Pharmacokinetics and ex vivo anti-inflammatory effects of oral misoprostol in horses

Abstract

Background: Misoprostol is an E prostanoid (EP) 2, 3 and 4 receptor agonist that is anecdotally used to treat and prevent NSAID-induced GI injury in horses. Misoprostol elicits anti-inflammatory effects in vivo in men and rodents, and inhibits TNFα production in equine leucocytes in vitro.

Objective: Define the pharmacokinetic parameters of oral misoprostol in horses, and determine the inhibitory effect of oral misoprostol administration on equine leucocyte TNFα production in an ex vivo inflammation model.

Study design: Pharmacokinetic study, ex vivo experimental study.

Methods: Six healthy adult horses of mixed breeds were used. In phase one, horses were given 5 μg/kg misoprostol orally, and blood was collected at predetermined times for determination of misoprostol free acid (MFA) by UHPLC-MS/MS. Pharmacokinetic parameters were calculated. In phase two, horses were dosed as in phase one, and blood was collected at T0, 0.5, 1 and 4 h following misoprostol administration for leucocyte isolation. Leucocytes were stimulated with 100 ng/mL LPS, and TNFα mRNA concentrations were determined via quantitative real-time PCR.

Results: About 5 μg/kg oral misoprostol produced a rapid time to maximum concentration (T_max ) of 23.4 ± 2.4 min, with a maximum concentration (C_max ) of 0.29 ± 0.07 ng/mL and area under the curve (AUC_0-∞ ) of 0.4 ± 0.12 h ng/mL. LPS stimulation of equine leucocytes ex vivo significantly increased TNFα mRNA concentrations, and there was no significant effect of misoprostol even at the T_max .

Main limitations: Only a single dose was used, and sample size was small.

Conclusions: Misoprostol is rapidly absorbed following oral administration in horses, and a single 5 μg/kg dose had no significant inhibitory effect on ex vivo LPS-stimulated TNFα mRNA production in leucocytes. Further studies analysing different dosing strategies, including repeat administration or combination with other anti-inflammatory drugs, are warranted.

Keywords: E prostanoid receptor agonist; horse; inflammation; leucocyte; pharmacokinetics; tumour necrosis factor-α.

Figure 1

Mean plasma concentration of misoprostol free acid (ng/mL) vs. time (hours) after oral administration of misoprostol at a dose of 5 μg/kg. Data are represented as mean ± s.d. and include five different horses.

Figure 2

Effect of oral misoprostol administration on LPS‐stimulated equine leucocyte TNFα mRNA concentrations ex vivo. Following collection of whole blood (see methods section), one leucocyte‐rich plasma (LRP) sample per horse was immediately processed as a baseline measurement of TNFα mRNA concentrations. Remaining samples of LRP was incubated with either LPS (100 ng/mL) or vehicle (PBS) for 2 h. mRNA was isolated for real‐time PCR. a) LPS significantly increased TNFα mRNA concentrations at each time point evaluated. Data are presented as mean fold change TNFα mRNA ± s.e.m. vs. unstimulated baseline cells (not shown, equal to 1) and represent six horses. *P<0.05 vs. time‐matched vehicle treated cells (white bar) via paired t test. b) Misoprostol did not exert a statistically significant effect on LPS‐stimulated TNFα mRNA concentrations in equine leucocyte‐rich plasma. Data are presented as mean fold change TNFα mRNA ± s.e.m. vs. time‐matched vehicle‐treated unstimulated cells (not shown, equal to 1) and represent six different horses. c) Inter‐horse variability of the effect of misoprostol on LPS‐stimulated TNFα mRNA concentrations. Data are presented as in b).