Salivary Proteomics Reveals Significant Changes in Relation to Alzheimer’s Disease and Aging

Abstract

Background: Aging is a risk factor for several pathologies as Alzheimer’s disease (AD). Great interest exists, therefore, in discovering diagnostic biomarkers and indicators discriminating biological aging and health status. To this aim, omic investigations of biological matrices, as saliva, whose sampling is easy and non-invasive, offer great potential. Objective: Investigate the salivary proteome through a statistical comparison of the proteomic data by several approaches to highlight quali-/quantitative variations associated specifically either to aging or to AD occurrence, and, thus, able to classify the subjects. Methods: Salivary proteomic data of healthy controls under-70 (adults) and over-70 (elderly) years old, and over-70 AD patients, obtained by liquid chromatography/mass spectrometry, were analyzed by multiple Mann-Whitney test, Kendall correlation, and Random-Forest (RF) analysis. Results: Almost all the investigated proteins/peptides significantly decreased in relation to aging in elderly subjects, with or without AD, in comparison with adults. AD subjects exhibited the highest levels of α-defensins, thymosin β4, cystatin B, S100A8 and A9. Correlation tests also highlighted age/disease associated differences. RF analysis individuated quali-/quantitative variations in 20 components, as oxidized S100A8 and S100A9, α-defensin 3, P-B peptide, able to classify with great accuracy the subjects into the three groups. Conclusion: The findings demonstrated a strong change of the salivary protein profile in relation to the aging. Potential biomarkers candidates of AD were individuated in peptides/proteins involved in antimicrobial defense, innate immune system, inflammation, and in oxidative stress. RF analysis revealed the feasibility of the salivary proteome to discriminate groups of subjects based on age and health status.

Top-Down Proteomics Detection of Potential Salivary Biomarkers for Autoimmune Liver Diseases Classification

“…RF analysis, one of several up-to-date machine learning methods, evidenced a panel of proteins/peptides present in saliva able to correctly classify PBCp group with respect to HCs with an 83.3% accuracy, AIHp group with respect to HCs with an 79.9% accuracy and PBCp with respect to AIHp with an 80.2% accuracy. The same statistical approach has been successfully used to classify groups of subjects based on age and health status highlighting the feasibility of the salivary proteome to discriminate groups of subjects based on physiological or pathological condition not only confined to oral cavity [ 75 ]. RF results were confirmed by LDA which provided a very good classification of subject based on PBC or AIH occurrence and with respect to HCs.…”

Characterization of Cystatin B Interactome in Saliva from Healthy Elderly and Alzheimer’s Disease Patients

“…This explorative proteomic study, based on an AP-MS approach, was inspired by the results that we obtained in previous top-down proteomic investigations on the salivary proteome associated with Alzheimer’s disease. We previously demonstrated that some salivary proteins/peptides, including S-modified proteoforms of cystatin B, significantly varied in abundance between AD patients and healthy controls [ 14 , 15 ]. Due to the relevance of developing proteomic studies in peripheral biofluids to the AD-associated biomarker discovery and the known ability of cystatin B to interact in multi-protein complexes [ 24 , 25 , 26 , 27 ] and considering the significance of PPI studies in the field of neurodegenerative disease, we decided to investigate the possible PPI of cystatin B in AD saliva.…”

“…Altered levels of cystatin B have been observed in the context of AD, where high amounts of the protein were found post-mortem in the brains of patients [ 13 ]. Our recent studies on the salivary proteome in AD patients evidenced significantly higher levels of cystatin B compared to age-matched elderly healthy individuals [ 14 , 15 ], suggesting the existence of a self-protection mechanism in the patients, which could reflect the neuronal protective role of cystatin B in the brain [ 14 , 15 ]. AD is the most common neurodegenerative disease (ND) afflicting the elderly population, and its principal hallmarks are misfolded beta-amyloid (Aβ) and tau proteins in the central nervous system (CNS) [ 16 ].…”

“…In vitro studies have shown that cystatin B oligomers can inhibit or facilitate Aβ fibril growth according to their size [ 25 ]. These properties of cystatin B, our previous insights on salivary proteoforms of cystatin B associated with AD [ 14 , 15 ], and the relevance of the characterization of peripheral potential biomarkers and of their interactome in AD led us to investigate the possible involvement of cystatin B in a PPI network in the oral cavity. The feasibility, non-invasiveness, painlessness, and low-cost of collecting saliva make this biofluid an optimal choice to discover protein biomarkers useful for the diagnosis and risk assessment of several diseases, including AD [ 28 , 29 ], as opposed to blood and CSF [ 20 ].…”

“…In addition, in AD patients, the function of the major salivary glands and their secretion can be altered [ 28 , 30 , 31 ]. Saliva is a biofluid that has been investigated in the AD field [ 14 , 15 , 28 , 30 , 31 ], and to date, several salivary biomarkers of AD have been studied by different approaches, even if with contrasting outcomes [ 20 , 28 , 31 , 32 ].…”