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Characterization of minocycline transport by human neutrophils
1Section of Periodontology, College of Dentistry, The Ohio State University Health Sciences Center, Columbus, OH 43210, USA. walters2@osu.edu
Background:
Tetracyclines are used in periodontal therapy as antimicrobial agents and as inhibitors of matrix metalloproteinases. Neutrophils appear to accumulate minocycline and other tetracyclines through a mechanism that has not been fully characterized.
Methods:
The transport of minocycline and other tetracyclines by isolated human neutrophils was characterized by measuring the increase in cell-associated fluorescence.
Results:
Quiescent neutrophils took up minocycline through a saturable, concentrative, sodium-dependent mechanism with a Michaelis constant (K(m)) of 153 micro g/ml (501 microM) and a maximal velocity of 240 ng/minute/10(6) cells. The efficiency of minocycline transport was not influenced significantly by a two-unit variation in extracellular pH and was not enhanced upon cell activation with phorbol myristate acetate. Neutrophil incubation in medium containing 10 micro g/ml minocycline, doxycycline, or tetracycline yielded steady-state intracellular/extracellular concentration ratios of approximately 64.0, 7.5, or 1.8, respectively. The dilution of extracellular minocycline or doxycycline triggered efflux from cells loaded with these antibiotics. Minocycline transport was competitively inhibited by the organic cations carnitine, diphenhydramine, and verapamil, but penicillin and other organic anions failed to produce inhibition.
Conclusion:
Transport of tetracyclines by neutrophils could potentially enhance the effectiveness of these agents in periodontal therapy by enhancing or sustaining their therapeutic levels at inflammatory sites and by enhancing the killing of phagocytosed bacterial pathogens.
Insights
Neutrophils actively transport tetracyclines like minocycline via a sodium-dependent mechanism. This uptake could improve periodontal therapy by increasing drug levels at inflammatory sites.
Area of Science:
- Pharmacology
- Cell Biology
- Periodontology
Background:
- Tetracyclines are utilized in periodontal therapy for their antimicrobial and matrix metalloproteinase inhibitory properties.
- The precise mechanism by which neutrophils accumulate tetracyclines remains incompletely understood.
Purpose of the Study:
- To characterize the transport mechanism of minocycline and other tetracyclines in isolated human neutrophils.
- To investigate factors influencing tetracycline uptake and efflux by neutrophils.
Main Methods:
- Quantified minocycline and tetracycline transport by measuring increased cell-associated fluorescence in isolated human neutrophils.
- Assessed the influence of extracellular pH, cell activation, and competitive inhibitors on tetracycline transport.
Main Results:
- Neutrophils exhibit saturable, sodium-dependent minocycline uptake with a K(m) of 153 µg/ml.
- Uptake was unaffected by pH variations or activation with phorbol myristate acetate.
- High intracellular/extracellular concentration ratios were observed for minocycline (64.0), doxycycline (7.5), and tetracycline (1.8).
- Efflux of minocycline and doxycycline was triggered by dilution, and transport was competitively inhibited by organic cations.
Conclusions:
- Neutrophil tetracycline transport can be characterized as a concentrative, sodium-dependent process.
- This transport mechanism may enhance the efficacy of tetracyclines in periodontal therapy.
- Sustained therapeutic levels and improved bacterial pathogen killing are potential benefits.
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