1995
DOI: 10.1128/aac.39.8.1676
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Role of extracellular calcium in in vitro uptake and intraphagocytic location of macrolides

Abstract: We compared the uptakes and intracellular locations of four 14-membered-ring macrolides (roxithromycin, dirithromycin, erythromycin, and erythromycylamine) in human polymorphonuclear neutrophils (PMNs) in vitro. Intracellular location was assessed by cell fractionation and uptake kinetics in cytoplasts (granule-poor PMNs). Trapping of dirithromycin within PMN granules (up to 80% at 30 min) was significantly more marked than the intracellular trapping of the other drugs (erythromycylamine, 45% ؎ 5.1%; erythromy… Show more

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Cited by 44 publications
(45 citation statements)
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“…The present experiments have been limited to J774 macrophages, which were used as a model, and generalization of our conclusions to other phagocytic cells therefore remains unwarranted. In particular, the presence of a functional P-glycoprotein and its potential role in macrolide handling in polymorphonuclear leukocytes are controversial (4,14,21,38). It is interesting, however, that human KB and G-185 cells overexpressing P-glycoprotein show decreased levels of macrolide accumulation and correspondingly reduced levels of antimicrobial activity against intracellular forms of Listeria monocyto- genes (22).…”
Section: Discussionmentioning
confidence: 99%
“…The present experiments have been limited to J774 macrophages, which were used as a model, and generalization of our conclusions to other phagocytic cells therefore remains unwarranted. In particular, the presence of a functional P-glycoprotein and its potential role in macrolide handling in polymorphonuclear leukocytes are controversial (4,14,21,38). It is interesting, however, that human KB and G-185 cells overexpressing P-glycoprotein show decreased levels of macrolide accumulation and correspondingly reduced levels of antimicrobial activity against intracellular forms of Listeria monocyto- genes (22).…”
Section: Discussionmentioning
confidence: 99%
“…23 Cells were maintained in HBSS. The viability, measured by the release of lactate dehydrogenase (LDH) 29 and purity of the final preparation were greater than 99 and 95%, respectively. PMN degranulation was obtained by addition of formyl-methionyl-leucyl-phenylalanine (fMLP) at 10 Ϫ6 M for 10 minutes, at 37°C (500 l of conditioned medium corresponding to the release of 5 ϫ 10 5 activated PMN were used throughout the experimental procedure).…”
Section: In Vitro Studies Of Smc-pmn-fibrin Interactionsmentioning
confidence: 99%
“…Aliquots of, 2.5 ϫ 10 6 PMN were loaded with the drugs at 10 g/ml (30 min at 37°C) and were centrifuged through the oil cushion. The cell pellet was sonicated in the presence of 0.5% Triton X-100 or 0.73 M sucrose to protect granules (17,23,24). After centrifugation, the amounts of lysozyme (a granule marker) and radiolabeled drugs in the pellet and the supernatant were determined as previously described (18).…”
Section: Human Neutrophils (Pmn)mentioning
confidence: 99%
“…The following experimental conditions were varied to study the mechanism of uptake: pretreatment of PMN with 10% formaldehyde followed with two washes in HBSS (for cell viability); pH from 6.5 to 8.5; temperatures of 0, 20, 37, and 40°C; extracellular concentrations of 2.5 to 100 g/ml; pretreatment for 15 min with various metabolic inhibitors (NaF, KCN, or 2,4-dinitrophenol; 1 mM) or with various activators and/or inhibitors of PMN functions which have been reported to interfere with macrolide uptake (17,23,24), namely Ni 2ϩ , a blocker of the Na ϩ -Ca 2ϩ exchanger, at 1.25 to 5 mM; addition of phorbol myristate acetate (PMA), a protein kinase C (PKC) activator, at 100 and 10 ng/ml; addition of H89, a PKA inhibitor, at 50 M; and addition of verapamil, a Ca 2ϩ channel blocker, at 125 and 250 M. We also studied the inhibitory effect of various macrolides and ketolides; the concentrations chosen were the mean K m (23,24). PMN were incubated at 37°C for 5 min with unlabeled azithromycin (51 g/ml), HMR 3004 (22 g/ml), HMR 3647 (117 g/ml), or HBSS.…”
Section: Human Neutrophils (Pmn)mentioning
confidence: 99%
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