For the understanding of pathological states of bone tissues in oral surgery, it would be desirable to have the possibility to simulate these processes on bone cell models in vitro. These cultures, similarly to bone tissues, contain numerous proteins entrapped in the insoluble matrix. The major goal of this study was to verify whether a method based on direct in-matrix protein digestion could be suitable for the discrimination between different induced pathological states of bone cell models cultivated in vitro. Using in-sample specific protein digestion with trypsin followed by liquid chromatography-tandem mass spectrometry analysis of released peptides, 446 proteins (in average per sample) were identified in a bone cell in vitro model with induced cancer, 440 proteins were found in a model with induced inflammation, 451 proteins were detected in control in vitro culture, and 491 proteins were distinguished in samples of vestibular laminas of maxillary bone tissues originating from six different patients. Subsequent partial least squares - discrimination analysis of obtained liquid chromatography-tandem mass spectrometry data was able to discriminate among in vitro cultures with induced cancer, with induced inflammation, and control cultivation. Thus, the direct in-sample protein digestion by trypsin followed by liquid chromatography-tandem mass spectrometry analysis of released specific peptide fragments from the insoluble matrix and mathematical analysis of the mass spectrometry data seems to be a promising tool for the routine proteomic characterization of in vitro human bone models with induced different pathological states.
- MeSH
- chromatografie kapalinová metody MeSH
- lidé MeSH
- peptidy analýza MeSH
- proteiny chemie MeSH
- proteolýza MeSH
- proteomika metody MeSH
- stomatochirurgické výkony * MeSH
- tandemová hmotnostní spektrometrie * metody MeSH
- trypsin chemie MeSH
- zánět MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
At present, Alzheimer's disease is detected mainly using psychological tests, which can only confirm the disease in its more advanced phases. Therefore, bioanalytical possibilities for detecting this disease earlier are being investigated. To date, the results of analyses, which focus mainly on the study of lipids and proteins either in cerebrospinal fluid or much less often in blood plasma, do not provide satisfactory results. In addition, cerebrospinal fluid sampling is uncomfortable for the patients and involves many health risks. In this work, we deal with proteomic analysis using Matrix-Assisted Laser Desorption/Ionisation-Time of Flight and Liquid Chromatography coupled to tandem Mass Spectrometry of blood plasma with a focus on various ways of preanalytical sample treatments. This should lead to results improvement and facilitate the subsequent evaluation using principal component analysis and partial least squares discriminant analysis. The obtained results indicate the direction of further research, namely the study of interactions between proteins and lipids contained in blood plasma. These substances may be regarded as potential biomarkers allowing for the diagnosis of Alzheimer ́s disease even in its early stages.
- MeSH
- Alzheimerova nemoc * krev diagnóza MeSH
- biologické markery krev MeSH
- chromatografie kapalinová metody MeSH
- krevní plazma chemie MeSH
- krevní proteiny analýza MeSH
- lidé středního věku MeSH
- lidé MeSH
- lipidy krev MeSH
- metabolismus lipidů MeSH
- proteomika metody MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice metody MeSH
- tandemová hmotnostní spektrometrie metody MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- cytodiagnostika metody přístrojové vybavení MeSH
- hmotnostní spektrometrie metody přístrojové vybavení MeSH
- klinické laboratorní techniky MeSH
- monoklonální protilátky izolace a purifikace MeSH
- nádory mozku * diagnóza imunologie patologie MeSH
- regulace genové exprese u nádorů MeSH
- Publikační typ
- práce podpořená grantem MeSH
- MeSH
- biochemie * metody trendy MeSH
- chemické techniky analytické metody přístrojové vybavení trendy využití MeSH
- fluorescenční mikroskopie metody přístrojové vybavení MeSH
- publikování MeSH
- soudní lékařství * metody trendy MeSH
- spektrofotometrie infračervená metody přístrojové vybavení MeSH
- studium vysokoškolské MeSH
- výchova a vzdělávání * ekonomika organizace a řízení trendy MeSH
- Publikační typ
- práce podpořená grantem MeSH
Aortic stenosis is one of the most common heart diseases that occur in developed countries. The disease has many causes; among the most discussed is excessive intake of fluorides. The aim of our work is to clarify the biochemical nature of biomineralizations in cardial tissue as well as to select an appropriate medication and to mitigate the disease. Peptide mapping combined with UHPLC and MS were used in the study of the mineralized cardial tissue. We managed to identify proteins such as alkaline phosphatase, biglycan, mimecan, osteopontin, periostin and proteoglycan, which are probably related to mineralization of the valves.
- MeSH
- aortální stenóza * etiologie MeSH
- chromatografie kapalinová využití MeSH
- fluor škodlivé účinky MeSH
- hmotnostní spektrometrie * metody přístrojové vybavení využití MeSH
- kalcinóza * patofyziologie MeSH
- lidé MeSH
- peptidové mapování * metody trendy využití MeSH
- tandemová hmotnostní spektrometrie využití MeSH
- vitamin K chemie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
Proper identification of proteinaceous binders in artworks is essential for specification of the painting technique and thus also for selection of the restoration method; moreover, it might be helpful for the authentication of the artwork. This paper is concerned with the optimisation of analysis of the proteinaceous binders contained in the colour layers of artworks. Within this study, we worked out a method for the preparation and analysis of solid samples from artworks using tryptic cleavage and subsequent analysis of the acquired peptide mixture by matrix-assisted laser desorption/ionisation time of flight mass spectrometry. To make this approach rational and efficient, we created a database of commonly used binders (egg yolk, egg white, casein, milk, curd, whey, gelatine, and various types of animal glues); certain peaks in the mass spectra of these binders, formed by rich protein mixtures, were matched to amino acid sequences of the individual proteins that were found in the Internet database ExPASy; their cleavage was simulated by the program Mass-2.0-alpha4. The method developed was tested on model samples of ground layers prepared by an independent laboratory and then successfully applied to a real sample originating from a painting by Edvard Munch.
