Intracellular accumulation, subcellular distribution and efflux of tilmicosin in swine phagocytes

Abstract

Tilmicosin is a semi-synthetic macrolide antibiotic, currently approved for veterinary use in cattle and swine respiratory disease. As the concentrations of tilmicosin are generally low in swine lung tissue, the interaction of tilmicosin with three types of swine phagocytes (monocyte-macrophages, alveolar macrophages, and neutrophils) was evaluated to provide an understanding of clinical efficacy. After incubation with radiolabelled tilmicosin, uptake was determined and expressed as the ratio of the intracellular (Ci) to the extracellular (Ce) drug concentration (Ci/Ce). Tilmicosin was avidly accumulated by the swine phagocytes (Ci/Ce 48-69 at 4 h incubation) with 51 to 85% localized in the lysosomes. Uptake was dependent on cell viability, temperature and pH, but was not influenced by the metabolic inhibitors, sodium cyanide or potassium fluoride. However, lipopolysaccharide (LPS) exposure increased tilmicosin uptake by the swine phagocytes. In neutrophils, upon removal of extracellular tilmicosin, 60% of the intracellular tilmicosin was effluxed within the first 30 min, but after 4 h of incubation in drug-free medium, 25% remained cell-associated. In contrast, after 4 h of incubation in drug-free medium, 60% and 45% of tilmicosin remained cell-associated, within alveolar macrophages and monocyte-derived macrophages, respectively. Tilmicosin uptake was observed to increase lysosomal enzyme (acid phosphatase, lysozyme and beta-glucuronidase) production. Finally, neutrophils were shown to transport and efflux bioactive tilmicosin in a test system measuring both neutrophil chemotaxis under agarose and a bioassay measuring inhibition of bacterial growth in the presence of antibiotic in agar. These in vitro interactions of tilmicosin with swine phagocytes suggest an integral role in effecting clinical efficacy.

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“…The intracellular uptake method was similar to that described by (Scorneaux and Shryock, 1998). N1E-115 mice neuroblastoma cells (500,000 per plate) were cultivated in DMEM supplemented with 10% FCS and 100 units/ml penicillin/0.1 mg/ ml streptomycin at 37 C in a humid atmosphere of 95% air and 5% CO 2 in T225 bottles.…”

Tilmicosin and tylosin have anti-inflammatory properties via modulation of COX-2 and iNOS gene expression and production of cytokines in LPS-induced macrophages and monocytes

“…Tilmicosin is a semi-synthetic macrolide antibiotic, derived from tylosin, proven to be effective in treating cattle [8,9], sheep [10], swine [11] and inflammatory airway diseases and mastitis in ruminant animals [12]. The efficacy of tilmicosin is attributed to its pharmacodynamic character [13,14] and low inhibitory concentrations [15,16]. There is some evidence to suggest that tilmicosin may exert its anti-inflammatory effect through actions other than its antimicrobial properties, for example immunomodulatory actions on neutrophil apoptosis [17][18][19] and modulated production of inflammatory factors including prostaglandin E 2 (PGE 2 ) in macrophages [20].…”

Kinetics and intrapulmonary disposition of tilmicosin after single and repeated oral bolus administrations to rabbits

“…Clinical studies carried out in swine and calves by TIM oral administration have confirmed its good efficacy against pathogen pulmonary bacteria and its ability to modulate the inflammatory process by reducing inflammatory mediators in lung tissue (Scorneaux and Shryock 1998;Chin et al 1998). TIM can easily accumulate in pulmonary alveolar macrophages (PAM) because of the ion-trapping process (Honeybourne 1997).…”