The effects of
tiotropium bromide on ERK 1/2, SMAD 2/3 and NFκB signaling in bronchial submucosal gland (SMG) cells of sheep after
smoke inhalation and
burn injury (S + B) were studied. We hypothesized that
tiotropium would modify intracellular signaling processes within SMG cells after injury. Bronchial tissues were obtained from uninjured (
sham, n = 6), S + B injured sheep 48 h after injury (n = 6), and injured sheep nebulized with
tiotropium (n = 6). The percentage (mean ± SD) of cells showing nuclear localization of phosphorylated ERK 1/2, pSMAD 2/3, and NFκB (p65) was determined by immunohistochemistry. Nuclear pERK 1/2 staining was increased in injured animals as compared to
sham, (66 ± 20 versus 14 ± 9), p = 0.0022, as was nuclear pSMAD, 84 ± 10 versus 20 ± 10, p = 0.0022. There was a significant decrease in pERK 1/2 labeling in the
tiotropium group compared to the injured group (31 ± 20 versus 66 ± 20, p = 0.013), and also a decrease in pSMAD labeling, 62 ± 17 versus 84 ± 10, p = 0.04. A significant increase for NFκB (p65) was noted in injured animals as compared to
sham (73 ± 16 versus 7 ± 6, p = 0.0022).
Tiotropium-treated animals showed decreased p65 labeling as compared to injured (35 ± 17 versus 74 ± 16, p = 0.02). The decrease in nuclear expression of pERK, pSMAD and NFκB molecules in SMG cells with
tiotropium treatment is suggestive that their activation after injury is mediated in part through
muscarinic receptors.