Adult patients with periodontal disease exhibit tooth malocclusions such as flaring of the anterior segment. This unaesthetic appearance and/or dysfunction are the primary reasons for undergoing periodontal-orthodontic treatment using a multidisciplinary approach
. Combined orthodontic and periodontal treatment does not compromise the therapeutic effectiveness of the latter
. Indeed, this combination may be beneficial since it leads to increased bone resorption stability, a lack of incisor flaring, and improvements in bone defects
. However, some authors argue that orthodontic treatment is unsuitable in chronic periodontal patients because bacterial plaque is an etiologic factor in the development of periodontitis and the presence of orthodontic appliances facilitates plaque growth and maturation. Thus, the periodontal condition could worsen after orthodontic treatment
. Therefore, we examined periodontal-orthodontic patients with a view to elucidating the differences between them and non-periodontal orthodontic patients.
Such differences could be caused by inflammatory factors related to periodontitis or other protein factors
, which may represent candidate biomarkers. A candidate biomarker is defined as an informative signal associated with a specific condition. The specificity and sensitivity of a biomarker describe its usefulness in diagnosing a specific condition or predicting its progress
. An effective biomarker should be measurable in an accessible body fluid, such as serum, urine, or saliva
. Saliva contains abundant proteins, peptides, and other small molecules
. Thus, the salivary peptide spectrum may be applied widely in the diagnosis and monitoring of oral diseases. Progress has been made in screening for not only oral diseases such as oral cancer
, but also systematic conditions such as gastric cancer
 and breast cancer
. Studies of periodontitis using proteomics
[32, 33] have been performed, and the demand for periodontal-orthodontic combination treatment is increasing. However, little research on the salivary peptide spectrum in the periodontal-orthodontic area has been conducted.
In the present study, we used MALDI-TOF MS-based proteomic methods and WCX magnetic beads to examine all 24 saliva samples. Nine peaks that differed significantly were found (Figure
2), of which two (3163.4 and 1454.2 Da) exhibited the most significant differences (p <0.01). Orthodontic treatment is a prolonged procedure. And in our previous study, alterations were found in salivary proteins due to different orthodontic treatment durations
. In this study, the peak intensities of the three different groups showed differential expression of salivary peptides, indicating the orthodontic treatment could contribute to the change of salivary peptidome. Noticeably, significant differences were investigated between the two orthodontic groups. This suggested that differential expression of salivary peptidome profile did exist between orthodontic patients with and without periodontitis. Thus, this method provides a new tool for analyzing the effect of periodontitis on orthodontic treatment.
The peptide sequence identifications made in this study have led to interesting speculations. The 3163.4-Da peptide was identified as F2 prothrombin precursor. Thrombin is a ‘trypsin-like’ serine protease protein encoded by the F2 gene. Beyond its key role in the dynamic process of thrombus formation, thrombin has the potential to exert actions such as inflammation and leukocyte recruitment
. Moreover, isoform 1 of fibrinogen alpha chain precursor (FGA), which was the predicted identity of the 3154.4- and 2621.9-Da peptides, is encoded by FGA. It is cleaved by thrombin to form fibrin, indicating that it interacts with thrombin. Some studies
 suggested epithelial thrombomodulin (TM), which binds to thrombin and converts it from a procoagulant protease to an anticoagulant enzyme, increased in gingival crevicular fluid of individuals with chronic periodontitis. Moreover, gingipains, the major periodontopathic bacterium Porphyromonas gingivalis-derived cysteine proteases, lead to the degradation of endothelial TM. In addition, reduced expression of TM was found in gingival microvascular endothelia in patients with periodontitis
, and this may be involved in the pathogenesis of periodontitis. Thus, these differential expression patterns of altered proteins may have originated from periodontitis-associated inflammation or differences in bone metabolism between orthodontic patients with and without periodontitis. Ultimately, our aim is to determine the protein or gene from which a peptide is derived; however, this is complex. When the distribution of the peptides with low molecular weight were intended to match to the mass spectrometry spectra relative peak area, it should be noted that these peptides had complicated origins. Peptides in saliva could be secreted peptides and proteolytic fragments of related proteins
. Moreover, these components are subject to secondary modifications from distinctive protein families. Thus, a peptide sequence usually does not exclusively define a single protein.
As these specific peptides were investigated using this relatively new combined method with no validation using other techniques, thus, a larger sample size and repeated trial or trial using other techniques are needed to confirm the significant differences in peptide mass peaks found in this study and its reproducibility. The establishment of a relatively complete protein-peptide spectrum database will facilitate the determination of both the source of the salivary protein profile variation and the mechanism thereof. Moreover, the analysis of saliva is inherently challenging because of the wide protein concentration range therein
 and the presence of multiple post-translational modifications
. Further research into the composition of saliva will likely provide novel tools for investigations of physiological and pathophysiological states.