Non-genomic action of beclomethasone dipropionate on bronchoconstriction caused by leukotriene C4 in precision cut lung slices in the horse

Abstract

Background: Glucocorticoids have been proven to be effective in the therapy of recurrent airway obstruction (RAO) in horses via systemic as well as local (inhalative) administration. Elective analysis of the effects of this drug on bronchoconstriction in viable lung tissue offers an insight into the mechanism of action of the inflammatory cascade occurring during RAO which is still unclear. The mechanism of action of steroids in treatment of RAO is thought to be induced through classical genomic pathways. We aimed at electively studying the effects of the glucocorticoid beclomethasone dipropionate on equine precision-cut lung slices (PCLS).PCLS were used to analyze ex-vivo effects of beclomethasone on inhibiting bronchoconstriction in the horse. The inhibiting effect was measured through instillation of a known mediator of inflammation and bronchoconstriction, leukotriene C4. For this, the accessory lobes of 13 horses subjected to euthanasia for reasons unrelated to the respiratory apparatus were used to obtain viable lung slices.

Results: After 30 minutes of PCLS incubation, beclomethasone showed to significantly inhibit the contraction of the bronchioles after instillation with leukotriene C4. The EC50 values of the two contraction curves (LTC4 with and without BDP) differed significantly from each other (p = 0.002). The possibility of a non-genomic rapid mechanism of action seems likely since transcriptional activities require a longer lag period.

Conclusions: In human neuroendocrinology, high levels of glucocorticoids have been proven to function via a non-genomic mechanism of membrane receptors. The concentration of beclomethasone used on the lung slices in our study can be considered as high. This allows speculation about similar rapid non-genomic mechanisms of high-dosage inhaled glucocorticoids in the lower airways of horses. However, further assessment on a molecular basis is needed to confirm this.

Figure 1

Comparison between BDP-treated and untreated PCLS, and their reaction to instillation of LTC₄(10^-8 mol/l). Top row: untreated slices (slice A and B). Bottom row: after instillation with LTC₄, with slice A previously treated with BDP at 10^-5 mol/l, slice B untreated. Magnification 100x (Zeiss® Axiostar Microscope Universal Digicam Adapter).

Figure 2

Calculated EC₅₀values for each horse before and after treatment with 10^-5 mol/l beclomethasone dipropionate. Means of the calculated EC₅₀ values of the two contraction series (LTC₄ with and without BDP) differed significantly from each other (p = 0.002). The number indicates the negative power of 10 (mol/l) as the expression of the LTC₄ concentration needed to reach 50% of lumen reduction.

Figure 3

Graphical representation of inhibitory effect of beclomethasone dipropionate on LTC₄-induced bronchoconstriction. Indication of the bronchial lumen (given in % of initial value) in dependency of LTC₄ concentrations (given in 10^-x mol/l). Data shown are means ± SD of 13 horses.