpha, IL-1beta, and IL-6, yet the stimuli and sources of TNF-alpha and IL-1beta remain largely unknown. We tested the role of oxidative stress and the potential contribution of monocytes in this cytokine (especially IL-1beta) response in previously untrained individuals. Six healthy nonathletes performed two 45-min bicycle exercise sessions at 70% of Vo(2 max) before and after a combination of antioxidants (vitamins E, A, and C for 60 days; allopurinol for 15 days; and N-acetylcysteine for 3 days). Blood was drawn at baseline, end-exercise, and 30 and 120 min postexercise. Plasma cytokines were determined by ELISA and monocyte intracellular cytokine level by flow cytometry. Before antioxidants, TNF-alpha increased by 60%, IL-1beta by threefold, and IL-6 by sixfold secondary to exercise (P < 0.05). After antioxidants, plasma IL-1beta became undetectable, the TNF-alpha response to exercise was abolished, and the IL-6 response was significantly blunted (P < 0.05). Exercise did not increase the percentage of monocytes producing the cytokines or their mean fluorescence intensity. We conclude that in untrained humans oxidative stress is a major stimulus for exercise-induced cytokine production and that monocytes play no role in this process.

Strenuous resistive breathing induces plasma cytokines: role of antioxidants and monocytes.

Inspiratory resistive breathing increases plasma cytokines, yet the stimulus (or stimuli) and source(s) remain unknown. We tested the role of reactive oxygen species as stimuli and of monocytes as sources of resistive breathing-induced cytokines. Six healthy subjects performed two resistive breathing sessions at 75% of maximum inspiratory pressure before and after a combination of antioxidants (vitamin E 200 mg, vitamin A 50,000 IU, and vitamin C 1,000 mg per day for 60 days, allopurinol 600 mg/day for 15 days, and N-acetylcysteine 2 g/day for 3 days before the second session). Blood was drawn before, at the end, and at 30 and 120 minutes after resistive breathing. Before antioxidants, plasma cytokine levels (determined by enzyme-linked immunosorbent assay) increased secondary to resistive breathing (tumor necrosis factor-alpha and interleukin [IL]-6 by twofold and IL-1beta by threefold). After antioxidants, plasma IL-1beta became undetectable. The tumor necrosis factor-alpha response to resistive breathing was abolished, and the IL-6 response was significantly blunted. Intracellular cytokine detection (by flow cytometry) showed no change in either the percentage of monocytes producing the cytokines or their mean fluorescence intensity both before and after antioxidants. We conclude that oxidative stress is a major stimulus for the resistive breathing-induced cytokine production and that monocytes play no role in this process.