Vitamin E, γ-tocopherol, reduces airway neutrophil recruitment after inhaled endotoxin challenge in rats and in healthy volunteers

Abstract

Epidemiologic studies suggest that dietary vitamin E is an important candidate intervention for asthma. Our group has shown that daily consumption of vitamin E (γ-tocopherol, γT) has anti-inflammatory actions in both rodent and human phase I studies. The objective of this study was to test whether γT supplementation could mitigate a model of neutrophilic airway inflammation in rats and in healthy human volunteers. F344/N rats were randomized to oral gavage with γT versus placebo, followed by intranasal LPS (20μg) challenge. Bronchoalveolar lavage fluid and lung histology were used to assess airway neutrophil recruitment. In a phase IIa clinical study, 13 nonasthmatic subjects completed a double-blinded, placebo-controlled crossover study in which they consumed either a γT-enriched capsule or a sunflower oil placebo capsule. After 7 days of daily supplementation, they underwent an inhaled LPS challenge. Induced sputum was assessed for neutrophils 6 h after inhaled LPS. The effect of γT compared to placebo on airway neutrophils post-LPS was compared using a repeated-measures analysis of variance. In rats, oral γT supplementation significantly reduced tissue infiltration (p<0.05) and accumulation of airway neutrophils (p<0.05) that are elicited by intranasal LPS challenge compared to control rats. In human volunteers, γT treatment significantly decreased induced sputum neutrophils (p=0.03) compared to placebo. Oral supplementation with γT reduced airway neutrophil recruitment in both rat and human models of inhaled LPS challenge. These results suggest that γT is a potential therapeutic candidate for prevention or treatment of neutrophilic airway inflammation in diseased populations.

Figure 1. Pre-clinical studies using rats

A) Time Course of treatment with γT (black triangles) and intranasal LPS (up-arrows). Treatment related effects of LPS and γT on B) BALF and C) tissue PMNs collected 24h after the last LPS challenge.

Figure 2. Lung histology in rats

Photomicrographs of lung tissues stained with hematoxylin &eosin from rats after airway treatment with A) saline, B) endotoxin or C) endotoxin and γT. ap=alveolar parenchyma; bv = blood vessel; sa = small airway; arrows indicate inflammatory cell infiltrates. Magnification bar = 20mm.

Figure 3. Phase IIa study design in healthy human volunteers

Randomized, placebo controlled crossover study of a γT-enriched supplement or sunflower oil placebo in 13 healthy volunteers. Subjects were challenged with 20,000 endotoxin units of Clinical Center Reference Endotoxin (CCRE), followed by sputum induction 6 hours after the challenge.

Figure 4. Serum levels of Tocopherols and γ-CEHC during the placebo and active study periods

A). serum γT concentrations B). serum γ-CEHC concentrations C). serum 5-nitro- γT concentrations.

Figure 5. Blood and induced sputum leukocytes

Blood A). Total white blood cell (WBC) count B). Absolute neutrophil count (ANC). Induced sputum C). %neutrophils (%PMNS) and D). %eosinophils. Values were calculated by subtracting the baseline (pre-LPS) %PMN or %eosinophil value to the post-LPS value to assess the deltas. Repeated measures ANOVA was used to calculate the treatment effect.