The Potential of Telomeric G-Quadruplexes Containing Modified Oligoguanosine Overhangs in Activation of Bacterial Phagocytosis and Leukotriene Synthesis in Human Neutrophils

Abstract

Human neutrophils are the first line of defense against bacterial and viral infections. They eliminate pathogens through phagocytosis, which activate the 5-lipoxygenase (5-LOX) pathway resulting in synthesis of leukotrienes. Using HPLC analysis, flow cytometry, and other biochemical methods, we studied the effect of synthetic oligodeoxyribonucleotides (ODNs) able to fold into G-quadruplex structures on the main functions of neutrophils. Designed ODNs contained four human telomere TTAGGG repeats (G4) including those with phosphorothioate oligoguanosines attached to the end(s) of G-quadruplex core. Just modified analogues of G4 was shown to more actively than parent ODN penetrate into cells, improve phagocytosis of Salmonella typhimurium bacteria, affect 5-LOX activation, the cytosol calcium ion level, and the oxidative status of neutrophils. As evident from CD and UV spectroscopy data, the presence of oligoguanosines flanking G4 sequence leads to dramatic changes in G-quadruplex topology. While G4 folds into a single antiparallel structure, two main folded forms have been identified in solutions of modified ODNs: antiparallel and dominant, more stable parallel. Thus, both the secondary structure of ODNs and their ability to penetrate into the cytoplasm of cells are important for the activation of neutrophil cellular effects. Our results offer new clues for understanding the role of G-quadruplex ligands in regulation of integral cellular processes and for creating the antimicrobial agents of a new generation.

Magic Peptide: Unique Properties of the LRR11 Peptide in the Activation of Leukotriene Synthesis in Human Neutrophils

“…We have previously shown [ 2 , 3 ] that synthetic oligonucleotides of various types containing phosphorothioate fragments, to one degree or another, induce the generation of reactive oxygen species by neutrophilic granulocytes. Leukotriene synthesis is regulated by multiple mechanisms, and reactive oxygen species not only provide some peroxide tone required to activate 5-LOX, but through the initiation of NADPH-oxidase-dependent superoxide synthesis, lead to a decrease in NOS activity and protect against NO-induced inhibition of 5-LOX [ 22 , 36 ].…”

“…To purify the water/methanol extracts, solid-phase extraction was used on Sep-Pak C18 cartridges (500 mg; Macherey-Nagel, Dueren, Germany), which were conditioned first with methanol, and then with water. Major 5-LOX metabolites were identified by comparing retention times with those of known compounds, as previously described [ 2 ].…”

“…They eliminate pathogens through phagocytosis, which activates the 5-lipoxygenase (5-LOX)-mediated pathway leading to the synthesis of leukotrienes. Recently, it was found that the leukotriene synthesis in human neutrophils is regulated by genome G-rich DNA fragments and bacterial DNA degradation products—oligodeoxyribonucleotides (ODNs) containing unmethylated CpG motifs (CpG-ODNs) that are released from the inflammatory region [ 1 , 2 , 3 ]. Using synthetic oligonucleotides that mimic these DNA fragments, it was shown that G-rich ODNs capable of folding into non-B form G-quadruplex structures activated bacterial phagocytosis and the leukotriene synthesis in human neutrophils, both the secondary structure of ODNs and their ability to penetrate into cells being important for the induction of immune responses [ 2 ].…”

Modulation of the Innate Immune Response by Targeting Toll-like Receptors: A Perspective on Their Agonists and Antagonists

“…111,112 This event culminates in the development of an innate immune response, which can be applied to limit the spread of pathogens and enhance adaptive immunity, which can be exploited to improve vaccines. 113 Taken together, these considerations make TLR9 one of the most promising targets for fighting against infectious diseases. Chuang et al reported that several synthetic CpG-ODNs generate potent immune stimuli by inducing the production of pro-inflammatory cytokines and a T H 1-dependent immune response.…”

“…304 Noteworthy, 34, acting via TLR4-MyD88/ TRIF-NF-κB signaling pathway, was able to reduce traumatic brain injury-induced neurons loss, brain edema, and neurobehavioral impairment in rats, subsequently inhibiting neuronal autophagy and neuroinflammation responses in the hippocampus. 305 In inflammation-induced primary microglia, dihydrotestosterone (113) acted as a TLR4 antagonist by limiting LPS-mediated activation, NF-κB, and MAPK p38 signaling pathways and attenuating the release of TNF-α, IL-1β, IL-6, iNOS, COX-2, NO, and PGE2. 306 In another work, the cyanobacteria-derivative TLR4 inhibitor VB3323 was able to decrease microglial activation and morphological alterations of spinal cord neurons.…”