Proteomics Disclose the Potential of Gingival Crevicular Fluid (GCF) as a Source of Biomarkers for Severe Periodontitis

Abstract

Periodontal disease is a widespread disorder comprising gingivitis, a mild early gum inflammation, and periodontitis, a more severe multifactorial inflammatory disease that, if left untreated, can lead to the gradual destruction of the tooth-supporting apparatus. To date, effective etiopathogenetic models fully explaining the clinical features of periodontal disease are not available. Obviously, a better understanding of periodontal disease could facilitate its diagnosis and improve its treatment. The purpose of this study was to employ a proteomic approach to analyze the gingival crevicular fluid (GCF) of patients with severe periodontitis, in search of potential biomarkers. GCF samples, collected from both periodontally healthy sites (H-GCF) and the periodontal pocket (D-GCF), were subjected to a comparison analysis using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). A total of 26 significantly different proteins, 14 up-regulated and 12 down-regulated in D-GCF vs. H-GCF, were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). The main expressed proteins were inflammatory molecules, immune responders, and host enzymes. Most of these proteins were functionally connected using the STRING analysis database. Once validated in a large scale-study, these proteins could represent a cluster of promising biomarkers capable of making a valuable contribution for a better assessment of periodontitis.

Keywords: SDS-PAGE; biomarkers; gingival crevicular fluid; mass spectrometry; periodontal disease; periodontitis; proteomics.

Figure 1

The representative SDS-PAGE image. First lane, MW: molecular weight marker (kDa), Dual Blue Precision Plus Protein Standard. Lanes 2–3, H-GCF: representative pools of H-GCF samples. Lanes 4–5, D-GCF: representative pools of D-GCF samples. The differentially expressed bands are enclosed within lanes. Gel separation: gradient Bolt 4–12% Bis-Tris Plus, Coomassie Blue stain method.

Figure 2

The functional interaction network obtained using STRING. Proteins are indicated with their gene names. Blue nodes = up-regulated proteins in D-GCF: CALR, calreticulin; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HP, haptoglobin; HPX, hemopexin; SERPINA3, alpha-1-antichymotrypsin; A2M, alpha-2-macroglobulin; SERPINB1, leukocyte elastase inhibitor. Red nodes = down-regulated proteins in D-GCF: ENO1, alpha-enolase; SERPINA1, alpha-1-antitrypsin; KRT19, keratin, type I cytoskeletal 19; C3, complement C3; GC, Vitamin-D-binding protein; KRT14, keratin, type I cytoskeletal 14; KRT16, keratin, type I cytoskeletal 16; KRT6A, keratin, type II cytoskeletal 6A; KRT13, keratin, type I cytoskeletal 13; KRT4, keratin, type II cytoskeletal 4.