Macrophages, critical modulators of the immune response, polarize into various phenotypes, including M1 and M2. M1 macrophages are typically activated by lipopolysaccharide and produce proinflammatory cytokines. Conversely, M2 macrophages are activated by stimulation with interleukin 4 (IL)−4 and promote tissue remodeling and anti-inflammatory reactions. Recently, polyunsaturated fatty acids (PUFAs) have been shown to play important roles in the regulation of inflammation. Docosahexaenoic acid (DHA), a PUFA, has antiinflammatory effects on chronic inflammatory disease, but its role in macrophage polarization remains unclear. In this study, we clarified the effects of DHA on macrophage polarization using U937 cells. Treatment with DHA resulted in upregulation of M2 macrophage markers and increased secretion of anti-inflammatory cytokines by U937 cells. IL-4, but not DHA, triggered phosphorylation of signal transducer and activator of transcription 6 (STAT6). DHA enhanced the expression of krüppel-like factor-4 (KLF4), a transcription factor involved in the regulation of macrophage polarization and increased the phosphorylation of p38 mitogen-activated protein kinase (MAPK). A selective inhibitor of p38 MAPK downregulated the expression of CD206 in DHA-treated U937 cells. Moreover, inhibitors of autophagy suppressed the phosphorylation of p38 MAPK and the expression of CD206 in DHA-treated U937 cells.Expression of microtubule-associated protein light chain 3-II, which is involved in autophagosome formation, was enhanced in DHA-treated U937 cells. Taken together, these results indicated that DHA enhanced the expression of M2 macrophage markers through the p38 MAPK signaling pathway and autophagy, suggesting that DHA regulates M2 macrophage polarization and plays an important role in innate immunity. K E Y W O R D Sautophagy, docosahexaenoic acid, macrophage, p38 mitogen-activated protein kinase, polarization

Persister cells are difficult to eliminate because they are tolerant to antibiotic stress. In the present study, using artificially induced Escherichia coli persister cells, we found that reactive oxygen species (ROS) have greater effects on persister cells than on exponential cells. Thus, we examined which types of ROS could effectively eliminate persister cells and determined the mechanisms underlying the effects of these ROS. Ultraviolet (UV) light irradiation can kill persister cells, and bacterial viability is markedly increased under UV shielding. UV induces the production of ROS, which kill bacteria by moving toward the shielded area. Electron spin resonance-based analysis confirmed that hydroxyl radicals are produced by UV irradiation, although singlet oxygen is not produced. These results clearly revealed that ROS sterilizes persister cells more effectively compared to the sterilization of exponential cells (* * p < 0.01). These ROS do not injure the bacterial cell wall but rather invade the cell, followed by cell killing. Additionally, the sterilization effect on persister cells was increased by exposure to oxygen plasma during UV irradiation. However, vapor conditions decreased persister cell sterilization by reducing the levels of hydroxyl radicals. We also verified the effect of ROS against bacteria in biofilms that are more resistant than planktonic cells. Although UV alone could not completely sterilize the biofilm bacteria, UV with ROS achieved complete sterilization. Our results demonstrate that persister cells strongly resist the effects of antibiotics and starvation stress but are less able to withstand exposure to ROS. It was shown that ROS does not affect the cell membrane but penetrates it and acts internally to kill persister cells. In particular, it was clarified that the hydroxy radical is an effective sterilizer to kill persister cells.

Background Bacteria survive in various environments by forming biofilms. Bacterial biofilms often cause significant problems to medical instruments and industrial processes. Techniques to inhibit biofilm formation are essential and have wide applications. In this study, we evaluated the ability of two types of biosurfactants (rhamnolipids and surfactin) to inhibit growth and biofilm formation ability of oral pathogenic bacteria such as Aggregatibacter actinomycetemcomitans, Streptococcus mutans, and Streptococcus sanguinis. Results Rhamnolipids inhibited the growth and biofilm formation ability of all examined oral bacteria. Surfactin showed effective inhibition against S. sanguinis ATCC10556, but lower effects toward A. actinomycetemcomitans Y4 and S. mutans UA159. To corroborate these results, biofilms were observed by scanning electron microscopy (SEM) and confocal microscopy. The observations were largely in concordance with the biofilm assay results. We also attempted to determine the step in the biofilm formation process that was inhibited by biosurfactants. The results clearly demonstrated that rhamnolipids inhibit biofilm formation after the initiation process, however, they do not affect attachment or maturation. Conclusions Rhamnolipids inhibit oral bacterial growth and biofilm formation by A. actinomycetemcomitans Y4, and may serve as novel oral drug against localized invasive periodontitis.

Magnesium hydroxide nanoparticles are widely used in medicinal and hygiene products because of their low toxicity, environment-friendliness, and low cost. Here, we studied the effects of three different sizes of magnesium hydroxide nanoparticles on antibacterial activity: NM80, NM300, and NM700. NM80 (D50 = 75.2 nm) showed a higher bactericidal effect against Escherichia coli than larger nanoparticles (D50 = 328 nm (NM300) or 726 nm (NM700)). Moreover, NM80 showed a high bactericidal effect against not only exponential cells but also persister cells, which are difficult to eliminate owing to their high tolerance to antibiotics. NM80 eliminated strains in which magnesium-transport genes were knocked out and exhibited a bactericidal effect similar to that observed in the wild-type strain. The bactericidal action involved physical cell damage, as confirmed using scanning electron microscopy, which showed that E. coli cells treated with NM80 were directly injured.

Although various caries-preventive agents have been developed, dental caries is still a leading global disease, mostly caused by biological factors such as mutans streptococci. Magnesium hydroxide nanoparticles have been reported to exhibit antibacterial effects; however, they are rarely used in oral care practical applications. In this study, we examined the inhibitory effect of magnesium hydroxide nanoparticles on biofilm formation by Streptococcus mutans and Streptococcus sobrinus—two typical caries-causing bacteria. Three different sizes of magnesium hydroxide nanoparticles (NM80, NM300, and NM700) were studied, all of which inhibited biofilm formation. The results showed that the nanoparticles were important for the inhibitory effect, which was not influenced by pH or the presence of magnesium ions. We also determined that the inhibition process was mainly contact inhibition and that medium (NM300) and large (NM700) sizes were particularly effective in this regard. The findings of our study demonstrate the potential applications of magnesium hydroxide nanoparticles as caries-preventive agents.