{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,16]],"date-time":"2026-10-16T20:14:59Z","timestamp":1792181699681,"version":"4.4.0"},"reference-count":60,"publisher":"MDPI AG","issue":"21","license":[{"start":{"date-parts":[[2021,11,6]],"date-time":"2021-11-06T00:00:00Z","timestamp":1636156800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Molecules"],"abstract":"<jats:p>Our studies aimed to explore the protein components of the matrix of human submandibular gland sialoliths. A qualitative analysis was carried out based on the filter aided sample preparation (FASP) methodology. In the protein extraction process, we evaluated the applicability of the standard demineralization step and the use of a lysis buffer containing sodium dodecyl sulfate (SDS) and dithiothreitol (DTT). The analysis of fragmentation spectra based on the human database allowed for the identification of 254 human proteins present in the deposits. In addition, the use of multi-round search in the PEAKS Studio program against the bacterial base allowed for the identification of 393 proteins of bacterial origin present in the extract obtained from sialolith, which so far has not been carried out for this biological material. Furthermore, we successfully applied the SWATH methodology, allowing for a relative quantitative analysis of human proteins present in deposits. The obtained results correlate with the classification of sialoliths proposed by Tretiakow. The performed functional analysis allowed for the first time the selection of proteins, the levels of which differ between the tested samples, which may suggest the role of these proteins in the calcification process in different types of sialoliths. These are preliminary studies, and drawing specific conclusions requires research on a larger group, but it provides us the basis for the continuation of the work that has already begun.<\/jats:p>","DOI":"10.3390\/molecules26216725","type":"journal-article","created":{"date-parts":[[2021,11,7]],"date-time":"2021-11-07T20:41:14Z","timestamp":1636317674000},"page":"6725","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":11,"title":["Trial Proteomic Qualitative and Quantitative Analysis of the Protein Matrix of Submandibular Sialoliths"],"prefix":"10.3390","volume":"26","author":[{"given":"Paulina","family":"Czaplewska","sequence":"first","affiliation":[{"name":"Intercollegiate Faculty of Biotechnology, University of Gdansk, 80-307 Gdansk, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7750-7585","authenticated-orcid":false,"given":"Aleksandra E.","family":"Bogucka","sequence":"additional","affiliation":[{"name":"Intercollegiate Faculty of Biotechnology, University of Gdansk, 80-307 Gdansk, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9883-7965","authenticated-orcid":false,"given":"Natalia","family":"Musia\u0142","sequence":"additional","affiliation":[{"name":"Intercollegiate Faculty of Biotechnology, University of Gdansk, 80-307 Gdansk, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9251-4006","authenticated-orcid":false,"given":"Dmitry","family":"Tretiakow","sequence":"additional","affiliation":[{"name":"Department of Otolaryngology, Medical University of Gdansk, 80-214 Gdansk, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0042-6314","authenticated-orcid":false,"given":"Andrzej","family":"Skorek","sequence":"additional","affiliation":[{"name":"Department of Otolaryngology, Medical University of Gdansk, 80-214 Gdansk, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dominik","family":"Stodulski","sequence":"additional","affiliation":[{"name":"Department of Otolaryngology, Medical University of Gdansk, 80-214 Gdansk, Poland"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2021,11,6]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"329","DOI":"10.1006\/abbi.1994.1450","article-title":"Kappa-Casein and beta-Caseins in Human-Milk Micelles: Structural Studies","volume":"314","author":"Dev","year":"1994","journal-title":"Arch. Biochem. Biophys."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"374","DOI":"10.1111\/j.1399-3011.1989.tb00705.x","article-title":"Complete primary structure of statherin, a potent inhibitor of calcium phosphate precipitation, from the saliva of the monkey, Macaca arctoides","volume":"34","author":"Schlesinger","year":"2009","journal-title":"Int. J. Pept. Protein Res."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"951","DOI":"10.1001\/archotol.129.9.951","article-title":"Sialolithiasis Management: The state of the art","volume":"129","author":"Marchal","year":"2003","journal-title":"Arch. Otolaryngol. Head Neck Surg."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"1879","DOI":"10.1002\/lary.25377","article-title":"Nearly 3000 salivary stones: Some clinical and epidemiologic aspects","volume":"125","author":"Sigismund","year":"2015","journal-title":"Laryngoscope"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"E23","DOI":"10.1038\/sj.bdj.2014.1054","article-title":"Salivary stones: Symptoms, aetiology, biochemical composition and treatment","volume":"217","author":"Kraaij","year":"2014","journal-title":"Br. Dent. J."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"2089","DOI":"10.1007\/s00405-013-2463-7","article-title":"Algorithm changes in treatment of submandibular gland sialolithiasis","volume":"270","author":"Wierzbicka","year":"2013","journal-title":"Eur. Arch. Otorhinolaryngol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"983","DOI":"10.1111\/joor.12711","article-title":"An experimental chronic obstructive sialadenitis model by partial ligation of the submandibular duct characterised by sialography, histology, and transmission electron microscopy","volume":"45","author":"Li","year":"2018","journal-title":"J. Oral Rehabil."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"79","DOI":"10.14748\/bmr.v9.138","article-title":"Immunoregulation by the salivary glands","volume":"9","author":"Sabbadini","year":"1998","journal-title":"Biomed. Rev."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"764","DOI":"10.1016\/j.ijom.2016.02.007","article-title":"Association of serum electrolytes and smoking with salivary gland stone formation","volume":"45","author":"Yiu","year":"2016","journal-title":"Int. J. Oral Maxillofac. Surg."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"530","DOI":"10.1046\/j.1365-2559.1998.00432.x","article-title":"Microlithiasis in parotid sialadenosis and chronic submandibular sialadenitis is related to the microenvironment: An ultrastructural and microanalytical investigation","volume":"32","author":"Triantafyllou","year":"1998","journal-title":"Histopathology"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"935","DOI":"10.1177\/0194599811415489","article-title":"Etiologic Factors in Sialolithiasis","volume":"145","author":"Huoh","year":"2011","journal-title":"Otolaryngol. Neck Surg."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"2095","DOI":"10.1039\/c2an15924d","article-title":"Analytical investigation of salivary calculi, by mid-infrared spectroscopy","volume":"137","author":"Sabot","year":"2012","journal-title":"Analyst"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"1709","DOI":"10.1007\/s00784-012-0870-6","article-title":"Proteomic and scanning electron microscopic analysis of submandibular sialoliths","volume":"17","author":"Szalma","year":"2012","journal-title":"Clin. Oral Investig."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"299","DOI":"10.1016\/j.jds.2016.02.007","article-title":"Biochemical structure, symptoms, location and treatment of sialoliths","volume":"11","author":"Stelmach","year":"2016","journal-title":"J. Dent. Sci."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"219","DOI":"10.1080\/01913123.2020.1744784","article-title":"Ultrastructural analysis of the submandibular sialoliths: Raman spectroscopy and electron back-scatter studies","volume":"44","author":"Tretiakow","year":"2020","journal-title":"Ultrastruct. Pathol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1711","DOI":"10.1111\/odi.13708","article-title":"Classification of submandibular salivary stones based on ultrastructural studies","volume":"27","author":"Tretiakow","year":"2020","journal-title":"Oral Dis."},{"key":"ref_17","first-page":"109","article-title":"Salivary calculi microbiota: New insights into microbial networks and pathogens reservoir","volume":"21","author":"Capaccio","year":"2018","journal-title":"Microbes Infect."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"51","DOI":"10.1353\/hpu.0.0257","article-title":"Cystatin Superfamily","volume":"21","author":"Ochieng","year":"2010","journal-title":"J. Health Care Poor Underserved"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"234","DOI":"10.1111\/j.1601-0825.1999.tb00307.x","article-title":"Functional aspects of the human salivary cystatins in the oral environment","volume":"5","author":"Baron","year":"2008","journal-title":"Oral Dis."},{"key":"ref_20","first-page":"82","article-title":"Influence of point mutations on the stability, dimerization, and oligomerization of human cystatin C and its L68Q variant","volume":"5","author":"Jankowska","year":"2012","journal-title":"Front. Mol. Neurosci."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"513","DOI":"10.1002\/jmr.2220","article-title":"Interaction of serum amyloid A with human cystatin C-identification of binding sites","volume":"25","author":"Spodzieja","year":"2012","journal-title":"J. Mol. Recognit."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"415","DOI":"10.1002\/jmr.2283","article-title":"Interaction of serum amyloid A with human cystatin C-assessment of amino acid residues crucial for hCC-SAA formation (part II)","volume":"26","author":"Spodzieja","year":"2013","journal-title":"J. Mol. Recognit."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"e2581","DOI":"10.1002\/jmr.2581","article-title":"Characteristics of C-terminal, \u03b2-amyloid peptide binding fragment of neuroprotective protease inhibitor, cystatin C","volume":"30","author":"Spodzieja","year":"2016","journal-title":"J. Mol. Recognit."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"70","DOI":"10.1016\/j.archoralbio.2017.09.037","article-title":"Reactivation of peroxidase activity in human saliva samples by polyphenols","volume":"85","author":"Gau","year":"2018","journal-title":"Arch. Oral Biol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1364","DOI":"10.1128\/IAI.00825-12","article-title":"Host Defense Proteins Derived from Human Saliva Bind to Staphylococcus aureus","volume":"81","author":"Heo","year":"2013","journal-title":"Infect. Immun."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"647240","DOI":"10.1100\/2012\/647240","article-title":"Proteomic Analysis of Saliva Identifies Potential Biomarkers for Orthodontic Tooth Movement","volume":"2012","author":"Ellias","year":"2012","journal-title":"Sci. World J."},{"key":"ref_27","first-page":"73","article-title":"Detection of potential markers for systemic disease in saliva of pigs by proteomics: A pilot study","volume":"151","author":"Soler","year":"2012","journal-title":"Vet. Immunol. Immunopathol."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"857","DOI":"10.1177\/00220345840630060901","article-title":"Basic Biological Sciences Relationship between Concentration of Human Salivary Statherin and Inhibition of Calcium Phosphate Precipitation in Stimulated Human Parotid Saliva","volume":"63","author":"Hay","year":"1984","journal-title":"J. Dent. Res."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"268","DOI":"10.1177\/0022034510388653","article-title":"Histatin 1 Resists Proteolytic Degradation when Adsorbed to Hydroxyapatite","volume":"90","author":"McDonald","year":"2010","journal-title":"J. Dent. Res."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"403","DOI":"10.1159\/000094286","article-title":"Statherin and Histatin 1 Reduce Parotid Saliva-Promoted Streptococcus mutans Strain MT8148 Adhesion to Hydroxyapatite Surfaces","volume":"40","author":"Shimotoyodome","year":"2006","journal-title":"Caries Res."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"14","DOI":"10.1186\/s13075-017-1228-x","article-title":"Identification of potential saliva and tear biomarkers in primary Sj\u00f6gren\u2019s syndrome, utilising the extraction of extracellular vesicles and proteomics analysis","volume":"19","author":"Aqrawi","year":"2017","journal-title":"Arthritis Res."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1371","DOI":"10.1021\/pr060520e","article-title":"Proteomic Analysis of Salivary Acidic Proline-Rich Proteins in Human Preterm and At-Term Newborns","volume":"6","author":"Inzitari","year":"2007","journal-title":"J. Proteome Res."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1007\/BF00223503","article-title":"Salivary proline-rich proteins","volume":"45","author":"Bennick","year":"1982","journal-title":"Mol. Cell. Biochem."},{"key":"ref_34","unstructured":"Taylor, T.A., and Unakal, C.G. (2021, May 06). Staphylococcus Aureus, StatPearls, Available online: http:\/\/www.ncbi.nlm.nih.gov\/books\/NBK441868\/."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"787","DOI":"10.1515\/pjvs-2015-0102","article-title":"Biofilm-formation by Staphylococcus aureus and Staphylococcus epidermidis isolates from subclinical mastitis in conditions mimicking the udder environment","volume":"18","author":"Seixas","year":"2015","journal-title":"Pol. J. Vet. Sci."},{"key":"ref_36","unstructured":"(2021, May 06). Staphylococcus aureus and Staphylococcus epidermidis Virulence Strains as Causative Agents of Persistent Infections in Breast Implants, Available online: https:\/\/www.ncbi.nlm.nih.gov\/pmc\/articles\/PMC4727902\/."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"4592","DOI":"10.1128\/JCM.43.9.4592-4601.2005","article-title":"Phenotypic and Genotypic Characteristics of Streptococcus porcinus Isolated from Human Sources","volume":"43","author":"Duarte","year":"2005","journal-title":"J. Clin. Microbiol."},{"key":"ref_38","first-page":"4135246","article-title":"Streptococcus pseudoporcinus: Case Reports and Review of the Literature","volume":"2020","author":"Khan","year":"2020","journal-title":"Case Rep. Infect. Dis."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"2910","DOI":"10.3389\/fmicb.2019.02910","article-title":"Quantification of Human Oral and Fecal Streptococcus parasanguinis by Use of Quantitative Real-Time PCR Targeting the groEL Gene","volume":"10","author":"Chen","year":"2019","journal-title":"Front. Microbiol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"3199","DOI":"10.3389\/fmicb.2018.03199","article-title":"Streptococcus infantis, Streptococcus mitis, and Streptococcus oralis Strains with Highly Similar cps5 Loci and Antigenic Relatedness to Serotype 5 Pneumococci","volume":"9","author":"Pimenta","year":"2019","journal-title":"Front. Microbiol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"915","DOI":"10.2217\/fmb-2018-0043","article-title":"Streptococcus sanguinis biofilm formation & interaction with oral pathogens","volume":"13","author":"Zhu","year":"2018","journal-title":"Future Microbiol."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"2949","DOI":"10.1038\/s41598-017-02399-4","article-title":"Distinct Biological Potential of Streptococcus gordonii and Streptococcus sanguinis Revealed by Comparative Genome Analysis","volume":"7","author":"Zheng","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"745","DOI":"10.1038\/s41579-018-0089-x","article-title":"The oral microbiota: Dynamic communities and host interactions","volume":"16","author":"Lamont","year":"2018","journal-title":"Nat. Rev. Genet."},{"key":"ref_44","unstructured":"(2021, May 06). Clinical, Phenotypic, and Genotypic Evidence for Streptococcus sinensis as the Common Ancestor of Anginosus and Mitis Groups of Streptococci\u2014ScienceDirect. Available online: https:\/\/www.sciencedirect.com\/science\/article\/pii\/S0306987705004433?via%3Dihub."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"3591","DOI":"10.1128\/JCM.01481-12","article-title":"Evaluation of Methods for Identification and Determination of the Taxonomic Status of Strains Belonging to the Streptococcus porcinus-Streptococcus pseudoporcinus Complex Isolated from Animal, Human, and Dairy Sources","volume":"50","author":"Shewmaker","year":"2012","journal-title":"J. Clin. Microbiol."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"3041","DOI":"10.1128\/JCM.00802-09","article-title":"Thumb Infection Caused by Streptococcus pseudoporcinus","volume":"47","author":"Mahlen","year":"2009","journal-title":"J. Clin. Microbiol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"280","DOI":"10.5582\/irdr.2018.01110","article-title":"A case of leg cellulitis caused by multidrug-resistant Streptococcus pseudoporcinus","volume":"7","author":"Sawamura","year":"2018","journal-title":"Intractable Rare Dis. Res."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"e01985-19","DOI":"10.1128\/mBio.01985-19","article-title":"Identification of Virulence-Associated Properties by Comparative Genome Analysis of Streptococcus pneumoniae, S. pseudopneumoniae, S. mitis, Three, S. oralis Subspecies, and S. infantis","volume":"10","author":"Kilian","year":"2019","journal-title":"mBio"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"923","DOI":"10.1128\/JCM.44.3.923-927.2006","article-title":"Characteristics of Streptococcus pseudopneumoniae Isolated from Purulent Sputum Samples","volume":"44","author":"Keith","year":"2006","journal-title":"J. Clin. Microbiol."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"1570","DOI":"10.1099\/mic.0.056077-0","article-title":"A genomic approach to bacterial taxonomy: An examination and proposed reclassification of species within the genus Neisseria","volume":"158","author":"Bennett","year":"2012","journal-title":"Microbiology"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"e000019","DOI":"10.1099\/acmi.0.000019","article-title":"Phylogenetic insights into the diversity of Chryseobacterium species","volume":"1","author":"Siddaramappa","year":"2019","journal-title":"Access Microbiol."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"47","DOI":"10.1186\/s13099-016-0130-4","article-title":"Whole genome sequencing uncovers a novel IND-16 metallo-\u03b2-lactamase from an extensively drug-resistant Chryseobacterium indologenes strain J31","volume":"8","author":"Wang","year":"2016","journal-title":"Gut Pathog."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"12","DOI":"10.1186\/s12014-020-09275-w","article-title":"A comprehensive analysis of sialolith proteins and the clinical implications","volume":"17","author":"Busso","year":"2020","journal-title":"Clin. Proteom."},{"key":"ref_54","unstructured":"(2020, November 06). AZGP1 Is Androgen Responsive and Involved in AR-Induced Prostate Cancer Cell Proliferation and Metastasis\u2014PubMed, Available online: https:\/\/pubmed.ncbi.nlm.nih.gov\/30820960\/."},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"5674","DOI":"10.1021\/pr900748n","article-title":"Combination of FASP and StageTip-Based Fractionation Allows In-Depth Analysis of the Hippocampal Membrane Proteome","volume":"8","author":"Zougman","year":"2009","journal-title":"J. Proteome Res."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"1896","DOI":"10.1038\/nprot.2007.261","article-title":"Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips","volume":"2","author":"Rappsilber","year":"2007","journal-title":"Nat. Protoc."},{"key":"ref_57","doi-asserted-by":"crossref","unstructured":"Lewandowska, A., Fel, A., Thiel, M., Czaplewska, P., \u0141ukaszuk, K., Wi\u015bniewski, J., and O\u0142dziej, S. (2021). Compatibility of Distinct Label-Free Proteomic Workflows in Absolute Quantification of Proteins Linked to the Oocyte Quality in Human Follicular Fluid. Int. J. Mol. Sci., 22.","DOI":"10.3390\/ijms22147415"},{"key":"ref_58","doi-asserted-by":"crossref","unstructured":"(2021, April 30). PRIDE Database and Related Tools and Resources in 2019: Improving Support for Quantification Data|Nucleic Acids Research|Oxford Academic. Available online: https:\/\/academic.oup.com\/nar\/article\/47\/D1\/D442\/5160986.","DOI":"10.1093\/nar\/gky1106"},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"2498","DOI":"10.1101\/gr.1239303","article-title":"Cytoscape: A Software Environment for Integrated Models of Biomolecular Interaction Networks","volume":"13","author":"Shannon","year":"2003","journal-title":"Genome Res."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"D607","DOI":"10.1093\/nar\/gky1131","article-title":"STRING v11: Protein\u2013protein association networks with increased coverage, supporting functional discovery in genome-wide experimental datasets","volume":"47","author":"Szklarczyk","year":"2018","journal-title":"Nucleic Acids Res."}],"container-title":["Molecules"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1420-3049\/26\/21\/6725\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:26:51Z","timestamp":1760167611000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1420-3049\/26\/21\/6725"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,11,6]]},"references-count":60,"journal-issue":{"issue":"21","published-online":{"date-parts":[[2021,11]]}},"alternative-id":["molecules26216725"],"URL":"https:\/\/doi.org\/10.3390\/molecules26216725","relation":{},"ISSN":["1420-3049"],"issn-type":[{"value":"1420-3049","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,11,6]]}}}