RESUMO
Implant insertion can evoke excessive inflammation which disrupts the healing process and potentially leads to complications such as implant rejection. Neutrophils and macrophages play a vital role in the early inflammatory phase of tissue repair, necessitating the study of cellular responses in host-implant interactions. In order to deepen the knowledge about these interactions, the response of neutrophils and macrophages to contact with selected biomaterials was examined in vitro on the basis of secretory response as well as reactive oxygen species/reactive nitrogen species (ROS/RNS) generation. Porcine neutrophils exposed to hydroxyapatite (HA) released more enzymes and generated higher levels of ROS/RNS compared to the control group. The addition of AMPNE diminished these responses. Although the results from porcine cells can provide valuable preliminary data, further validation using human cells or clinical studies would be necessary to fully extrapolate the findings to human medicine. Our study revealed that human neutrophils after contact of with HA increased the production of nitric oxide (NO) (10.00 ± 0.08 vs. control group 3.0 ± 0.11 µM, p < 0.05), while HAP or FAP did not elicit a significant response. Human macrophages cultured with HA produced more superoxide and NO, while HAP or FAP had a minimal effect, and curdlan reduced ROS/RNS generation. The addition of AMPNE to cultures with all biomaterials, except curdlan, reduced neutrophil activity, regardless of the peptides' origin. These results highlight the potential of antimicrobial peptides in modulating excessive biomaterial/host cell reactions involving neutrophils and macrophages, enhancing our understanding of immune reactions, and suggesting that AMPNE could regulate leukocyte response during implantation.
RESUMO
Cationic antimicrobial peptides are molecules with potential applications for treating infections due to their antimicrobial and immunomodulatory properties. The aim of this work was to explore the antimicrobial activity and mechanisms of action of a porcine neutrophil cathelicidin mixture (MPPN). Gram-positive and Gram-negative bacteria were used to determine the minimum inhibitory concentration (MIC) and experiments of both time-kill kinetics and effects on growth curves were performed. Planar black lipid bilayer conductance was measured to analyze the interaction of MPPN with lipid bilayers. Visualization of bacterial surfaces and membrane alterations was achieved using atomic force microscopy and transmission electron microscopy. The effects on the activity of efflux pumps (EPs) were studied with an intracellular accumulation of acridine orange (AO) assay. In E. coli, MPPN behaves as a bactericide at high concentrations and as a bacteriostatic at lower concentrations. The bacteriostatic effect was also observed for slightly shorter periods in S. enterica. The mixture was not active on S. aureus. The increase in AO accumulation in the presence of MPPN indicates that, at least in E. coli, the mixture causes inhibition of the EP function. Observed and detected variable conductance events demonstrate a strong MPPN effect on lipid bilayers. Damage to the structure of treated E. coli indicates that MPPN induces alterations in the bacterial surface. The use of AMPs capable of inhibiting EP can be seen as a good tool to combat antimicrobial resistance since they could be used alone or in combination with other conventional antibiotics to which bacteria have become resistant.
