Study of the immunomodulatory properties of gamithromycin and dexamethasone in a lipopolysaccharide inflammation model in calves.

The aim of this study was to define the in vivo immunomodulatory properties of the macrolide antibiotic gamithromycin in calves, with respect to the acute phase response. Additionally, the corticosteroid dexamethasone was included as a positive control immunomodulatory drug. Both drugs, as well as their combination,were studied in a previously developed inflammation model,which was initiated by an intravenous lipopolysaccharide (LPS) challenge (0.5 µg/kg body weight). Twenty-four 4-week-old male Holstein Friesian calves were randomized into four groups: no pharmacological treatment (n = 6) or a pharmacological treatment with gamithromycin (n= 6), dexamethasone (n= 6) or their combination (n= 6) 1 h prior to LPS administration. Blood collection and clinical scoring were performed at regular time points until 72 h post LPS challenge. Plasma concentrations of selected cytokines (tumour necrosis factor-α (TNF-α) and interleukin 6 (IL-6)) and acute phase proteins (serum amyloid A and haptoglobin) were subsequently determined. Gamithromycin did not have any beneficial effect on the LPS-induced clinical signs (dyspnea, fever, anorexia and depression), nor on the studied inflammatory mediators. In the dexamethasone and combination groups, the occurrence of dyspnea and fever was not prominently influenced, although the calves recovered significantly faster from the challenge. Moreover, dexamethasone significantly inhibited the levels of TNF-α and IL-6, suggesting a key role for these cytokines in sickness behaviour. In conclusion, unlike dexamethasone, gamithromycin did not directly reduce cytokine release in an LPS inflammation model in calves.

Multiplex analysis of pro-inflammatory cytokines in serum of Actinobacillus pleuropneumoniae-infected pigs.

Porcine pleuropneumonia is a severe respiratory disease caused by Actinobacillus (A.) pleuropneumoniae. The aim of the present study was to analyze serum samples of A. pleuropneumoniae-infected pigs for TNF-α, IL-1ß and IL-6 using a cytometric bead array (CBA) 3-plex assay and additionally for IL-6 using ELISA. The CBA 3-plex assay was successfully validated for use in serum. The limits of detection varied between 0.012 and 0.333 ng/mL, and the inter- and inter-assay coefficients of variation were <5% and <10%, respectively. Increased levels were observed for all 3 cytokines following experimental infection with A. pleuropneumoniae. Mean peak concentrations of TNF-α and IL-6 were recorded at 12h and at 10h p.i., respectively. For IL-6, similar concentration-time profiles were observed with CBA and ELISA. It is proposed that this immuno-assay can be applied for the screening of immunomodulatory properties of drugs and vaccine adjuvants in infection, inflammation and vaccination.

In vivo porcine lipopolysaccharide inflammation models to study immunomodulation of drugs.

Lipopolysaccharide (LPS), a structural part of the outer membrane of Gram-negative bacteria, is one of the most effective stimulators of the immune system and has been widely applied in pigs as an experimental model for bacterial infection. For this purpose, a variety of Escherichia coli serotypes, LPS doses, routes and duration of administration have been used. LPS administration induces the acute phase response (APR) and is associated with dramatic hemodynamic, clinical and behavioral changes in pigs. Pro-inflammatory cytokines, including tumor necrosis factor α (TNF-α), interleukin (IL)-1 and IL-6 are involved in the induction of the eicosanoid pathway and the hepatic production of acute phase proteins, including C-reactive protein (CRP), haptoglobin (Hp) and pig major acute phase protein (pig-MAP). Prostaglandin E2 (PGE2) and thromboxane A2 (TXA2) play a major role in the development of fever and pulmonary hypertension in LPS-challenged pigs, respectively. The LPS-induced APR can be modulated by drugs. Steroidal and nonsteroidal anti-inflammatory drugs ((N)SAIDs) possess anti-inflammatory, antipyretic and analgesic properties through (non)-selective central and peripheral cyclooxygenase (COX) inhibition. Antimicrobial drugs, especially macrolide antibiotics, which are commonly used in veterinary medicine for the treatment of bacterial respiratory diseases, have been recurrently reported to exert clinically important immunomodulatory effects in human and murine research. To investigate the influence of these drugs on the clinical response, production of pro-inflammatory cytokines, acute phase proteins (APP) and the course of the febrile response in pigs, in vivo LPS inflammation models can be applied. Yet, to date, in vivo research on the immunomodulatory properties of antimicrobial drugs in these models in pigs is largely lacking. This review provides acritical overview of the use of in vivo porcine E. coli LPS inflammation models for the study of the APR, as well as the potential immunomodulatory properties of anti-inflammatory and antimicrobial drugs in pigs.

Enantioselective pharmacokinetics of ketoprofen in calves after intramuscular administration of a racemic mixture.

The pharmacokinetic properties of ketoprofen were determined in 4-week-old calves after intramuscular (i.m.) injection of a racemic mixture at a dose of 3 mg/kg body weight. Due to possible enantioselective disposition kinetics and chiral inversion, the plasma concentrations of the R(-) and S(+) enantiomer were quantified separately, using a stereospecific HPLC-UV assay. A distinct predominance of the S(+) enantiomer was observed, as well as significantly different pharmacokinetic parameters between R(-) and S(+) ketoprofen. More in specific, a greater value for the mean area under the plasma concentration-time curve (AUC(0→∞)) (46.92 ± 7.75 and 11.13 ± 2.18 µg·h/mL for the S(+) and R(-) enantiomer, respectively), a lower apparent clearance (Cl/F) (32.8 ± 5.7 and 139.0 ± 25.1 mL/h·kg for the S(+) and R(-) enantiomer, respectively) and a lower apparent volume of distribution (V(d)/F) (139 ± 14.7 and 496 ± 139.4 mL/kg for the S(+) and R(-) enantiomer, respectively) were calculated for the S(+) enantiomer, indicating enantioselective pharmacokinetics for ketoprofen in calves following i.m. administration.

Pharmacokinetics of gamithromycin after intravenous and subcutaneous administration in pigs.

The aim of this study was to investigate the pharmacokinetic properties of gamithromycin in pigs after an intravenous (i.v.) or subcutaneous (s.c.) bolus injection of 6 mg/kg body weight. The plasma concentrations of gamithromycin were determined using a validated high-performance liquid chromatography-tandem mass spectrometry method, and the pharmacokinetics were noncompartmentally analysed. Following i.v. administration, the mean area under the plasma concentration-time curve extrapolated to infinity (AUCinf) and the mean elimination half-life (t1/2λz) were 3.67 ± 0.75 µg.h/mL and 16.03 h, respectively. The volume of distribution at steady state (Vss) and the plasma clearance were 31.03 ± 6.68 L/kg and 1.69 ± 0.33 L/h.kg, respectively. The mean residence time (MRTinf) was 18.84 ± 4.94 h. Gamithromycin administered subcutaneously to pigs demonstrated a rapid and complete absorption, with a mean maximal plasma concentration (Cmax) of 0.41 ± 0.090 µg/ml at 0.63 ± 0.21 h and a high absolute bioavailability of 118%. None of the reported pharmacokinetic variables significantly differed between both administration routes.

Pharmacokinetics of gamithromycin after intravenous and subcutaneous administration in broiler chickens.

Gamithromycin is a new macrolide antibiotic that is only registered for use in cattle to treat respiratory disorders such as bovine respiratory disease. The aim of this study was to determine the pharmacokinetics of gamithromycin in broiler chickens. Gamithromycin (6 mg/kg of BW) was injected intravenously (IV) or subcutaneously (SC) to six 4-wk-old chickens in a parallel study design, and blood was collected at different time points postadministration. Quantification of gamithromycin in plasma was performed using an in-house validated liquid chromatography-tandem mass spectrometry method and the pharmacokinetics analyzed according to a 2-compartmental model. Following IV administration, the mean area under the plasma concentration-time curve (AUC0→∞), and α and ß half-life of elimination (t1/2el α and t1/2el ß) were 3,998 h•ng/mL, 0.90 h, and 14.12 h, respectively. Similar values were obtained after a SC bolus injection, i.e., 4,095 h•ng/mL, 0.34 h, and 11.63 h, for AUC0→∞, t1/2el α, and t1/2el ß, respectively. The mean maximum plasma concentration (889.46 ng/mL) appeared at 0.13 h. Gamithromycin showed a large volume of distribution after IV as well as SC administration, 27.08 and 20.89 L/kg, respectively, and a total body clearance of 1.61 and 1.77 L/h•kg, respectively. The absolute bioavailability was 102.4%, showing that there is a complete absorption of gamithromycin after a SC bolus injection of 6 mg/kg of BW.