Monitoring of in vitro bioavailability and uptake of glycosylated food allergens using cell-based models
Praćenje in vitro biousvojivosti i preuzimanja glikozilovanih alergena hrane upotrebom ćelijskih modela
Аутори
Krstić-Ristivojević, MajaОстала ауторства
Ćirković-Veličković, Tanjavan Hage, Marianne
Stojadinović, Marija M.
Jovanović, Vesna B.
Burazer, Lidija M.
Докторска теза (Објављена верзија)
Метаподаци
Приказ свих података о документуАпстракт
extensive engagement of researchers in the elucidation of the mechanismsunderlying processes of food digestion, allergen transport, and uptake by the immunecells and its effector immune responses. The development of the in vitro assays andcell-based models has allowed bridging the problem of food allergy research and therespect of the ethical norms in used research procedures.The red meat allergy is a novel type of food allergy characterized by theproduction of an IgE antibody against the carbohydrate galactose-α-1,3-galactose (α-Gal). Glycoproteins from non-primate mammals are rich with an α-Gal as posttranslationalmodification. Also, red meat allergy is characterized by the delayed onsetof symptoms which may be related to the mechanism and the fate of α-Gal carryingproteins in the human gastrointestinal tract. Furthermore, the uptake, processing, andmechanisms of presentation of α-Gal by the immune cells are still unknown. Thereforethis doctoral dissertation aimed to investigate how ...protein glycosylation by α-Galaffects their susceptibility to gastric digestion, does α-Gal conjugated to proteins affectstheir transport through the Caco-2 cell monolayer, which mimics the gastrointestinallayer, and to examine the influence of α-Gal epitopes on the protein surface on theiruptake and processing by immature monocyte-derived dendritic cells (iMDDCs). Thestudy revealed that the presence of the α-Gal glycosylation on protein surface had animpact on their susceptibility to gastric digestion and the digestion pattern of theobtained protein fragments upon pepsinolysis. Prolonged survival, up to 2h of digestion,was characteristic of the large proteins fragments bearing the α-Gal epitope.Importantly, transport through the Caco-2 monolayer of proteins conjugated to α-Galwas hampered in comparison to unconjugated proteins. Furthermore, differentialcentrifugation of Caco-2 cell lysates upon transport experiments revealed that α-Galcould be detected on the intact protein in the endosomal fraction of the cells...
Alergije na hranu su rastući problem u ljudskoj populaciji širom sveta irešavanje ovog problema zahteva opsežno angažovanje istraživača u rasvetljavanjumehanizama uključenih u procese varenja hrane, transporta alergena i njihovog unosaod strane imunih ćelija odgovornih za efektorske mehanizme imunoloških odgovora.Ovo je nezamislivo bez razvoja in vitro testova i ćelijskih modela koji premošćujuproblem istraživanja alergija na hranu uz poštovanja etičkih normi u korišćenimistraživačkim metodama.Novu vrstu alergije na hranu, alergiju na crveno meso, karakteriše sintezaimunoglobulina E kao odgovor na prisustvo šećera galaktoza-α-1,3-galaktoza (α-Gal),koji je prisutan na površini glikoproteina primata. Takođe, alergiju na crveno mesokarakteriše odložena pojava simptoma što može biti rezultat promena u mehanizmuobrade proteina koji nose α-Gal u gastrointestinalnom traktu čoveka. Dalje, unos,obrada i mehanizmi prezentacije α-Gal šećera od strane imunih ćelija još uvek nisupoznati. Stoga cilj...evi ove doktorske disertacije su ispitivanje kako α-Gal glikozilacijaproteina utiče na njihovu digestiju od strane pepsina, da li α-Gal glikozilacija proteinautiče na njihov transport kroz monosloj Caco-2 ćelija, koji oponaša gastrointestinalniepitel, kao i ispitivanje uticaja α-Gal glikozilacije na površini proteina na njihov unos iobradu od strane nezrelih dendritičnih ćelijama kultivisanih iz monocita (iMDDC). Izdobijenih rezultata moze se zaključiti da prisustvo α-Gal glikozilacije na površiniproteina utiče na njihovu podložnost na digestiju a najviše na obrazac dobijenihfragmenata proteina nakon pepsinolize. Veliki fragmenti proteina koji nose α-Galprisutni su čak i do 2 sata digestije. Takođe, važno je istaći da je transport α-Galglikozilovanih proteina kroz Caco-2 monosloj otežan u poređenju sa neglikozilovanimproteinima. Dalje, diferencijalnim centrifugiranjem lizata aco-2 ćelija nakontranscitoze pokazalo je da je α-Gal prisutan na intaktnim proteinima u endozomalnimfrakcijama ćelija...
Кључне речи:
α-Gal / bioavailability / transcytosis / glycoprotein / Caco-2 cells / mammalianmeat allergy / iMDDCs / uptake / cell-based models / a-Gal / biousvojivost transcitoza glikoprotein aco-2 ćelije alergija na crveno meso / iMDDCs / usvajanje / ćelijski modeliИзвор:
Универзитет у Београду, 2020Издавач:
- Универзитет у Београду, Хемијски факултет
URI
http://eteze.bg.ac.rs/application/showtheses?thesesId=7891https://fedorabg.bg.ac.rs/fedora/get/o:23228/bdef:Content/download
http://vbs.rs/scripts/cobiss?command=DISPLAY&base=70036&RID=30780681
https://nardus.mpn.gov.rs/handle/123456789/17803
https://cherry.chem.bg.ac.rs/handle/123456789/4362
Колекције
Институција/група
Hemijski fakultet / Faculty of ChemistryTY - THES AU - Krstić-Ristivojević, Maja PY - 2020 UR - http://eteze.bg.ac.rs/application/showtheses?thesesId=7891 UR - https://fedorabg.bg.ac.rs/fedora/get/o:23228/bdef:Content/download UR - http://vbs.rs/scripts/cobiss?command=DISPLAY&base=70036&RID=30780681 UR - https://nardus.mpn.gov.rs/handle/123456789/17803 UR - https://cherry.chem.bg.ac.rs/handle/123456789/4362 AB - extensive engagement of researchers in the elucidation of the mechanismsunderlying processes of food digestion, allergen transport, and uptake by the immunecells and its effector immune responses. The development of the in vitro assays andcell-based models has allowed bridging the problem of food allergy research and therespect of the ethical norms in used research procedures.The red meat allergy is a novel type of food allergy characterized by theproduction of an IgE antibody against the carbohydrate galactose-α-1,3-galactose (α-Gal). Glycoproteins from non-primate mammals are rich with an α-Gal as posttranslationalmodification. Also, red meat allergy is characterized by the delayed onsetof symptoms which may be related to the mechanism and the fate of α-Gal carryingproteins in the human gastrointestinal tract. Furthermore, the uptake, processing, andmechanisms of presentation of α-Gal by the immune cells are still unknown. Thereforethis doctoral dissertation aimed to investigate how protein glycosylation by α-Galaffects their susceptibility to gastric digestion, does α-Gal conjugated to proteins affectstheir transport through the Caco-2 cell monolayer, which mimics the gastrointestinallayer, and to examine the influence of α-Gal epitopes on the protein surface on theiruptake and processing by immature monocyte-derived dendritic cells (iMDDCs). Thestudy revealed that the presence of the α-Gal glycosylation on protein surface had animpact on their susceptibility to gastric digestion and the digestion pattern of theobtained protein fragments upon pepsinolysis. Prolonged survival, up to 2h of digestion,was characteristic of the large proteins fragments bearing the α-Gal epitope.Importantly, transport through the Caco-2 monolayer of proteins conjugated to α-Galwas hampered in comparison to unconjugated proteins. Furthermore, differentialcentrifugation of Caco-2 cell lysates upon transport experiments revealed that α-Galcould be detected on the intact protein in the endosomal fraction of the cells... AB - Alergije na hranu su rastući problem u ljudskoj populaciji širom sveta irešavanje ovog problema zahteva opsežno angažovanje istraživača u rasvetljavanjumehanizama uključenih u procese varenja hrane, transporta alergena i njihovog unosaod strane imunih ćelija odgovornih za efektorske mehanizme imunoloških odgovora.Ovo je nezamislivo bez razvoja in vitro testova i ćelijskih modela koji premošćujuproblem istraživanja alergija na hranu uz poštovanja etičkih normi u korišćenimistraživačkim metodama.Novu vrstu alergije na hranu, alergiju na crveno meso, karakteriše sintezaimunoglobulina E kao odgovor na prisustvo šećera galaktoza-α-1,3-galaktoza (α-Gal),koji je prisutan na površini glikoproteina primata. Takođe, alergiju na crveno mesokarakteriše odložena pojava simptoma što može biti rezultat promena u mehanizmuobrade proteina koji nose α-Gal u gastrointestinalnom traktu čoveka. Dalje, unos,obrada i mehanizmi prezentacije α-Gal šećera od strane imunih ćelija još uvek nisupoznati. Stoga ciljevi ove doktorske disertacije su ispitivanje kako α-Gal glikozilacijaproteina utiče na njihovu digestiju od strane pepsina, da li α-Gal glikozilacija proteinautiče na njihov transport kroz monosloj Caco-2 ćelija, koji oponaša gastrointestinalniepitel, kao i ispitivanje uticaja α-Gal glikozilacije na površini proteina na njihov unos iobradu od strane nezrelih dendritičnih ćelijama kultivisanih iz monocita (iMDDC). Izdobijenih rezultata moze se zaključiti da prisustvo α-Gal glikozilacije na površiniproteina utiče na njihovu podložnost na digestiju a najviše na obrazac dobijenihfragmenata proteina nakon pepsinolize. Veliki fragmenti proteina koji nose α-Galprisutni su čak i do 2 sata digestije. Takođe, važno je istaći da je transport α-Galglikozilovanih proteina kroz Caco-2 monosloj otežan u poređenju sa neglikozilovanimproteinima. Dalje, diferencijalnim centrifugiranjem lizata aco-2 ćelija nakontranscitoze pokazalo je da je α-Gal prisutan na intaktnim proteinima u endozomalnimfrakcijama ćelija... PB - Универзитет у Београду, Хемијски факултет T2 - Универзитет у Београду T1 - Monitoring of in vitro bioavailability and uptake of glycosylated food allergens using cell-based models UR - https://hdl.handle.net/21.15107/rcub_nardus_17803 ER -
@phdthesis{ author = "Krstić-Ristivojević, Maja", year = "2020", abstract = "extensive engagement of researchers in the elucidation of the mechanismsunderlying processes of food digestion, allergen transport, and uptake by the immunecells and its effector immune responses. The development of the in vitro assays andcell-based models has allowed bridging the problem of food allergy research and therespect of the ethical norms in used research procedures.The red meat allergy is a novel type of food allergy characterized by theproduction of an IgE antibody against the carbohydrate galactose-α-1,3-galactose (α-Gal). Glycoproteins from non-primate mammals are rich with an α-Gal as posttranslationalmodification. Also, red meat allergy is characterized by the delayed onsetof symptoms which may be related to the mechanism and the fate of α-Gal carryingproteins in the human gastrointestinal tract. Furthermore, the uptake, processing, andmechanisms of presentation of α-Gal by the immune cells are still unknown. Thereforethis doctoral dissertation aimed to investigate how protein glycosylation by α-Galaffects their susceptibility to gastric digestion, does α-Gal conjugated to proteins affectstheir transport through the Caco-2 cell monolayer, which mimics the gastrointestinallayer, and to examine the influence of α-Gal epitopes on the protein surface on theiruptake and processing by immature monocyte-derived dendritic cells (iMDDCs). Thestudy revealed that the presence of the α-Gal glycosylation on protein surface had animpact on their susceptibility to gastric digestion and the digestion pattern of theobtained protein fragments upon pepsinolysis. Prolonged survival, up to 2h of digestion,was characteristic of the large proteins fragments bearing the α-Gal epitope.Importantly, transport through the Caco-2 monolayer of proteins conjugated to α-Galwas hampered in comparison to unconjugated proteins. Furthermore, differentialcentrifugation of Caco-2 cell lysates upon transport experiments revealed that α-Galcould be detected on the intact protein in the endosomal fraction of the cells..., Alergije na hranu su rastući problem u ljudskoj populaciji širom sveta irešavanje ovog problema zahteva opsežno angažovanje istraživača u rasvetljavanjumehanizama uključenih u procese varenja hrane, transporta alergena i njihovog unosaod strane imunih ćelija odgovornih za efektorske mehanizme imunoloških odgovora.Ovo je nezamislivo bez razvoja in vitro testova i ćelijskih modela koji premošćujuproblem istraživanja alergija na hranu uz poštovanja etičkih normi u korišćenimistraživačkim metodama.Novu vrstu alergije na hranu, alergiju na crveno meso, karakteriše sintezaimunoglobulina E kao odgovor na prisustvo šećera galaktoza-α-1,3-galaktoza (α-Gal),koji je prisutan na površini glikoproteina primata. Takođe, alergiju na crveno mesokarakteriše odložena pojava simptoma što može biti rezultat promena u mehanizmuobrade proteina koji nose α-Gal u gastrointestinalnom traktu čoveka. Dalje, unos,obrada i mehanizmi prezentacije α-Gal šećera od strane imunih ćelija još uvek nisupoznati. Stoga ciljevi ove doktorske disertacije su ispitivanje kako α-Gal glikozilacijaproteina utiče na njihovu digestiju od strane pepsina, da li α-Gal glikozilacija proteinautiče na njihov transport kroz monosloj Caco-2 ćelija, koji oponaša gastrointestinalniepitel, kao i ispitivanje uticaja α-Gal glikozilacije na površini proteina na njihov unos iobradu od strane nezrelih dendritičnih ćelijama kultivisanih iz monocita (iMDDC). Izdobijenih rezultata moze se zaključiti da prisustvo α-Gal glikozilacije na površiniproteina utiče na njihovu podložnost na digestiju a najviše na obrazac dobijenihfragmenata proteina nakon pepsinolize. Veliki fragmenti proteina koji nose α-Galprisutni su čak i do 2 sata digestije. Takođe, važno je istaći da je transport α-Galglikozilovanih proteina kroz Caco-2 monosloj otežan u poređenju sa neglikozilovanimproteinima. Dalje, diferencijalnim centrifugiranjem lizata aco-2 ćelija nakontranscitoze pokazalo je da je α-Gal prisutan na intaktnim proteinima u endozomalnimfrakcijama ćelija...", publisher = "Универзитет у Београду, Хемијски факултет", journal = "Универзитет у Београду", title = "Monitoring of in vitro bioavailability and uptake of glycosylated food allergens using cell-based models", url = "https://hdl.handle.net/21.15107/rcub_nardus_17803" }
Krstić-Ristivojević, M.. (2020). Monitoring of in vitro bioavailability and uptake of glycosylated food allergens using cell-based models. in Универзитет у Београду Универзитет у Београду, Хемијски факултет.. https://hdl.handle.net/21.15107/rcub_nardus_17803
Krstić-Ristivojević M. Monitoring of in vitro bioavailability and uptake of glycosylated food allergens using cell-based models. in Универзитет у Београду. 2020;. https://hdl.handle.net/21.15107/rcub_nardus_17803 .
Krstić-Ristivojević, Maja, "Monitoring of in vitro bioavailability and uptake of glycosylated food allergens using cell-based models" in Универзитет у Београду (2020), https://hdl.handle.net/21.15107/rcub_nardus_17803 .