Pathogen-targeted hydroxyl radical generation during melanization in insect hemolymph: EPR study of a probable cytotoxicity mechanism

• Published in: Applied magnetic resonance Vol. 35; no. 4; pp. 495 - 501
• Main Authors: Komarov, Denis A., Ryazanova, A. D., Slepneva, I. A., Khramtsov, V. V., Dubovskiy, I. M., Glupov, V. V.
• Format: Journal Article
• Language: English
• Published: Vienna Springer Vienna 01-05-2009; Springer Nature B.V
• Subjects: Atoms and Molecules in Strong Fields; Catalase; Cytotoxicity; Destruction; Electron paramagnetic resonance; Electron spin; Encapsulation; Galleria mellonella; Hydrogen peroxide; Hydroxyl radicals; Immune system; Insects; Laser Matter Interaction; Organic Chemistry; Oxygen; Pathogens; Physical Chemistry; Physics; Physics and Astronomy; Sodium azides; Solid State Physics; Spectroscopy/Spectrometry; Toxicity; Pathogenic Organism; Spin Trap; Electron Paramagnetic Resonance; Xylenol Orange; Electron Paramagnetic Resonance Spectrum
• ISSN: 0937-9347; 1613-7507
• DOI: 10.1007/s00723-009-0180-6

The main component of the insect immune system is melanotic encapsulation of pathogenic organisms. Molecular mechanisms of destruction of an encapsulated pathogen are poorly understood. Reactive oxygen species (ROS) are considered as probable cytotoxic agents responsible for destruction of pathogenic organisms in insect hemolymph. In the present work the formation of H
2
O
2
during melanization in
Galleria mellonella
hemolymph in the presence of catalase inhibitor NaN
3
was detected. Enhanced rates of H
2
O
2
generation were observed in the hemolymph of insects activated by injection of bacterial cells. Using spin trapping technique in combination with electron paramagnetic resonance spectroscopy we demonstrated that production of H
2
O
2
in the hemolymph causes the formation of highly toxic reactive oxygen species, hydroxyl radical. However, neither H
2
O
2
nor hydroxyl radical were detected in the absence of NaN
3
in agreement with the high catalase activity in the hemolymph. These observations allow us to propose a unique mechanism of pathogen-targeted cytotoxicity based on localized hydroxyl radical generation within a melanotic capsule.