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Immune mechanisms of induction and expression of contact hypersensitivity reaction in rats

dc.contributor.advisorMirkov, Ivana
dc.contributor.advisorKataranovski, Milena
dc.contributor.otherMedenica, Ljiljana
dc.contributor.otherVučević, Dragana
dc.creatorPopov Aleksandrov, Aleksandra D.
dc.date.accessioned2017-11-23T08:23:46Z
dc.date.available2017-11-23T08:23:46Z
dc.date.issued2013
dc.identifier.urihttp://eteze.bg.ac.rs/application/showtheses?thesesId=809
dc.identifier.urihttps://fedorabg.bg.ac.rs/fedora/get/o:7142/bdef:Content/download
dc.identifier.urihttp://vbs.rs/scripts/cobiss?command=DISPLAY&base=70036&RID=1024629938
dc.identifier.urihttp://nardus.mpn.gov.rs/123456789/2108
dc.identifier.urihttps://ibiss-r.rcub.bg.ac.rs/handle/123456789/2408
dc.descriptionAlergijski kontaktni dermatitis (AKD) je česta zapaljenska bolest kože kod ljudi, koja pogađa 15-20% svetske populacije. Velika zastupljenost hemijskih agenasa (haptena) koji izazivaju AKD, uz odsustvo adekvatne konvencionalne imunosupresivne terapije, čine ovu bolest velikim izazovom za javno zdravlje, i savremenu imunologiju. Razvoj životinjskog modela AKD, reakcije kontaktne preosetljivosti, omogućio je napredak u razumevanju njene patogeneze. Iako je reakcija kontaktne preosetljivosti (KP) dosta ispitivana, pretežno na mišjem modelu, mehanizmi u osnovi razvoja bolesti nisu dovoljno poznati. Takođe, mali je broj podataka o reakciji KP kod drugih eksperimentalnih životinja, osim miševa, koji bi mogli da unaprede znanje o imunobiologiji kontaktnih alergija. U svetlu toga, u ovom radu su ispitane karakteristike reakcije KP kod pacova, korišćenjem dva soja pacova, Dark Agouti (DA) i Albino Oxford (AO), za koje je pokazano da se razlikuju u reaktivnosti na pojedine zapaljenske stimuluse. Indukcija (senzibilizacija) i izazivanje (elicitacija) reakcije KP vršene su sa haptenom dinitrohlorobenzenom (DNCB), koji je primenjen u dve različite koncentracije (režim senzibilizacije/elicitacije 0.4%/0.13% i 4%/1.3% DNCB). Mehanizmi KP kod pacova ispitani su u fazi senzibilizacije i elicitacije reakcije, praćenjem dinamike promena aktivnosti ćelija drenirajućih limfnih čvorova (dLČ) u fazi senzibilizacije, i ispitivanjem produkcije inflamatornih/ anti-inflamatornih medijatora u kondicioniranom medijumu organokulture uha, i promena aktivnosti ćelija dLČ u fazi elicitacije reakcije (24 časa nakon elicitacije). Rezultati su pokazali povećanje in vivo (otok uha) i ex vivo (patohistološka analiza isečaka kože uha) parametara reakcije KP kod oba soja pacova nakon tretmana sa višom dozom haptena, ali je reakcija bila intenzivnija kod DA u odnosu AO soj. Nakon senzibilizacije dolazi do: povećanja broja ćelija u dLČ kod oba soja (izraženije kod DA jedinki), promene zastupljenosti CD4+ i CD8+ ćelija i njihovog odnosa (povećanja odnosa CD4+/CD8+ ćelija kod DA soja, zbog povećanja zastupljenosti CD4+ ćelija a pada zastupljenosti CD8+ ćelija, i smanjenja odnosa CD4+/CD8+ kod AO soja, zbog pada zastupljenosti CD4+ ćelija a povećanja CD8+ ćelija), povećanja ex vivo proliferacije izolovanih ćelija i povećanja spontane i specifične (nakon in vitro stimulacije ćelija sa haptenom) produkcije inflamatornihcitokina IFN-γ i IL-17, i Th2/anti-inflamatornog citokina IL-10 (izraženije kod DA soja)...sr
dc.descriptionAllergic contact dermatitis (ACD) is a common skin inflammatory disease in humans, affecting 15-20% of the general population. Large number of chemical agents that induce ACD (called haptens), together with lack of adequate conventional immunosuppressive therapy, make this disease a major challenge for public health and modern immunology. Pathophysiology of ACD is studied on animal models referred as contact hypersensitivity. Although the reaction of contact hypersensitivity (CHS) has been much studied, mostly in mice, the knowledge of its mechanisms is still far from complete. In addition, there is paucity of data about CHS in other experimental animals, beside mice, which might increase our understanding of the immunobiology of contact allergy. In light of this, in the present study the characteristics of CHS were analyzed in rats, using two strains, Dark Agouti (DA) i Albino Oxford (AO), which have been shown to differ in reactivity to certain inflammatory stimuli. CHS to hapten dinitrochlorobenzene (DNCB) was induced by applying two different hapten concentrations in sensitization and elicitation phases (sensitization/elicitation regime 0.4%/0.13% and 4%/1.3% DNCB). Mechanisms of CHS were studied in induction (sensitization) and expression (elicitation) phases, by evaluating dynamics in activity changes of draining lymph node (dLN) cells in sensitization phase and by evaluating production of inflammatory/anti-inflammatory mediators in conditioned medium of ear skin organ culture and activity changes of dLN cells in elicitation phase (24 hours after elicitation). The results showed increased in vivo (ear thickness) and ex vivo (patohistology analysis of ear skin) parameters of CHS reaction, in both rat strains following the treatment with high DNCB dose, but the reaction was more intensive in DA compared to AO rats. In sensitization phase it was showed: increased dLN cell number in both strains (higher in DA rats), changes in ratio of CD4+ and CD8+ subsets (increased ratio of CD4+/CD8+ cells in DA rats, because of increased relative number of CD4+ and decreased relative number of CD8+ cells and decreased CD4+/CD8+ ratio in AO rats, because of decreased relative number of CD4+ cells and increased CD8+ cells relative number), increased ex vivo proliferation of dLN cells and increased spontaneousand specific (induced by in vitro cell stimulation with hapten) production of inflammatory cytokines IFN-γ and IL-17 and Th2/anti-inflammatory cytokine IL-10 (more pronounced in DA rats)...en
dc.description.abstractAlergijski kontaktni dermatitis (AKD) je česta zapaljenska bolest kože kod ljudi, koja pogađa 15-20% svetske populacije. Velika zastupljenost hemijskih agenasa (haptena) koji izazivaju AKD, uz odsustvo adekvatne konvencionalne imunosupresivne terapije, čine ovu bolest velikim izazovom za javno zdravlje, i savremenu imunologiju. Razvoj životinjskog modela AKD, reakcije kontaktne preosetljivosti, omogućio je napredak u razumevanju njene patogeneze. Iako je reakcija kontaktne preosetljivosti (KP) dosta ispitivana, pretežno na mišjem modelu, mehanizmi u osnovi razvoja bolesti nisu dovoljno poznati. Takođe, mali je broj podataka o reakciji KP kod drugih eksperimentalnih životinja, osim miševa, koji bi mogli da unaprede znanje o imunobiologiji kontaktnih alergija. U svetlu toga, u ovom radu su ispitane karakteristike reakcije KP kod pacova, korišćenjem dva soja pacova, Dark Agouti (DA) i Albino Oxford (AO), za koje je pokazano da se razlikuju u reaktivnosti na pojedine zapaljenske stimuluse. Indukcija (senzibilizacija) i izazivanje (elicitacija) reakcije KP vršene su sa haptenom dinitrohlorobenzenom (DNCB), koji je primenjen u dve različite koncentracije (režim senzibilizacije/elicitacije 0.4%/0.13% i 4%/1.3% DNCB). Mehanizmi KP kod pacova ispitani su u fazi senzibilizacije i elicitacije reakcije, praćenjem dinamike promena aktivnosti ćelija drenirajućih limfnih čvorova (dLČ) u fazi senzibilizacije, i ispitivanjem produkcije inflamatornih/ anti-inflamatornih medijatora u kondicioniranom medijumu organokulture uha, i promena aktivnosti ćelija dLČ u fazi elicitacije reakcije (24 časa nakon elicitacije). Rezultati su pokazali povećanje in vivo (otok uha) i ex vivo (patohistološka analiza isečaka kože uha) parametara reakcije KP kod oba soja pacova nakon tretmana sa višom dozom haptena, ali je reakcija bila intenzivnija kod DA u odnosu AO soj. Nakon senzibilizacije dolazi do: povećanja broja ćelija u dLČ kod oba soja (izraženije kod DA jedinki), promene zastupljenosti CD4+ i CD8+ ćelija i njihovog odnosa (povećanja odnosa CD4+/CD8+ ćelija kod DA soja, zbog povećanja zastupljenosti CD4+ ćelija a pada zastupljenosti CD8+ ćelija, i smanjenja odnosa CD4+/CD8+ kod AO soja, zbog pada zastupljenosti CD4+ ćelija a povećanja CD8+ ćelija), povećanja ex vivo proliferacije izolovanih ćelija i povećanja spontane i specifične (nakon in vitro stimulacije ćelija sa haptenom) produkcije inflamatornih citokina IFN-γ i IL-17, i Th2/anti-inflamatornog citokina IL-10 (izraženije kod DA soja). U ovoj fazi reakcije kod oba soja nije zapažena promena u produkciji i ekspresiji Th2/anti-inflamatornog citokina IL-4. U fazi ekspresije (elicitacije) reakcije pokazano je da je povećanje otoka uha praćeno povećanjem produkcije TNF-α, NO, MPO, IFN-γ i IL-17 u kondicioniranom medijumu organokulture eksplanata kože uha, kao i da su razlike u ekspresiji reakcije između dva soja praćene razlikama u intenzitetu promena (veća koncentracija NO, MPO, IFN-γ, IL-17 kod DA soja) i tipu imunskog odgovora (povećana produkcija IL-4 samo kod AO soja). U ovoj fazi reakcije pokazano je povećanje aktivnosti ćelija dLČ (spontane i specifične proliferacije, uz povećanje ekspresije inducibilnog α lanca receptora za faktor rasta IL-2, kao i produkcije efektorskih citokina IFN-γ i IL-17, i ekspresije gena za p35, p40 i T-bet, koji su pokazatelji diferencijacije Th1/tip1 ćelija) kod DA soja nakon tretmana sa DNCB-om, u odnosu na kontrole. Odsustvo promena u aktivnosti efektorskih mehanizama u dLČ kod jedinki AO soja je praćeno povećanom produkcijom IL-4, i povećanim brojem CD4+CD25+Foxp3+ ćelija, koje fenotipski odgovaraju regulatornim T ćelijama (Treg). Reakcija KP na DNCB kod pacova (slično mišjem modelu) karakteriše se aktivacijom Th1/tip1 i Th17/tip imunskog odgovora, a razlike u intenzitetu odgovora (u fazi senzibilizacije i elicitacije) ili tipu odgovora (u fazi elicitacije), mogu biti u osnovi zabeleženih međusojnih razlika u intenzitetu ekspresije reakcije. Komparativno ispitivanje kontaktne preosetljivosti kod reaktivnijeg DA soja i manje reaktivnog AO soja, može doprineti saznanjima o imunskim mehanizmima koji su u osnovi razvoja i ekspresije, kao i regulacije ove reakcije. Ovakva saznanja mogu dati dobru osnovu za razvoj novih terapeutskih modaliteta u lečenju alergijskog kontaktnog dermatitisa, ali i osnovu za ispitivanje imunotoksičnog potencijala hemijskih agenasa, posebno onih koji dospevaju u organizam preko kože.sr
dc.description.abstractAllergic contact dermatitis (ACD) is a common skin inflammatory disease in humans, affecting 15-20% of the general population. Large number of chemical agents that induce ACD (called haptens), together with lack of adequate conventional immunosuppressive therapy, make this disease a major challenge for public health and modern immunology. Pathophysiology of ACD is studied on animal models referred as contact hypersensitivity. Although the reaction of contact hypersensitivity (CHS) has been much studied, mostly in mice, the knowledge of its mechanisms is still far from complete. In addition, there is paucity of data about CHS in other experimental animals, beside mice, which might increase our understanding of the immunobiology of contact allergy. In light of this, in the present study the characteristics of CHS were analyzed in rats, using two strains, Dark Agouti (DA) i Albino Oxford (AO), which have been shown to differ in reactivity to certain inflammatory stimuli. CHS to hapten dinitrochlorobenzene (DNCB) was induced by applying two different hapten concentrations in sensitization and elicitation phases (sensitization/elicitation regime 0.4%/0.13% and 4%/1.3% DNCB). Mechanisms of CHS were studied in induction (sensitization) and expression (elicitation) phases, by evaluating dynamics in activity changes of draining lymph node (dLN) cells in sensitization phase and by evaluating production of inflammatory/anti-inflammatory mediators in conditioned medium of ear skin organ culture and activity changes of dLN cells in elicitation phase (24 hours after elicitation). The results showed increased in vivo (ear thickness) and ex vivo (patohistology analysis of ear skin) parameters of CHS reaction, in both rat strains following the treatment with high DNCB dose, but the reaction was more intensive in DA compared to AO rats. In sensitization phase it was showed: increased dLN cell number in both strains (higher in DA rats), changes in ratio of CD4+ and CD8+ subsets (increased ratio of CD4+/CD8+ cells in DA rats, because of increased relative number of CD4+ and decreased relative number of CD8+ cells and decreased CD4+/CD8+ ratio in AO rats, because of decreased relative number of CD4+ cells and increased CD8+ cells relative number), increased ex vivo proliferation of dLN cells and increased spontaneous and specific (induced by in vitro cell stimulation with hapten) production of inflammatory cytokines IFN-γ and IL-17 and Th2/anti-inflammatory cytokine IL-10 (more pronounced in DA rats). Similarly to DA rats, no changes in production and expression of Th2/anti-inflammatory cytokine IL-4 were noted in AO rats in sensitization phase. Increased ear thickness in elicitation phase was accompanied by increased production of TNF-α, NO, MPO, IFN-γ and IL-17 in conditioned medium of ear skin organ culture. Strain-dependent differences in CHS expression were related to differences in intensity (higher NO, MPO, IFN-γ, IL-17 in DA strain) and type of immune response (increased production of IL-4 only in AO rats). In addition, in this phase of the reaction it was showed the increase in dLN activity in DNCB-treated DA rats (increased spontaneous and specific proliferation, concomitantly with increased expression of growth factor IL-2 receptor α chain, as well as production of effector cytokines IFN-γ and IL-17, and gene expression of p35, p40 and T-bet, which are indicators of differentiation of Th1/type1 cells), compared to controls. The absence of changes in the activity of effector mechanisms in dLN of AO strain is accompanied by increased production of IL-4 and increased numbers of CD4+CD25+Foxp3+ cells, that phenotypically correspond to regulatory T cells (Treg). CHS to DNCB in rats (similarly to mouse model) is characterized by activation of Th1/type1 and Th17/type17 immune response, and differences in response intensity (in induction and expression phase) and response type (in expression phase) could be underlying mechanism of strain-dependent difference. Comparative analysis of CHS in more reactive (DA) and less reactive (AO) strains, can contribute to the further knowledge of the immunological mechanisms underlying the development and expression, as well as regulation of this reaction. Such findings might be useful in exploring the potential of therapeutic modalities to reduce skin damage caused by contact allergy, as well as in the elucidation of immunomodulatory potential of different chemicals, particularly those that gain access to the host system via skin.en
dc.formatapplication/pdf
dc.languagesr
dc.publisherBelgrade: University of Belgrade, Faculty of Biology
dc.rightsopenAccess
dc.sourceUniversity of Belgrade, Faculty of Biology
dc.subjectReakcija kontaktne preosetljivostisr
dc.subjectDNCBsr
dc.subjectFaza senzibilizacijesr
dc.subjectFaza elicitacije reakcijesr
dc.subjectDA i AO pacovisr
dc.subjectDrenirajući limfni čvorovisr
dc.subjectOrganokultura eksplantata kože uhasr
dc.subjectContact hypersensitivity reactionen
dc.subjectDNCBen
dc.subjectSensitization phaseen
dc.subjectElicitation phaseen
dc.subjectDA and AO ratsen
dc.subjectDraining lymph nodesen
dc.subjectEar skin organ cultureen
dc.titleImunski mehanizmi indukcije i ekspresije reakcije kontaktne preosetljivosti kod pacovasr
dc.titleImmune mechanisms of induction and expression of contact hypersensitivity reaction in ratsen
dc.typedoctoralThesis
dc.rights.licenseBY-NC-ND
dcterms.abstractМирков, Ивана; Катарановски,Милена; Меденица, Љиљана; Вучевић, Драгана; Попов Aлександров, Aлександра Д.; Имунски механизми индукције и експресије реакције контактне преосетљивости код пацова; Имунски механизми индукције и експресије реакције контактне преосетљивости код пацова;
dc.citation.apaPopov Aleksandrov, A. (2013). Imunski mehanizmi indukcije i ekspresije reakcije kontaktne preosetljivosti kod pacova. University of Belgrade, Faculty of Biology. p. 147.
dc.citation.vancouverPopov Aleksandrov A. Imunski mehanizmi indukcije i ekspresije reakcije kontaktne preosetljivosti kod pacova [dissertation]. Belgrade: University of Belgrade, Faculty of Biology; 2013. 147 p.
dc.citation.spage1
dc.citation.epage147
dc.type.versionpublishedVersionen
dc.identifier.fulltexthttp://ibiss-r.rcub.bg.ac.rs//bitstream/id/212/Popov_Aleksandrov_Aleksandra_dissertation.pdf


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