Related Experiment Videos
Intracellular accumulation, subcellular distribution and efflux of tilmicosin in swine phagocytes
1Animal science Research, Elanco Animal Health, Greenfield, IN 46140, USA. Scorneaux_Bernard@Lilly.com
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.
Insights
Tilmicosin, an antibiotic for swine, is effectively accumulated by swine phagocytes, including neutrophils and macrophages. This uptake and subsequent release suggest a key role in the drug
Area of Science:
- Veterinary Pharmacology
- Immunology
- Antimicrobial Agents
Background:
- Tilmicosin is a semi-synthetic macrolide antibiotic used in veterinary medicine for cattle and swine respiratory diseases.
- Low concentrations of tilmicosin in swine lung tissue necessitate understanding its interaction with phagocytic cells for clinical efficacy.
Purpose of the Study:
- To evaluate the interaction of tilmicosin with swine phagocytes (monocyte-macrophages, alveolar macrophages, neutrophils).
- To elucidate the role of these interactions in the clinical efficacy of tilmicosin in swine.
Main Methods:
- Incubation of swine phagocytes with radiolabelled tilmicosin.
- Measurement of tilmicosin uptake (intracellular to extracellular concentration ratio, Ci/Ce) and localization (lysosomes).
- Assessment of uptake dependency on cell viability, temperature, pH, metabolic inhibitors, and lipopolysaccharide (LPS) exposure.
- Evaluation of tilmicosin efflux from neutrophils, alveolar macrophages, and monocyte-derived macrophages.
- Measurement of lysosomal enzyme production and assessment of bioactive tilmicosin transport and efflux by neutrophils.
Main Results:
- Swine phagocytes avidly accumulated tilmicosin (Ci/Ce 48-69 at 4 h), with 51-85% localized in lysosomes.
- Uptake was dependent on cell viability, temperature, and pH, but enhanced by LPS exposure.
- Neutrophils exhibited significant efflux of tilmicosin, while macrophages retained higher intracellular concentrations.
- Tilmicosin uptake increased lysosomal enzyme production, and neutrophils transported and effluxed bioactive tilmicosin.
Conclusions:
- Tilmicosin is actively taken up and retained by swine phagocytes, with differential efflux rates between neutrophils and macrophages.
- The drug's interaction with phagocytes, including lysosomal enzyme induction and bioactive efflux, suggests a significant role in its clinical efficacy against swine respiratory diseases.