Characterization and comparison of neutrophil extracellular traps in gingival samples of periodontitis and gingivitis: A pilot study

Abstract

Background and Objective Neutrophil extracellular traps (NETs) are a recently discovered antimicrobial mechanism used by neutrophils that have been proposed as an intervention in the pathogenesis of periodontitis. The objective of our study was to characterize the expression of NETs in gingival tissues with periodontitis and controls and to compare the expression of these traps in gingival tissue samples of patients with gingivitis and periodontitis. Material and Methods An observational cross‐sectional study was conducted on patients with periodontitis, gingivitis, and controls that needed tooth extraction. Gingival tissue biopsies were gathered after clinical examination and tooth extraction. Electron microscopy and immunofluorescence were performed to characterize NETs, comparing periodontitis and control patients. Immunohistochemical analysis was performed to quantify neutrophil extracellular trap expression through extracellular citrullinated histone H3 and myeloperoxidase in biopsies from patients with gingivitis and periodontitis. Results Thirteen biopsies were gathered from 13 patients: five gingivitis, six periodontitis, and two controls. Electron microscopy and immunofluorescence imaging showed greater expression of neutrophils present in periodontal inflamed tissue compared with controls. Release of nuclear content to the extracellular space was observed, compatible with the formation of NETs. The expression of citrullinated histone H3 was higher in gingivitis samples than periodontitis samples (P = 0.0106). Myeloperoxidase expression was higher in periodontitis than gingivitis, but without achieving statistical significance. Conclusion Neutrophil extracellular traps were found in tissue samples of periodontitis as extracellular components of chromatin, along with neutrophil enzymes, that were not present in healthy controls. The comparison of NETs expression in periodontitis and gingivitis showed higher expression in gingivitis, associating them to acute phases of the periodontal inflammatory process.

Neutrophil Extracellular Traps in Periodontitis

“…The comparison of periodontitis and gingivitis showed that NET composition changed, and the general expression of citrullinated histone H3 was found to be higher in gingivitis. These findings suggested that the potential role of NETs in periodontitis may be associated with early and more acute phases of the inflammatory process [63]. With regard to previous results published by our group, we were able to characterize NETs in tissue samples with periodontitis and gingivitis using immunofluorescence, immunohistochemistry and electron microscopy analysis (Figure 2).…”

“…The comparison of periodontitis and gingivitis showed that NET composition changed, and the general expression of citrullinated histone H3 was found to be higher in gingivitis. These findings suggested that the potential role of NETs in periodontitis may be associated with early and more acute phases of the inflammatory process [ 63 ].…”

Influence of Myeloperoxidase Levels on Periodontal Disease: An Applied Clinical Study

“…In accordance with the results of our study, several studies have been demonstrated that situations which may cause an increase of oxidative stress, such as CVD and periodontal disease, cause the high release of CRP, which in turn, can arouse the production of MPO in saliva and serum in order to defend tissue damage determined by oxidative stress condition [36]. In agreement with the results of the present study, Magan-Fernandez et al [39] demonstrated that CRP and MPO levels were higher during active phases of periodontal disease.…”

“…However, there is no unanimous consensus in the literature on the effects of NO levels on tissue damage during periodontitis. Some reports have shown high levels of NO in periodontal tissue in the active periodontal period [39,42] while, on the other hand, other authors have shown lower levels of NO in saliva of subjects with periodontal disease [25,43]. However, results in the literature may have been determined by the different homogeneity of the patients enrolled in the studies, by the different age ranges of the patients analyzed, or by the excessive presence of patients who smoke; in fact it has been shown that smoking can cause a high increase in salivary NO salivary levels [44][45][46].…”

The Role of Neutrophil Extracellular Traps in Periodontitis

“…Therefore, AMPs associated with the DNA backbone would be co-labeled. Citrullinated histones 3 (CitH3), MPO, NET-associated MPO-DNA complexes, histone H1, CD-177, NE, and cathepsin-G (Hirschfeld et al, 2015;Kaneko et al, 2018;Magan-Fernandez et al, 2019) are supplementary NET markers for determining NET-DNA sequence. Although fluorometric detection is commonly used for quantifying NETs, the specificity and objectivity need to be verified, since the results are obtained in photographic form.…”