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Protective mechanism of arylpiperazine dopaminergic D2ligands on nitric oxide and 6-hydroxydopamine induced SH-SY5Y human neuroblastoma cell death

dc.contributor.advisorAnđus, Pavle
dc.contributor.advisorZogović, Nevena
dc.contributor.otherTrajković, Vladimir
dc.contributor.otherTomić, Mirko
dc.creatorTovilović-Kovačević, Gordana
dc.date.accessioned2017-11-23T08:24:15Z
dc.date.available2017-11-23T08:24:15Z
dc.date.issued2012
dc.identifier.urihttp://eteze.bg.ac.rs/application/showtheses?thesesId=167
dc.identifier.urihttps://fedorabg.bg.ac.rs/fedora/get/o:5401/bdef:Content/download
dc.identifier.urihttp://vbs.rs/scripts/cobiss?command=DISPLAY&base=70036&RID=1024560050
dc.identifier.urihttp://nardus.mpn.gov.rs/123456789/2081
dc.identifier.urihttps://ibiss-r.rcub.bg.ac.rs/handle/123456789/2424
dc.descriptionU ovom radu je ispitivan uticaj 20 novosintetisanih arilpiperazinskihdopaminergičkih liganada na vijabilnost humanih SH-SY5Y neuroblastomskih ćelijatretiranih donorom azot monoksida (engl. nitric oxide, NO) natrijum nitroprusidom(engl. sodium nitroprusside, SNP) i uzročnikom oksidativnog stresa dopaminergičkimneurotoksinom 6-hidroksidopaminom (6-OHDA). Supstanca koje je pružala najjačuzaštitu od donora NO je bio N-{4-[2-(4-fenil-piperazin-1-il)-etil]-fenil}-pikolinamid(arilpiperazin 6a), dok je N-{3-[2-(4-fenil-piperazin-1-il)-etil]-fenil}-pikolinamid(arilpiperazin 6b) najefikasnije štitio humanu neuroblastomsku ćelijsku liniju SH-SY5Yod 6-OHDA. Arilpiperazin 6a je delimično sprečavao povećanje sadržaja superoksidanjon radikala, smanjenje potencijala membrane mitohondrija i unutarćelijskog sadržajaadenozin-trifosfata (ATP), aktivaciju kaspaza i sledstvenu fragmentaciju DNK koje jeizazivao NO. Uočeno smanjenje unutarćelijske koncentracije superoksida nije biloposledica direktne interakcije ispitivanog arilpiperazina sa O2-∙, niti je supstanca 6auticala na akumulaciju NO unutar ćelije. Arilpiperazin 6a je sprečavao inhibicijuprotektivnog Akt, kao i aktivaciju proapoptotskih ERK, JNK i AMPK signalnih putevaizazvane SNP-om, ukazujući na bitnu ulogu ovih molekula u njegovom zaštitnomdelovanju. Potencijalni značaj arilpiperazina 6a u sprečavanjuneurodegenerativnih/neurozapaljenskih procesa posebno naglašava činjenica da je štitioneuronima slične SH-SY5Y ćelije i od citotoksičnog efekta NO-a poreklom odstimulisanih makrofaga. Slično, arilpiperazin 6b je sprečavao povećanje unutarćelijskogsadržaja superoksid anjon radikala (O2-∙), smanjenje potencijala membrane mitohondrija posledične apoptotske događaje – aktivaciju kaspaza i fragmentaciju DNK – koje jeizazivao 6-OHDA. Stabilizacija potencijala mitohondrijalne membrane pod dejstvomarilpiperazina 6b se vremenski poklapala sa smanjenjem unutarćelijskog sadržaja O2-∙,što ukazuje da je supstanca 6b inhibirala oslobađanje superoksida iz oštećenihmitohondrija stabilizacijom potencijala njihove membrane...sr
dc.descriptionWe investigated the protective ability of 20 novel arylpiperazine-baseddopaminergic ligands against nitric oxide (NO) and dopaminergic neurotoxin6-hydroxydopamine (6-OHDA)-mediated neurotoxicity. The most potentneuroprotective compound against NO-induced toxicity was N-{4-[2-(4-phenylpiperazin-1-yl)-ethyl]-phenyl}-picolinamide (arylpiperazine 6a), while N-{3-[2-(4-phenyl-piperazin-1-yl)-ethyl]-phenyl}-picolinamide (arylpiperazine 6b) mosteffectively protected SH-SY5Y human neuron-like cells from 6-OHDA-generatedoxidative injury. Arylpiperazine 6a diminished the proapoptotic action of NO donorsodium nitroprusside (SNP) by decreasing superoxide anion content, mitochondrialmembrane depolarization, decline in intracellular adenozine-triphosphate (ATP)content, caspase activation and subsequent phosphatydilserine externalization/DNAfragmentation. The observed decrease of intracellular superoxide concentration was notmediated by direct O2-∙ scavenging. Arylpiperazine 6a did not interfere with NOaccumulation within the cell. The protective effect of arylpiperazine 6a in NO-inducedstress was associated with activation of anti-apoptotic (Akt) and the inhibition ofproapoptotic (JNK, ERK, AMPK) signaling pathways. A potential therapeutic value ofthe arylpiperazine 6a in neurodegenerative/neuroinflammatory diseases prevention wasadditionally supported by the ability of this arylpiperazine to protect SH-SY5Y neuronlikecells from macrophage-derived NO. Similarly, arylpiperazine 6b prevented6-OHDA-induced increase in superoxide anion content, mitochondrial membrane depolarization and following apoptotic related events – caspase activation and DNAfragmentation. The stabilization of 6-OHDA-disrupted mitochondrial membranepotential by arylpiperazine 6b correlated with the decrease in intracellular superoxideanion (O2-∙) content, suggesting that decline in O2-∙ concentration resulted from mitohondrial membrane stabilizationen
dc.description.abstractU ovom radu je ispitivan uticaj 20 novosintetisanih arilpiperazinskih dopaminergičkih liganada na vijabilnost humanih SH-SY5Y neuroblastomskih ćelija tretiranih donorom azot monoksida (engl. nitric oxide, NO) natrijum nitroprusidom (engl. sodium nitroprusside, SNP) i uzročnikom oksidativnog stresa dopaminergičkim neurotoksinom 6-hidroksidopaminom (6-OHDA). Supstanca koje je pružala najjaču zaštitu od donora NO je bio N-{4-[2-(4-fenil-piperazin-1-il)-etil]-fenil}-pikolinamid (arilpiperazin 6a), dok je N-{3-[2-(4-fenil-piperazin-1-il)-etil]-fenil}-pikolinamid (arilpiperazin 6b) najefikasnije štitio humanu neuroblastomsku ćelijsku liniju SH-SY5Y od 6-OHDA. Arilpiperazin 6a je delimično sprečavao povećanje sadržaja superoksid anjon radikala, smanjenje potencijala membrane mitohondrija i unutarćelijskog sadržaja adenozin-trifosfata (ATP), aktivaciju kaspaza i sledstvenu fragmentaciju DNK koje je izazivao NO. Uočeno smanjenje unutarćelijske koncentracije superoksida nije bilo posledica direktne interakcije ispitivanog arilpiperazina sa O2 -∙, niti je supstanca 6a uticala na akumulaciju NO unutar ćelije. Arilpiperazin 6a je sprečavao inhibiciju protektivnog Akt, kao i aktivaciju proapoptotskih ERK, JNK i AMPK signalnih puteva izazvane SNP-om, ukazujući na bitnu ulogu ovih molekula u njegovom zaštitnom delovanju. Potencijalni značaj arilpiperazina 6a u sprečavanju neurodegenerativnih/neurozapaljenskih procesa posebno naglašava činjenica da je štitio neuronima slične SH-SY5Y ćelije i od citotoksičnog efekta NO-a poreklom od stimulisanih makrofaga. Slično, arilpiperazin 6b je sprečavao povećanje unutarćelijskog sadržaja superoksid anjon radikala (O2 -∙), smanjenje potencijala membrane mitohondrija i posledične apoptotske događaje – aktivaciju kaspaza i fragmentaciju DNK – koje je izazivao 6-OHDA. Stabilizacija potencijala mitohondrijalne membrane pod dejstvom arilpiperazina 6b se vremenski poklapala sa smanjenjem unutarćelijskog sadržaja O2 -∙, što ukazuje da je supstanca 6b inhibirala oslobađanje superoksida iz oštećenih mitohondrija stabilizacijom potencijala njihove membrane. Arilpiperazin 6b nije pokazao značajnu sposobnost direktnog uklanjanja superoksid anjon radikala. Zaštitni mehanizam supstance 6b nije uključivao interakciju sa JNK i ERK proapoptotskim putevima, koje je aktivirao 6-OHDA. Međutim, arilpiperazin 6b je izazvao dodatnu prolaznu aktivaciju preživljavajućeg Akt signalnog puta u ćelijama tretiranim 6-OHDA-om, pa se ne može isključiti doprinos Akt-a antiapoptotskom efektu arilpiperazina 6b. Pored apoptoze, 6-OHDA je izazivao i citotoksičnu autofagiju u SH-SY5Y ćelijama smanjujući aktivnost ključnog inhibitora autofagije u ćeliji − mTOR proteina. Arilpiperazin 6b je delimično sprečavao inhibiciju aktivnosti mTOR-a i posledičnu autofagiju zabeležene u prisustvu 6-OHDA. Ovaj efekat arilpiperazina 6b je bio nezavisan od aktivnosti pozitivnog regulatora autofagije AMPK. Iako su oba arilpiperazina pokazivala parcijalnu agonističku aktivnost na D2 receptorima njihov neuroprotektivni efekat je bio nezavisan od vezivanja za dopaminske receptore, jer ga visokoafinitetni blokatori D1/D2 receptora butaklamol (u slučaju 6a) ili selektivni blokator D2 receptora sulpirid (u slučaju 6b) nisu poništavali. Ovi rezultati ukazuju da arilpiperazini predstavljaju dobre kandidate za razvoj potencijalnih neuroprotektivnih lekova.sr
dc.description.abstractWe investigated the protective ability of 20 novel arylpiperazine-based dopaminergic ligands against nitric oxide (NO) and dopaminergic neurotoxin 6-hydroxydopamine (6-OHDA)-mediated neurotoxicity. The most potent neuroprotective compound against NO-induced toxicity was N-{4-[2-(4-phenylpiperazin- 1-yl)-ethyl]-phenyl}-picolinamide (arylpiperazine 6a), while N-{3-[2-(4- phenyl-piperazin-1-yl)-ethyl]-phenyl}-picolinamide (arylpiperazine 6b) most effectively protected SH-SY5Y human neuron-like cells from 6-OHDA-generated oxidative injury. Arylpiperazine 6a diminished the proapoptotic action of NO donor sodium nitroprusside (SNP) by decreasing superoxide anion content, mitochondrial membrane depolarization, decline in intracellular adenozine-triphosphate (ATP) content, caspase activation and subsequent phosphatydilserine externalization/DNA fragmentation. The observed decrease of intracellular superoxide concentration was not mediated by direct O2 -∙ scavenging. Arylpiperazine 6a did not interfere with NO accumulation within the cell. The protective effect of arylpiperazine 6a in NO-induced stress was associated with activation of anti-apoptotic (Akt) and the inhibition of proapoptotic (JNK, ERK, AMPK) signaling pathways. A potential therapeutic value of the arylpiperazine 6a in neurodegenerative/neuroinflammatory diseases prevention was additionally supported by the ability of this arylpiperazine to protect SH-SY5Y neuronlike cells from macrophage-derived NO. Similarly, arylpiperazine 6b prevented 6-OHDA-induced increase in superoxide anion content, mitochondrial membrane depolarization and following apoptotic related events – caspase activation and DNA fragmentation. The stabilization of 6-OHDA-disrupted mitochondrial membrane potential by arylpiperazine 6b correlated with the decrease in intracellular superoxide anion (O2 -∙) content, suggesting that decline in O2 -∙ concentration resulted from mitohondrial membrane stabilization. Arylpiperazine 6b did not posses significant O2 -∙ scavenging ability. The mechanism underlying the protection provided by the compound 6b was independent of JNK- and ERK-proapoptotic pathways, as 6-OHDA-provoked activation of these proapototic pathways was not modified by arylpiperazine 6b. However, arylpiperazine 6b caused additional transient activation of pro-survival Akt signaling pathway in 6-OHDA-treated cells, which probably contributed to its anti-apoptotic effect. In addition to apoptosis, 6-OHDA induced deleterious autophagy in SH-SY5Y cells by silencing the activity of mTOR, a key autophagy inhibitor in the cell. Arylpiperazine 6b partialy prevented 6-OHDA-triggered decrease of mTOR activity and autophagy in AMPK-independent manner. Although both compounds showed partial agonist activity on hD2 receptors, their neuroprotective action was independent of dopamine receptor binding, as it was not affected by the high-affinity D1/D2 receptor blocker butaclamol (6a) or selective D2 receptor blocker sulpiride (6b). These results support further study of arylpiperazines as potential neuroprotective drugs.en
dc.formatapplication/pdf
dc.languagesr
dc.publisherBelgrade: University of Belgrade, Faculty of Biology
dc.rightsopenAccess
dc.sourceUniversity of Belgrade, Faculty of Biology
dc.subjectArilpiperazinisr
dc.subjectNeuroprotekcijasr
dc.subjectAzot monoksidsr
dc.subjectOksidativni stressr
dc.subject6-hidroksidopaminsr
dc.subjectApoptozasr
dc.subjectAutofagijasr
dc.subjectDopaminski D2 receptorisr
dc.subjectArylpiperazinesen
dc.subjectNeuroprotectionen
dc.subjectNitric oxideen
dc.subjectOxidative stressen
dc.subject6-hydroxydopamineen
dc.subjectApoptosisen
dc.subjectAutophagyen
dc.subjectDopamine D2 receptorsen
dc.titleMehanizam zaštitnog dejstva arilpiperazinskih liganada za dopaminske D2 receptore u azot monoksidom i 6-hidroksidopaminom izazvanoj smrti SH-SY5Y ćelija humanog neuroblastomasr
dc.titleProtective mechanism of arylpiperazine dopaminergic D2ligands on nitric oxide and 6-hydroxydopamine induced SH-SY5Y human neuroblastoma cell deathen
dc.typedoctoralThesis
dc.rights.licenseBY-NC-ND
dcterms.abstractAнђус, Павле; Зоговић, Невена; Трајковић, Владимир; Томић, Мирко; Товиловић-Ковачевић, Гордана;
dc.citation.apaTovilović, G. (2012). Mehanizam zaštitnog dejstva arilpiperazinskih liganada za dopaminske D2 receptore u azot monoksidom i 6-hidroksidopaminom izazvanoj smrti SH-SY5Y ćelija humanog neuroblastoma. University of Belgrade, Faculty of Biology. 116 p.
dc.citation.vancouverTovilović G. Mehanizam zaštitnog dejstva arilpiperazinskih liganada za dopaminske D2 receptore u azot monoksidom i 6-hidroksidopaminom izazvanoj smrti SH-SY5Y ćelija humanog neuroblastoma [dissertation]. Belgrade: University of Belgrade, Faculty of Biology; 2012. p. 116.
dc.citation.spage1
dc.citation.epage116
dc.type.versionpublishedVersionen
dc.identifier.fulltexthttp://ibiss-r.rcub.bg.ac.rs//bitstream/id/260/Tovilovic_Gordana_dissertation.pdf


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