Salivary stones, also known as sialoliths, are formed in a pathological situation in the salivary glands. So far, neither the mechanism of their formation nor the factors predisposing to their formation are known despite several hypotheses. While they do not directly threaten human life, they significantly deteriorate the patient’s quality of life. Although this is not a typical research material, attempts are made to apply various analytical tools to characterise sialoliths and search for the biomarkers in their proteomes. In this work, we used mass spectrometry and SWATH-MS qualitative and quantitative analysis to investigate the composition and select proteins that may contribute to solid deposits in the salivary glands. Twenty sialoliths, previously characterized spectroscopically and divided into the following groups: calcified (CAL), lipid (LIP) and mixed (MIX), were used for the study. Proteins unique for each of the groups were found, including: for the CAL group among them, e.g. proteins from the S100 group (S100 A8/A12 and P), mucin 7 (MUC7), keratins (KRT1/2/4/5/13), elastase (ELANE) or stomatin (STOM); proteins for the LIP group—transthyretin (TTR), lactotransferrin (LTF), matrix Gla protein (MPG), submandibular gland androgen-regulated protein 3 (SMR3A); mixed stones had the fewest unique proteins. Bacterial proteins present in sialoliths have also been identified. The analysis of the results indicates the possible role of bacterial infections, disturbances in calcium metabolism and neutrophil extracellular traps (NETs) in the formation of sialoliths.

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.

Salivary gland tumors are highly variable in clinical presentation and histology. The World Health Organization (WHO) classifies 22 types of malignant and 11 types of benign tumors of the salivary glands. Diagnosis of salivary gland tumors is based on imaging (ultrasound, magnetic resonance imaging) and fine-needle aspiration biopsy, but the final diagnosis is based on histopathological examination of the removed tumor tissue. In this pilot study, we are testing a new approach to identifying peptide biomarkers in saliva that can be used to diagnose salivary gland tumors. The research material for the peptidomic studies was extracts from washings of neoplastic tissues and healthy tissues (control samples). At the same time, saliva samples from patients and healthy individuals were analyzed. The comparison of the peptidome composition of tissue extracts and saliva samples may allow the identification of potential peptide markers of salivary gland tumors in patients’ saliva. The peptidome compositions extracted from 18 tumor and 18 healthy tissue samples, patients’ saliva samples (11 samples), and healthy saliva samples (8 samples) were analyzed by LC-MS tandem mass spectrometry. A group of 109 peptides was identified that were present only in the tumor tissue extracts and in the patients’ saliva samples. Some of the identified peptides were derived from proteins previously suggested as potential biomarkers of salivary gland tumors (ANXA1, BPIFA2, FGB, GAPDH, HSPB1, IGHG1, VIM) or tumors of other tissues or organs (SERPINA1, APOA2, CSTB, GSTP1, S100A8, S100A9, TPI1). Unfortunately, none of the identified peptides were present in all samples analyzed. This may be due to the high heterogeneity of this type of cancer. The surprising result was that extracts from tumor tissue did not contain peptides derived from salivary gland-specific proteins (STATH, SMR3B, HTN1, HTN3). These results could suggest that the developing tumor suppresses the production of proteins that are essential components of saliva.

Salivary gland tumors are highly variable in clinical presentation and histology. The World Health Organization (WHO) classifies 22 types of malignant and 11 types of benign tumors of the salivary glands. Diagnosis of salivary gland tumors is based on imaging (ultrasound, magnetic resonance imaging) and fine-needle aspiration biopsy, but the final diagnosis is based on histopathological examination of the removed tumor. In this pilot study, we are testing a new approach to identifying peptide biomarkers in saliva that can be used to diagnose salivary gland tumors. The research material for the peptidomic studies was extracts from washings of neoplastic tissues and healthy tissues (control samples). At the same time, saliva samples from patients and healthy individuals were analysed. The comparison of the peptidome composition of tissue extracts and saliva samples may allow the identification of potential peptide markers of salivary gland tumors in patients' saliva. The peptidome composition extracted from 18 tumor and 18 healthy tissue samples, patient (11 samples) and healthy saliva samples (8 samples) was analysed by LC-MS tandem mass spectrometry. A group of 109 peptides were identified that were present only in the tumor tissue extracts and in the patient saliva samples. Some of the identified peptides were derived from proteins previously suggested as potential biomarkers of salivary gland tumors (ANXA1, BPIFA2, FGB, GAPDH, HSPB1, IGHG1, VIM) or tumors of other tissues or organs (SERPINA1, APOA2, CSTB, GSTP1, S100A8, S100A9, TPI1). Unfortunately, none of the identified peptides were present in all samples analysed. This may be due to the high heterogeneity of this type of cancer. The surprising result was that extracts from tumor tissue did not contain peptides derived from salivary gland-specific proteins (STATH, SMR3B, HTN1, HTN3). These results could suggest that the developing tumor suppresses the production of proteins that are essential components of saliva.

Wstęp: Celem pracy było zbadanie kwestii, czy kierunek studiów i styl życia wpływają na równowagę u młodych dorosłych. Materiał i metody: Badaniem objęto 62 studentów fizjoterapii i systemów diagnostycznych w medycynie w wieku od 20 do 24 lat. Przeprowadzone zostało za pomocą autorskiej ankiety. Dodatkowo osoby zostały przebadane systemem Zebris pod kątem oceny równowagi. Uzyskane wyniki poddane zostały analizie statystycznej. Wyniki: Wykazano zależność pomiędzy wzrostem, wagą, płcią badanych oraz stylem życia a parametrami równowagi. Czynniki, takie jak masa ciała, wiek i aktywność fizyczna, wykazały również wpływ na rozkład symetrii nacisku kończyn dolnych u grupy badanej. Zaobserwowano statystycznie istotną średnią korelację ujemną wieku i całkowitego obciążenia kończyny dolnej prawej w grupie kobiet (R=-0,373; p=0,015). Zaobserwowano również statystycznie istotną średnią korelację dodatnią (p=0,029) wartości COP HD (Horizontal Deviation) w grupie kobiet w zależności od kierunku studiów. Wnioski: Najistotniejszymi czynnikami wpływającymi na parametry równowagi badanych jest wzrost, wiek oraz niektóre składowe stylu życia. Z kolei na rozkład sił nacisku i symetrię obciążenia kończyn dolnych wpływa przede wszystkim masa ciała mężczyzn, wiek kobiet oraz częstość palenia papierów i jakość snu.

Background: Salivary and kidney stone diseases are associated with pathological calcification, yet their molecular composition remains incompletely characterized. Proteomic analysis of saliva and stone material may provide valuable insights into these processes; however, such analyses are technically challenging and strongly dependent on efficient and reproducible sample preparation workflows. Methods: In this study, we systematically evaluated and optimised protein extraction and digestion conditions for proteomic analysis of saliva, salivary stones and kidney stones. Different lysis buffers, sample amounts, sonication parameters and digestion strategies were tested. Additional biological materials, including salivary gland tissue and Raoultella ornithinolytica , were analysed to assess the broader applicability of the developed workflows. Results: Optimised workflows significantly improved peptide and protein identification, reduced the proportion of missed cleavages and enhanced sequence coverage across multiple biological matrices. Sonication-assisted protocols proved particularly effective for stone-derived materials, although optimal conditions varied depending on the sample type. Conclusion: This pilot study demonstrates that careful optimisation of protein extraction and digestion is essential for reliable proteomic analysis of challenging biomaterials such as saliva and pathological calcifications. The workflows established here provide a robust methodological foundation for future large-scale and biomarker-oriented proteomic studies of stone disease and related calcification disorders.