Platelet microbicidal proteins and neutrophil defensin disrupt the Staphylococcus aureus cytoplasmic membrane by distinct mechanisms of action.

Abstract

Platelet microbicidal proteins (PMPs) are hypothesized to exert microbicidal effects via cytoplasmic membrane disruption. Transmission electron microscopy demonstrated a temporal association between PMP exposure, damage of the Staphylococcus aureus cytoplasmic membrane ultrastructure, and subsequent cell death. To investigate the mechanisms of action of PMPs leading to membrane damage, we used flow cytometry to compare the effects of two distinct PMPs (thrombin-induced PMP-1 [tPMP-1] or PMP-2) with human neutrophil defensin-1 (hNP-1) on transmembrane potential (Deltapsi), membrane permeabilization, and killing of S. aureus. Related strains 6850 (Deltapsi -150 mV) and JB-1 (Deltapsi -100 mV; a respiration-deficient menadione auxotroph of 6850) were used to assess the influence of Deltapsi on peptide microbicidal effects. Propidium iodide (PI) uptake was used to detect membrane permeabilization, retention of 3,3'-dipentyloxacarbocyanine (DiOC5) was used to monitor membrane depolarization (Deltapsi), and quantitative culture or acridine orange accumulation was used to measure viability. PMP-2 rapidly depolarized and permeabilized strain 6850, with the extent of permeabilization inversely related to pH. tPMP-1 failed to depolarize strain 6850, but did permeabilize this strain in a manner directly related to pH. Depolarization, permeabilization, and killing of strain JB-1 due to PMPs were significantly less than in strain 6850. Growth in menadione reconstituted Deltapsi of JB-1 to a level equivalent to 6850, and was associated with greater depolarization due to PMP-2, but not tPMP-1. Reconstitution of Deltapsi also enhanced permeabilization and killing of JB-1 due to tPMP-1 or PMP-2. Both PMP-2 and tPMP-1 caused significant reductions in viability of strain 6850. In contrast to tPMP-1 or PMP-2, defensin hNP-1 depolarized, permeabilized, and killed both strains 6850 and JB-1 equally, and in a manner directly related to pH. Collectively, these data indicate that membrane dysfunction and cell death due to tPMP-1, PMP-2, or hNP-1 likely involve different mechanisms. These findings may also reveal new insights into the microbicidal activities versus mammalian cell toxicities of antimicrobial peptides.

The use of autologous platelet–leukocyte gels to enhance the healing process in surgery, a review

“…Furthermore, Yeaman et al [57] and Tang et al [58] support the idea that platelets also are involved in microbicidal activity, suggesting that they play a role in the platelet host defense mechanism by releasing a variety of platelet microbicidal proteins. The platelet microbicidal proteins were shown to be released after platelet acti-vation, demonstrating potent activities against pathogens that have a tendency to enter the bloodstream [59].…”

Lipopeptides as anti-infectives: a practical perspective

“…Recently, it was demonstrated that telavancin's antibacterial activity derives from at least two mechanisms of action. The second mechanism of action of telavancin is related to perturbation of the electrochemical potential and permeability of the bacterial cell membrane, causing membrane permeabilization similarly to the mode of action of antimicrobial peptides [40,102,104,105]. Dissipation of the cell membrane potential is concentration-dependent, rapid (15 minutes), and can be detected starting with telavancin at its 10-fold MIC values [102].…”

Scorpion venom as antimicrobial peptides (AMPs): A review article

“…Microorganisms constitutively resist the action of AMPs by using different mechanisms, for example resistance strains of Staphylococcus aureuscontain high D-alanylation of lipoteichoic acid with positively charged amine groups resulted in lowering the membrane negatively charge so lack of electrostatic affinity for AMPs toward membranes [45], another mechanism by membrane energy alteration, studies reports that microorganisms with altered membrane energetic are more resistance to AMPs action than microorganisms with normal membrane energy status, such as Candida albicans a respiratory deficient mutant fungi with membrane energy altered by lowered mitochondrial ATP synthesis [46] and S. aureus with membrane energy altered by constitutively reduced transmembrane potential [47]. Electrostatic shielding mechanism used also by microorganisms to resist the action of AMPs in which membranes have been covered with highly anionic glycocalyx or special capsule resulted in shield the lipid bilayer from AMPs [19].…”