Previously we demonstrated that human adenocarcinoma gastric cells AGS are able to counteract the detrimental effects of the ROS-inducer diallyl-disulfide. Here we report that such resistance can be extended to other pro-oxidant molecules. Conversely, AGS cells were induced to apoptosis via the mitochondrial pathway upon treatment with thiols-oxidizing agents, such as diamide. Apoptosis is associated with persistent oxidative damages, as evidenced by the increase of carbonylated proteins and the expression/activation of DNA damage-sensitive proteins histone H2A.X and DNA-PK. Resistance to H2O2, as well as sensitivity to diamide are correlated with GSH redox state, with H2O2 transiently increasing protein-GSH mixed disulfides, and diamide dramatically elevating GSSG. Moreover, the inhibition of GSH neo-synthesis, by buthionine sulfoximine, or the enrichment of intracellular sulphydryl pool, by N-acetylcysteine, was able to protect cells. Nrf2 rapidly translocates to the nuclei when AGS cells are treated with H2O2; whereas p53 is activated in response to diamide treatment by the oxidative induction of Trx1/p38MAPK signaling pathway. Finally, experiments carried out with p53 siRNA or dominant-negative Nrf2, indicate that these transcription factors play an important role in cell death and resistance to it. Our results are of particular importance for inducing apoptosis in tumor histotypes resistant to ROS-producing chemotherapeutics.
Piccirillo, S. (2009). Attivazione di differenti vie di segnalazione redox nella risposta cellulare allo stress ossidativo: ruolo del glutatione e dello status tiolico.
Attivazione di differenti vie di segnalazione redox nella risposta cellulare allo stress ossidativo: ruolo del glutatione e dello status tiolico
PICCIRILLO, SARA
2009-07-07
Abstract
Previously we demonstrated that human adenocarcinoma gastric cells AGS are able to counteract the detrimental effects of the ROS-inducer diallyl-disulfide. Here we report that such resistance can be extended to other pro-oxidant molecules. Conversely, AGS cells were induced to apoptosis via the mitochondrial pathway upon treatment with thiols-oxidizing agents, such as diamide. Apoptosis is associated with persistent oxidative damages, as evidenced by the increase of carbonylated proteins and the expression/activation of DNA damage-sensitive proteins histone H2A.X and DNA-PK. Resistance to H2O2, as well as sensitivity to diamide are correlated with GSH redox state, with H2O2 transiently increasing protein-GSH mixed disulfides, and diamide dramatically elevating GSSG. Moreover, the inhibition of GSH neo-synthesis, by buthionine sulfoximine, or the enrichment of intracellular sulphydryl pool, by N-acetylcysteine, was able to protect cells. Nrf2 rapidly translocates to the nuclei when AGS cells are treated with H2O2; whereas p53 is activated in response to diamide treatment by the oxidative induction of Trx1/p38MAPK signaling pathway. Finally, experiments carried out with p53 siRNA or dominant-negative Nrf2, indicate that these transcription factors play an important role in cell death and resistance to it. Our results are of particular importance for inducing apoptosis in tumor histotypes resistant to ROS-producing chemotherapeutics.Campo DC | Valore | Lingua |
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dc.authority.academicField2000 | Settore BIO/11 - BIOLOGIA MOLECOLARE | en |
dc.authority.otherpeople | CIRIOLO, MARIA ROSA | en |
dc.authority.people | PICCIRILLO, SARA | en |
dc.cilea.antefix | yes | it |
dc.collection.id.s | e291c0df-b2ad-cddb-e053-3a05fe0aa144 | * |
dc.collection.name | 07 - Tesi di dottorato | * |
dc.coverage.academiccycle | 21. | en |
dc.coverage.academicyear | A.A. 2008/2009 | en |
dc.date.accessioned | 2009-07-07T14:08:27Z | it |
dc.date.available | 2009-07-07T14:08:27Z | it |
dc.date.issued | 2009-07-07 | - |
dc.description | 21. ciclo | en |
dc.description.abstracteng | Previously we demonstrated that human adenocarcinoma gastric cells AGS are able to counteract the detrimental effects of the ROS-inducer diallyl-disulfide. Here we report that such resistance can be extended to other pro-oxidant molecules. Conversely, AGS cells were induced to apoptosis via the mitochondrial pathway upon treatment with thiols-oxidizing agents, such as diamide. Apoptosis is associated with persistent oxidative damages, as evidenced by the increase of carbonylated proteins and the expression/activation of DNA damage-sensitive proteins histone H2A.X and DNA-PK. Resistance to H2O2, as well as sensitivity to diamide are correlated with GSH redox state, with H2O2 transiently increasing protein-GSH mixed disulfides, and diamide dramatically elevating GSSG. Moreover, the inhibition of GSH neo-synthesis, by buthionine sulfoximine, or the enrichment of intracellular sulphydryl pool, by N-acetylcysteine, was able to protect cells. Nrf2 rapidly translocates to the nuclei when AGS cells are treated with H2O2; whereas p53 is activated in response to diamide treatment by the oxidative induction of Trx1/p38MAPK signaling pathway. Finally, experiments carried out with p53 siRNA or dominant-negative Nrf2, indicate that these transcription factors play an important role in cell death and resistance to it. Our results are of particular importance for inducing apoptosis in tumor histotypes resistant to ROS-producing chemotherapeutics. | - |
dc.description.allpeople | Piccirillo, Sara | it |
dc.description.allpeopleoriginal | Piccirillo, Sara | it |
dc.description.course | Biologia cellulare e molecolare | en |
dc.description.doctschool | Dottorato in biologia cellulare e molecolare | en |
dc.description.fulltext | open | en |
dc.description.tableofcontents | Biologia dei tumori - Processi coinvolti in tumorigenesi - Il modello della cellula staminale tumorale - Basi molecolari dello sviluppo e della progressione dei tumori gastrici - Stress ossidativo intrinseco in cellule tumorali - Lo stress ossidativo - Le specie reattive dell’ossigeno (ROS) - Le difese antiossidanti: sistemi di difesa enzimatici e non enzimatici - Ruolo dei sulfidrili proteici nel mantenimento dell’ambiente redox intracellulare - Alterazioni redox nei processi di traduzione del segnale - Segnalazione redox nella biologia dei tumori - Meccanismi alla base dello stress ossidativo in cellule tumorali - Conseguenze dello stress ossidativo in cellule tumorali - Implicazioni terapeutiche e tumori gastrici - Proteine redox regolate coinvolte in tumorigenesi - I fattori di trascrizione - La tioredossina - Procedure sperimentali - Colture cellulari - Trattamenti cellulari - Modulazione del contenuto intracellulare di glutatione - Inibitori delle MAP-chinasi - Analisi del ciclo cellulare e dell’apoptosi - Determinazione della morte cellulare mediante conta con Trypan blue - Determinazione delle proteine totali - Determinazione del glutatione e dei disolfuri misti mediante HPLC - Immunoelettroforesi (Western blotting) - Preparazione dei campioni per dosaggio elettroforetico - Transfezioni - Preparazione delle cellule per microscopia a fluorescenza - Urea PAGE - Costruzione dei plasmidi e mutagenesi sito-diretta - Analisi statistica dei dati - Scopo del lavoro - Risultati - Cellule di adenocarcinoma gastrico AGS mostrano diversa suscettibilità al trattamento con agenti pro-ossidanti - Il trattamento delle cellule AGS con diammide determina l’attivazione dell’apoptosi attraverso la via mitocondriale - I processi di resistenza e di suscettibilità allo stress ossidativo sono correlati alla modulazione del pool intracellulare del glutatione - Le cellule AGS mostrano danno ossidativo solo in seguito al trattamento con diammide - H2O2 e diammide modulano specificamente l’attività dei fattori di trascrizione Nrf2 e p53 - L’inibizione di Nrf2 e di p53 reverte la risposta cellulare - L’ossidazione della Trx1 correla con la fosforilazione di p53 mediata da p38MAPK - La Trx1 è il discriminante molecolare alla base della risposta allo stress ossidativo tiolico - Conclusioni. - Bibliografia | en |
dc.format.extent | 1871063 bytes | it |
dc.format.mimetype | application/pdf | it |
dc.identifier.citation | Piccirillo, S. (2009). Attivazione di differenti vie di segnalazione redox nella risposta cellulare allo stress ossidativo: ruolo del glutatione e dello status tiolico. | en |
dc.identifier.uri | http://hdl.handle.net/2108/915 | it |
dc.identifier.uri | http://hdl.handle.net/http://hdl.handle.net/2108/915 | - |
dc.language.iso | ita | en |
dc.publisher.country | Italy | - |
dc.publisher.name | Università degli Studi di Roma "Tor Vergata" | - |
dc.subject.keywordseng | glutatione; stress ossidativo; vie di segnalazione; p53; stato redox; nrf2 | - |
dc.subject.singlekeyword | glutatione | * |
dc.subject.singlekeyword | stress ossidativo | * |
dc.subject.singlekeyword | vie di segnalazione | * |
dc.subject.singlekeyword | p53 | * |
dc.subject.singlekeyword | stato redox | * |
dc.subject.singlekeyword | nrf2 | * |
dc.title | Attivazione di differenti vie di segnalazione redox nella risposta cellulare allo stress ossidativo: ruolo del glutatione e dello status tiolico | en |
dc.title.alternative | Activation of different redox-sensitive signaling pathways in cell response to oxidative stress: role of glutathione and thiolic status | en |
dc.type | Tesi di dottorato | - |
dc.type.driver | info:eu-repo/semantics/doctoralThesis | - |
dc.type.full | Pubblicazioni::07 - Tesi di dottorato | it |
iris.orcid.lastModifiedDate | 2023/07/26 13:21:51 | * |
iris.orcid.lastModifiedMillisecond | 1690370511063 | * |
Appare nelle tipologie: | 07 - Tesi di dottorato |
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