Alpha-Defensin 1: An Emerging Periodontitis Biomarker

Abstract

Background: Research on the development of reliable diagnostic targets is being conducted to overcome the high prevalence and difficulty in managing periodontitis. However, despite the development of various periodontitis target markers, their practical application has been limited due to poor diagnostic accuracy. In this study, we present an improved periodontitis diagnostic target and explore its role in periodontitis. Methods: Gingival crevicular fluid (GCF) was collected from healthy individuals and periodontitis patients, and proteomic analysis was performed. The target marker levels for periodontitis were quantified in GCF samples by enzyme-linked immunosorbent assay (ELISA). Mouse bone marrow-derived macrophages (BMMs) were used for the osteoclast formation assay. Results: LC-MS/MS analysis of whole GCF showed that the level of alpha-defensin 1 (DEFA-1) was higher in periodontitis GCF than in healthy GCF. The comparison of periodontitis target proteins galactin-10, ODAM, and azurocidin proposed in other studies found that the difference in DEFA-1 levels was the largest between healthy and periodontitis GCF, and periodontitis was more effectively distinguished. The differentiation of RANKL-induced BMMs into osteoclasts was significantly reduced by recombinant DEFA-1 (rDEFA-1). Conclusions: These results suggest the regulatory role of DEFA-1 in the periodontitis process and the relevance of DEFA-1 as a diagnostic target for periodontitis.

Keywords: alpha-defensin 1; gingival crevicular fluids; inflammation; neutrophile; osteoclast; periodontitis biomarker.

Figure 1

Western blot analysis for DEFA-1. Twenty-five micrograms of protein from a GCF sample from healthy subjects (H-GCF) and patients with periodontitis (P-GCF) was separated on 15% acrylamide gel electrophoresis, and a Western blot was performed with DEFA-1-specific primary antibody.

Figure 2

ELISA analysis for DEFA-1, galectin-10, ODAM, and azurocidin. Concentrations of target proteins were quantified in GCF from healthy subjects (H-GCF, n = 85) and patients with periodontitis (P-GCF, n = 130). p value vs. H-GCF.

Figure 3

Effect of recombinant DEFA-1 protein (rDEFA) on RANKL-induced osteoclastic bone resorption. BMM cells were treated with M-CSF (30 ng/mL) and RANKL (100 ng/mL) with or without rDEFA (10 or 50 µg/mL) for 5 days and were stained to detect the expression of TRAP. The total number of TRAP-positive multinucleated (≥3 nuclei) osteoclasts (MNCs) per well is graphically presented. The data are expressed as the mean ± SD. p vs. M-CSF + RANKL condition. ns = non-specific.