Influence of the Protein Kinase C Activator Phorbol Myristate Acetate on the Intracellular Activity of Antibiotics against Hemin- and Menadione-Auxotrophic Small-Colony Variant Mutants of Staphylococcus aureus and Their Wild-Type Parental Strain in Human THP-1 Cells

Abstract

cIn a previous study (L. G. Garcia et al., Antimicrob. Agents Chemother. 56:3700 -3711, 2012), we evaluated the intracellular fate of menD and hemB mutants (corresponding to menadione-and hemin-dependent small-colony variants, respectively) of the parental COL methicillin-resistant Staphylococcus aureus strain and the pharmacodynamic profile of the intracellular activity of a series of antibiotics in human THP-1 monocytes. We have now examined the phagocytosis and intracellular persistence of the same strains in THP-1 cells activated by phorbol 12-myristate 13-acetate (PMA) and measured the intracellular activity of gentamicin, moxifloxacin, and oritavancin in these cells. Postphagocytosis intracellular counts and intracellular survival were lower in PMA-activated cells, probably due to their higher killing capacities. Gentamicin and moxifloxacin showed a 5-to 7-fold higher potency (lower static concentrations) against the parental strain, its hemB mutant, and the genetically complemented strain in PMA-activated cells and against the menD strain in both activated and nonactivated cells. This effect was inhibited when cells were incubated with N-acetylcysteine (a scavenger of oxidant species). In parallel, we observed that the MICs of these drugs were markedly reduced if bacteria had been preexposed to H 2 O 2 . In contrast, the intracellular potency of oritavancin was not different in activated and nonactivated cells and was not decreased by the addition of N-acetylcysteine, regardless of the phenotype of the strains. The oritavancin MIC was also unaffected by preincubation of the bacteria with H 2 O 2 . Thus, activation of THP-1 cells by PMA may increase the intracellular potency of certain antibiotics (probably due to synergy with reactive oxygen species), but this effect cannot be generalized to all antibiotics.

TheATG16L1–T300Aallele impairs clearance of pathosymbionts in the inflamed ileal mucosa of Crohn's disease patients

“…Interestingly, such a difference is not observed when monocytes were not exposed to PMA. PMA activates protein kinase C and triggers reactive oxygen species production, thereby causing oxidative stress and increased production of inflammatory cytokines, providing an in vitro model for inflammation 39. In addition, we show that exposure to either IL-1β or TNF-α enhanced the survival of AIEC in monocytes homozygous for the risk allele compared with monocytes homozygous for the protective allele, though these differences were not significant.…”

Pharmacodynamic Evaluation of the Intracellular Activity of Antibiotics towards Pseudomonas aeruginosa PAO1 in a Model of THP-1 Human Monocytes

“…A switch from apparent intracellular persistence to a net increase in the number of intracellular bacteria is probably related to an equilibrium among intrinsic bacterial growth properties, expression of virulence factors, and the level of host defenses. The latter are notoriously poor in THP-1 cells (32). Electron microscopy suggests that intracellular P. aeruginosa remains confined in vacuoles in THP-1 cells for at least 24 h and can actively replicate therein.…”

Antibiotic activity against small-colony variants of Staphylococcus aureus: review of in vitro, animal and clinical data

“…Second, this higher potency was seen for the haemin-dependent mutant and its parental strain, but not for the menadionedependent mutant. 35,36 The latter may not have been influenced by cell activation simply because the antibiotics already showed a higher potency towards this strain in non-activated cells. 36 Thus, these data suggest that the menadione-dependent strain is hypersusceptible to oxidant species (see Figure 1 for an illustration of the potential link between menadione dependence and susceptibility to oxidant species).…”