Reactive oxygen species and antimicrobial peptides are sequentially produced in silkworm midgut in response to bacterial infection

ABSTRACT All metazoans produce antimicrobial peptides (AMPs) that have both broad antimicrobial and immunomodulatory activity. Cathelicidins are AMPs that preferentially kill Gram-negative bacteria in vitro, purportedly by assembling into higher-order structures that perforate the membrane. We utilized high-resolution, single-cell fluorescence microscopy to examine their mechanism of action in real time. Engineered cathelicidins rapidly bound to Gram-negative and Gram-positive cells and penetrated the cytoplasmic membrane. Rapid failure of the peptidoglycan superstructure in regions of active turnover caused leakage of cytoplasmic contents and the formation of membrane-bound blebs. A mutation anticipated to destabilize interactions between cathelicidin subunits had no effect on bactericidal activity, suggesting that cathelicidins have activities beyond perforating the membrane. Nanomolar concentrations of cathelicidins, although not bactericidal, reduced the growth rate of Gram-negative and Gram-positive bacteria. The cells exhibited expression changes in multiple essential processes, including protein synthesis, peptidoglycan biosynthesis, respiration, and the detoxification of reactive oxygen species (ROS). Time-lapse imaging revealed that ROS accumulation preceded bleb formation, and treatments that reduced cellular ROS levels overcame these bactericidal effects. We propose that that the primary effect of cathelicidins is to induce the production of ROS that damage bacterial molecules, leading to slowed growth or cell death. Given their low circulating levels in vivo, AMPs may serve to slow bacterial population expansion so that cellular immunity systems can respond to and battle the infection. IMPORTANCE Antimicrobial peptides (AMPs) are an important part of the mammalian innate immune system in the battle against microbial infection. How AMPs function to control bacteria is not clear, as nearly all activity studies use nonphysiological levels of AMPs. We monitored peptide action in live bacterial cells over short time frames with single-cell resolution and found that the primary effect of cathelicidin peptides is to increase the production of oxidative molecules that cause cellular damage in Gram-positive and Gram-negative bacteria.

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Aryl hydrocarbon receptor protects against bacterial infection by promoting macrophage survival and reactive oxygen species production

International Immunology ◽

10.1093/intimm/dxt067 ◽

2013 ◽

Vol 26 (4) ◽

pp. 209-220 ◽

Cited By ~ 31

Author(s):

Akihiro Kimura ◽

Hiromi Abe ◽

Sanae Tsuruta ◽

Sayuri Chiba ◽

Yoshiaki Fujii-Kuriyama ◽

...

Keyword(s):

Reactive Oxygen Species ◽

Bacterial Infection ◽

Aryl Hydrocarbon Receptor ◽

Reactive Oxygen Species Production ◽

Reactive Oxygen ◽

Oxygen Species ◽

Aryl Hydrocarbon ◽

Species Production

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Manganese Dioxide Nanozyme for Reactive Oxygen Therapy of Bacterial Infection and Wound Healing

Biomaterials Science ◽

10.1039/d1bm00683e ◽

2021 ◽

Author(s):

Li Liu ◽

Cheng Wang ◽

Yuting Li ◽

Lin Qiu ◽

Shuwen Zhou ◽

...

Keyword(s):

Reactive Oxygen Species ◽

Wound Healing ◽

Soft Tissue ◽

Bacterial Infection ◽

Manganese Dioxide ◽

Soft Tissue Infection ◽

Oxygen Therapy ◽

Reactive Oxygen ◽

Oxygen Species ◽

Central Effector

Reactive oxygen species (ROS) are the weapon of neutrophiles against bacterial pathogens, and also the central effector in the reactive oxygen therapy for skin and soft tissue infection. Nanozymes that...

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The Stimulated Innate Resistance Event in Bordetella pertussis Infection Is Dependent on Reactive Oxygen Species Production

Infection and Immunity ◽

10.1128/iai.00336-13 ◽

2013 ◽

Vol 81 (7) ◽

pp. 2371-2378 ◽

Cited By ~ 8

Author(s):

E. Zurita ◽

G. Moreno ◽

A. Errea ◽

M. Ormazabal ◽

M. Rumbo ◽

...

Keyword(s):

Reactive Oxygen Species ◽

Bacterial Infection ◽

Bordetella Pertussis ◽

Reactive Oxygen ◽

Dependent Manner ◽

Oxygen Species ◽

Pertussis Infection ◽

Content Type ◽

Innate Resistance ◽

Species Production

ABSTRACTThe exacerbated induction of innate immune responses in airways can abrogate diverse lung infections by a phenomenon known as stimulated innate resistance (StIR). We recently demonstrated that the enhancement of innate response activation can efficiently impairBordetella pertussiscolonization in a Toll-like receptor 4 (TLR4)-dependent manner. The aim of this work was to further characterize the effect of lipopolysaccharide (LPS) on StIR and to identify the mechanisms that mediate this process. Our results showed that bacterial infection was completely abrogated in treated mice when the LPS ofB. pertussis(1 μg) was added before (48 h or 24 h), after (24 h), or simultaneously with theB. pertussischallenge (107CFU). Moreover, we detected that LPS completely cleared bacterial infection as soon as 2 h posttreatment. This timing suggests that the observed StIR phenomenon should be mediated by fast-acting antimicrobial mechanisms. Although neutrophil recruitment was already evident at this time point, depletion assays using an anti-GR1 antibody showed thatB. pertussisclearance was achieved even in the absence of neutrophils. To evaluate the possible role of free radicals in StIR, we performed animal assays using the antioxidantN-acetyl cysteine (NAC), which is known to inactivate oxidant species. NAC administration blocked theB. pertussisclearance induced by LPS. Nitrite concentrations were also increased in the LPS-treated mice; however, the inhibition of nitric oxide synthetases did not suppress the LPS-induced bacterial clearance. Taken together, our results show that reactive oxygen species (ROS) play an essential role in the TLR4-dependent innate clearance ofB. pertussis.

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Reactive oxygen species play no role in the candidacidal activity of the salivary antimicrobial peptide histatin 5

Biochemical Journal ◽

10.1042/bj20040208 ◽

2004 ◽

Vol 381 (2) ◽

pp. 447-452 ◽

Cited By ~ 56

Author(s):

Enno C. I. VEERMAN ◽

Kamran NAZMI ◽

Wim van 't HOF ◽

Jan G. M. BOLSCHER ◽

Alice L. den HERTOG ◽

...

Keyword(s):

Reactive Oxygen Species ◽

Antimicrobial Peptides ◽

Reactive Oxygen ◽

Inhibitory Effect ◽

Oxygen Species ◽

Histatin 5 ◽

A Cell ◽

Synthetic Derivative ◽

Ros Scavenger

The mechanism of action of antimicrobial peptides is still a matter of debate. The formation of ROS (reactive oxygen species) has been suggested to be the crucial step in the fungicidal mechanism of a number of antimicrobial peptides, including histatin 5 and lactoferrin-derived peptides. In the present study we have investigated the effects of histatin 5 and of a more amphipathic synthetic derivative, dhvar4, on the generation of ROS in the yeast Candida albicans, using dihydroethidium as an indicator for ROS. With both peptides, a substantial enhancement of fluorescence was observed. However, TEMPO (2,2,6,6-tetramethylpiperidine-N-oxyl), a cell-permeant ROS scavenger, did not have an inhibitory effect on killing or on the enhancement of fluorescence. Furthermore, antimycin and azide, which have been reported to induce ROS in vitro, were not able to enhance the dihydroethidium fluorescence, while chlorhexidine, a non-specific antiseptic agent, enhanced dihydroethidium fluorescence to the same extent as did the peptides. Fluorescence microscopy showed the fluorescence enhancement to be a consequence of the release of unbound preformed ethidium from the mitochondrial matrix within the cell. It is concluded that ROS do not play a role in the histatin 5-mediated killing of C. albicans.

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