Simvastatin inhibits smoke-induced airway epithelial injury: implications for COPD therapy

Abstract

Chronic obstructive pulmonary disease (COPD) is the third leading cause of death. The statin drugs may have therapeutic potential in respiratory diseases such as COPD, but whether they prevent bronchial epithelial injury is unknown. We hypothesised that simvastatin attenuates acute tobacco smoke-induced neutrophilic lung inflammation and airway epithelial injury. Spontaneously hypertensive rats were given simvastatin (20 mg·kg(-1) i.p.) daily for either 7 days prior to tobacco smoke exposure and during 3 days of smoke exposure, or only during tobacco smoke exposure. Pretreatment with simvastatin prior to and continued throughout smoke exposure reduced the total influx of leukocytes, neutrophils and macrophages into the lung and airways. Simvastatin attenuated tobacco smoke-induced cellular infiltration into lung parenchymal and airway subepithelial and interstitial spaces. 1 week of simvastatin pretreatment almost completely prevented smoke-induced denudation of the airway epithelial layer, while simvastatin given only concurrently with the smoke exposure had no effect. Simvastatin may be a novel adjunctive therapy for smoke-induced lung diseases, such as COPD. Given the need for statin pretreatment there may be a critical process of conditioning that is necessary for statins' anti-inflammatory effects. Future work is needed to elucidate the mechanisms of this statin protective effect.

FIGURE 1

Pre-treatment with simvastatin 1 week prior to tobacco smoke exposure, and continued throughout the exposure time-frame, reduced tobacco smoke-induced leukocyte recruitment to the lung, while treatment only on the smoke exposure days had no effect. a) Total leukocytes recovered in the bronchoalveolar lavage fluid (BALF). b) Total macrophages recovered in the BALF. c) Total neutrophils recovered in the BALF. d) Total lymphocytes recovered in the BALF. All groups were analysed by one-way ANOVA with Bonferroni corrections for multiple comparisons.

FIGURE 2

Epithelial and inflammatory changes to the bronchial wall of spontaneous hypertensive rats exposed to filtered air or tobacco smoke and treated with simvastatin. Exposure to only tobacco smoke was associated with significant airway epithelial injury and sloughing. Pre-treatment with simvastatin provided almost complete epithelial protection to tobacco smoke if given 1 week prior to and during exposure to tobacco smoke, but not if treatment only began concurrently with exposure to tobacco smoke. Haematoxilin and eosin staining of bronchial airways at interpulmonary generation 2 in rats exposed to: a, b) filtered air; c, d) tobacco smoke; e, f) smoke plus simvastatin with no pre-treatment; and g, h) smoke plus simvastatin with 1 week pre-treatment with simvastatin. i) Quantitative analysis of the percentage of airway epithelial sloughing, data were analysed using the Wilcoxon rank sum test. a, c, e and g) scale bars = 25 µm. b, d, f and h) scale bars = 10 µm.

FIGURE 3

Treatment with simvastatin for 1 week prior to smoke exposure and continued during the smoke-exposure period reduced lung compliance and increased lung elastance, while simvastatin treatment only on exposure days had no effect. a) Resistance, b) compliance, c) elastance. All groups were analysed by one-way ANOVA with Bonferroni corrections for multiple comparisons.

FIGURE 4

Treatment with simvastatin for 1 week prior to smoke exposure and continued during the exposure did not change localisation of Ras and Rho between the cytosolic (C) and membrane (M) fractions. a) Western blots of active Ras and c) membrane to cytosol ratio of Western blot densitometry. b) Western blots of active Rho and d) membrane to cytosol ratio of Western blot densitometry. All groups were analysed by one-way ANOVA.

FIGURE 5

Treatment with simvastatin for 1 week prior to smoke exposure and continued during the exposure had no effect on total cholesterol in rat whole lung homogenate. Total cholesterol in whole lung homogenate lysates was measured and normalised to total protein, as described in the Materials and Methods. All groups were analysed by one-way ANOVA.

FIGURE 6

Haem oxygenase (HO)-1 protein expression was measured by Western blot as a marker for oxidative stress. a) Western blot of HO-1 and α-actinin (as a control) from 100 µg of total proteins from whole lung homogenates. b) Densitometric analysis of HO-1 expression normalised to α-actinin control. Data are presented as mean±SEM. p-value determined by Wilcoxon Rank sum test.