Simultaneous quantification of selected glycosaminoglycans by butanolysis-based derivatization and LC-SRM/MS analysis for assessing glycocalyx disruption in vitro and in vivo

2022
journal article
article
4
dc.abstract.enGlycosaminoglycans (GAGs) constitute the main building blocks of the endothelial glycocalyx (GLX), and disruption of GLX initiates and promotes endothelial dysfunction. Here, we aimed to develop a novel, specific and accurate LC-SRM/MS-based method for glycosaminoglycans (GAGs) profiling. The method involved butanolysis derivatization to facilitate GAG-specific disaccharide generation and its subsequent retention in LC–reversed-phase mode followed by mass spectrometric detection performed in positive ion-selected reaction monitoring (SRM) mode. GAG contents were measured in media of endothelial cells (EA.hy926) subjected to various GAG-degrading enzymes, as well as in murine plasma and urine in apolipoprotein E/low‐density lipoprotein receptor‐deficient (ApoE/LDLR −/−) mice and age-matched wild-type C57BL/6 mice. Alternatively, GLX disruption was verified by atomic force microscopy (AFM)-based analysis of GLX thickness. The proposed assay to quantify GAG-specific disaccharides presented high sensitivity for each of the analytes (LLOQ: 0.05–0.1 μg/mL) as well as accuracy and precision (86.8–114.9% and 2.0–14.3%, respectively). In medium of EA.hy926 cells subjected to GAG-degrading enzymes various GAG-specific disaccharides indicating the degradation of keratan sulphate (KS), heparan sulphate (HS), chondroitin sulphate (CHS) or hyaluronan (HA) were detected as predicted based on the characteristics of individual enzyme activity. In turn, AFM-based assessment of GLX thickness was reduced to a similar extent by all single enzyme treatments, whereas the most prominent reduction of GLX thickness was detected following the enzyme mixture. Plasma measurements of GAGs revealed age- and hypercholesterolemia-dependent decrease in GAGs concentration. In summary, a novel LC-SRM/MS-based method for GAG profiling was proposed that may inform on GLX status in cell culture for both in vitro and in vivo conditions.pl
dc.affiliationPion Prorektora ds. badań naukowych : Jagiellońskie Centrum Rozwoju Lekówpl
dc.affiliationWydział Lekarski : Zakład Farmakologiipl
dc.affiliationWydział Farmaceutyczny : Zakład Toksykologiipl
dc.cm.id106705
dc.cm.idOmegaUJCM8bfab1b5268f4317ba6ed2b9b84b1c15pl
dc.contributor.authorMatyjaszczyk-Gwarda, Karolina - 178201 pl
dc.contributor.authorKij, Agnieszka - 215016 pl
dc.contributor.authorOlkowicz, Mariola - 428620 pl
dc.contributor.authorFels, Benediktpl
dc.contributor.authorKusche-Vihrog, Kristinapl
dc.contributor.authorWalczak, Maria - 133728 pl
dc.contributor.authorChłopicki, Stefan - 128995 pl
dc.date.accession2022-02-15pl
dc.date.accessioned2022-02-01T11:25:41Z
dc.date.available2022-02-01T11:25:41Z
dc.date.issued2022pl
dc.date.openaccess0
dc.description.accesstimew momencie opublikowania
dc.description.numberPart 1pl
dc.description.versionostateczna wersja wydawcy
dc.description.volume238pl
dc.identifier.articleid123008pl
dc.identifier.doi10.1016/j.talanta.2021.123008pl
dc.identifier.eissn1873-3573pl
dc.identifier.issn0039-9140pl
dc.identifier.urihttps://ruj.uj.edu.pl/xmlui/handle/item/287521
dc.identifier.weblinkhttps://www.sciencedirect.com/science/article/pii/S0039914021009309?via%3Dihubpl
dc.languageengpl
dc.language.containerengpl
dc.pbn.affiliationDziedzina nauk medycznych i nauk o zdrowiu : nauki farmaceutyczne
dc.pbn.affiliationDziedzina nauk medycznych i nauk o zdrowiu : nauki medyczne
dc.rightsUdzielam licencji. Uznanie autorstwa 4.0 Międzynarodowa*
dc.rights.licenceCC-BY
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/legalcode.pl*
dc.share.typeOtwarte czasopismo
dc.subject.enGlycosaminoglycanspl
dc.subject.enGlycocalyxpl
dc.subject.enButanolysispl
dc.subject.enLiquid chromatography-tandem mass spectrometrypl
dc.subject.enEndotheliumpl
dc.subject.enAtherosclerosispl
dc.subtypeArticlepl
dc.titleSimultaneous quantification of selected glycosaminoglycans by butanolysis-based derivatization and LC-SRM/MS analysis for assessing glycocalyx disruption in vitro and in vivopl
dc.title.journalTalantapl
dc.typeJournalArticlepl
dspace.entity.typePublication
dc.abstract.enpl
Glycosaminoglycans (GAGs) constitute the main building blocks of the endothelial glycocalyx (GLX), and disruption of GLX initiates and promotes endothelial dysfunction. Here, we aimed to develop a novel, specific and accurate LC-SRM/MS-based method for glycosaminoglycans (GAGs) profiling. The method involved butanolysis derivatization to facilitate GAG-specific disaccharide generation and its subsequent retention in LC–reversed-phase mode followed by mass spectrometric detection performed in positive ion-selected reaction monitoring (SRM) mode. GAG contents were measured in media of endothelial cells (EA.hy926) subjected to various GAG-degrading enzymes, as well as in murine plasma and urine in apolipoprotein E/low‐density lipoprotein receptor‐deficient (ApoE/LDLR −/−) mice and age-matched wild-type C57BL/6 mice. Alternatively, GLX disruption was verified by atomic force microscopy (AFM)-based analysis of GLX thickness. The proposed assay to quantify GAG-specific disaccharides presented high sensitivity for each of the analytes (LLOQ: 0.05–0.1 μg/mL) as well as accuracy and precision (86.8–114.9% and 2.0–14.3%, respectively). In medium of EA.hy926 cells subjected to GAG-degrading enzymes various GAG-specific disaccharides indicating the degradation of keratan sulphate (KS), heparan sulphate (HS), chondroitin sulphate (CHS) or hyaluronan (HA) were detected as predicted based on the characteristics of individual enzyme activity. In turn, AFM-based assessment of GLX thickness was reduced to a similar extent by all single enzyme treatments, whereas the most prominent reduction of GLX thickness was detected following the enzyme mixture. Plasma measurements of GAGs revealed age- and hypercholesterolemia-dependent decrease in GAGs concentration. In summary, a novel LC-SRM/MS-based method for GAG profiling was proposed that may inform on GLX status in cell culture for both in vitro and in vivo conditions.
dc.affiliationpl
Pion Prorektora ds. badań naukowych : Jagiellońskie Centrum Rozwoju Leków
dc.affiliationpl
Wydział Lekarski : Zakład Farmakologii
dc.affiliationpl
Wydział Farmaceutyczny : Zakład Toksykologii
dc.cm.id
106705
dc.cm.idOmegapl
UJCM8bfab1b5268f4317ba6ed2b9b84b1c15
dc.contributor.authorpl
Matyjaszczyk-Gwarda, Karolina - 178201
dc.contributor.authorpl
Kij, Agnieszka - 215016
dc.contributor.authorpl
Olkowicz, Mariola - 428620
dc.contributor.authorpl
Fels, Benedikt
dc.contributor.authorpl
Kusche-Vihrog, Kristina
dc.contributor.authorpl
Walczak, Maria - 133728
dc.contributor.authorpl
Chłopicki, Stefan - 128995
dc.date.accessionpl
2022-02-15
dc.date.accessioned
2022-02-01T11:25:41Z
dc.date.available
2022-02-01T11:25:41Z
dc.date.issuedpl
2022
dc.date.openaccess
0
dc.description.accesstime
w momencie opublikowania
dc.description.numberpl
Part 1
dc.description.version
ostateczna wersja wydawcy
dc.description.volumepl
238
dc.identifier.articleidpl
123008
dc.identifier.doipl
10.1016/j.talanta.2021.123008
dc.identifier.eissnpl
1873-3573
dc.identifier.issnpl
0039-9140
dc.identifier.uri
https://ruj.uj.edu.pl/xmlui/handle/item/287521
dc.identifier.weblinkpl
https://www.sciencedirect.com/science/article/pii/S0039914021009309?via%3Dihub
dc.languagepl
eng
dc.language.containerpl
eng
dc.pbn.affiliation
Dziedzina nauk medycznych i nauk o zdrowiu : nauki farmaceutyczne
dc.pbn.affiliation
Dziedzina nauk medycznych i nauk o zdrowiu : nauki medyczne
dc.rights*
Udzielam licencji. Uznanie autorstwa 4.0 Międzynarodowa
dc.rights.licence
CC-BY
dc.rights.uri*
http://creativecommons.org/licenses/by/4.0/legalcode.pl
dc.share.type
Otwarte czasopismo
dc.subject.enpl
Glycosaminoglycans
dc.subject.enpl
Glycocalyx
dc.subject.enpl
Butanolysis
dc.subject.enpl
Liquid chromatography-tandem mass spectrometry
dc.subject.enpl
Endothelium
dc.subject.enpl
Atherosclerosis
dc.subtypepl
Article
dc.titlepl
Simultaneous quantification of selected glycosaminoglycans by butanolysis-based derivatization and LC-SRM/MS analysis for assessing glycocalyx disruption in vitro and in vivo
dc.title.journalpl
Talanta
dc.typepl
JournalArticle
dspace.entity.type
Publication
Affiliations

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