Top-down HPLC-ESI–MS proteomic analysis of saliva of edentulous subjects evidenced high levels of cystatin A, cystatin B and SPRR3

Objective This study aims to analyze the salivary peptidome/proteome of edentulous subject with respect to dentate control subjects. Design Unstimulated whole saliva, collected from 11 edentulous subjects (age 60–76 years) and 11 dentate age-matched control subjects, was immediately treated with 0.2% aqueous trifluoroacetic acid and the acidic soluble fraction analyzed by High Performace Liquid Chromatography-Mass Spectrometry. The relative abundance of the salivary peptides/proteins was determined by measuring the area of the High Performace Liquid Chromatography-Mass Spectrometry eXtracted Ion Current peaks which is linearly proportional to peptide/protein concentration under identical experimental conditions. Levels of salivary peptides/proteins in the two groups were compared by the nonparametric Mann-Whitney test to evidence statistically significant differences. Results Levels of cystatin A, S-glutathionylated, S-cystenylated, S-S dimer derivatives of cystatin B and S-glutathionylated derivative of SPRR3, were found significantly higher in edentulous subjects with respect to dentate controls. The major peptides and proteins typically deriving from salivary glands did not show any statistically significant differences. Conclusions Cystatin A, S-glutathionylated, S-cystenylated, S-S dimer derivatives of cystatin B and S-glutathionylated derivative of SPRR3, which are mainly of intracellular origin and represent the major constituents of the cornified cell envelope are a clue of inflammation of mucosal epithelia

Top-down proteomic profiling of human saliva in multiple sclerosis patients

Multiple sclerosis is a chronic disease of the central nervous system characterized by inflammation, demyelination and neurodegeneration which is of undetermined origin. To date a single diagnostic test of multiple sclerosis does not exists and novel biomarkers are demanded for a more accurate and early diagnosis. In this study, we performed the quantitative analysis of 119 salivary peptides/proteins from 49 multiple sclerosis patients and 54 healthy controls by a mass spectrometry-based top-down proteomic approach. Statistical analysis evidenced different levels on 23 proteins: 8 proteins showed lower levels in multiple sclerosis patients with respect to controls and they were mono- and di-oxidized cystatin SN, mono- and di-oxidized cystatin S1, mono-oxidized cystatin SA and mono-phosphorylated statherin. 15 proteins showed higher levels in multiple sclerosis patients with respect to controls and they were antileukoproteinase, two proteoforms of Prolactin-Inducible Protein, P-C peptide (Fr.1-14, Fr. 26-44, and Fr. 36-44), SV1 fragment of statherin, cystatin SN Des1-4, cystatin SN P11 → L variant, and cystatin A T96 → M variant. The differences observed between the salivary proteomic profile of patients suffering from multiple sclerosis and healthy subjects is consistent with the inflammatory condition and altered immune response typical of the pathology. Data are available via ProteomeXchange with identifier PXD009440

Salivary Cystatins: Exploring New Post-Translational Modifications and Polymorphisms by Top-Down High-Resolution Mass Spectrometry

Cystatins are a complex family of cysteine peptidase inhibitors. In the present study, various proteoforms of cystatin A, cystatin B, cystatin S, cystatin SN, and cystatin SA were detected in the acid-soluble fraction of human saliva and characterized by a top-down HPLC–ESI–MS approach. Proteoforms of cystatin D were also detected and characterized by an integrated top-down and bottom-up strategy. The proteoforms derive from coding sequence polymorphisms and post-translational modifications, in particular, phosphorylation, N-terminal processing, and oxidation. This study increases the current knowledge of salivary cystatin proteoforms and provides the basis to evaluate possible qualitative/quantitative variations of these proteoforms in different pathological states and reveal new potential salivary biomarkers of disease. Data are available via ProteomeXchange with identifier PXD007170

Extensive characterization of the human salivary basic proline-rich protein family by top-down mass spectrometry

Human basic proline-rich proteins and basic glycosylated proline-rich proteins, encoded by the polymorphic PRB1-4 genes and expressed only in parotid glands, are the most complex family of adult salivary proteins. The family includes 11 parent peptides/proteins and more than 6 parent glycosylated proteins, but a high number of proteoforms with rather similar structures derive from polymorphisms and post-translational modifications. 55 new components of the family were characterized by top-down liquid chromatography-mass spectrometry and tandem-mass platforms, bringing the total number of proteoforms to 109. The new components comprise the three variants P-H S1→ A, P-Ko P36→ S, and P-Ko A41â†' S and several of their naturally occurring proteolytic fragments. The paper represents an updated reference for the peptides included in the heterogeneous family of proteins encoded by PRB1/PRB4. MS data are available via ProteomeXchange with the identifier PXD009813

Monitoring neonatal fungal infection with metabolomics

The objective of our study was to evaluate the capability of the metabolomics approach to identify the variations of urine metabolites over time related to the neonatal fungal septic condition. The study population included a clinical case of a preterm neonate with invasive fungal infection and 13 healthy preterm controls. This study showed a unique urine metabolic profile of the patient affected by fungal sepsis compared to urine of controls and it was also possible to evaluate the efficacy of therapy in improving patient health

PCA (A) and PLS-DA (B) of plasma analysis from patients with FMS vs controls.

<p>PLSA-DA values were: R²X = 0,345 R²Y = 0.901 Q² = 0,806 p-value = 1.1×10−12.</p

Binding free energies for the three ligand-receptor complexes after docking and after MD simulations.

<p>RMSD value in the last column refers to the average value of ligand during the whole MD simulation time.</p><p>Binding free energies for the three ligand-receptor complexes after docking and after MD simulations.</p

Demographic characteristics and clinical parameters of FMS and Controls.

<p>* Amitriptyline <15 mg/die).</p><p>** Selective Serotonin Reuptake Inhibitors (Citalopram 20 mg/die).</p><p>***Selective Serotonin Noradrenaline Reuptake Inhibitors (Duloxetin 30 mg/die).</p><p>Demographic characteristics and clinical parameters of FMS and Controls.</p

Best binding pose obtained for the ligands (i) PC(14∶0/0∶0) in green and (ii) PC(16∶0/0∶0) in red inside PAF receptor.

<p>The receptor is shown using cartoon representation with helix 3 in pink and helix 6 in orange. The ligands are shown using ball and stick representation.</p