Células dentríticas condicionadas com rapamicina e pulsadas com aloantígenos na sobrevida de transplantes cutâneos alógenos neurocirúrgicos em ratos
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Arquivos
Data
2007
Autores
Horibe, Elaine Kawano [UNIFESP]
Orientadores
Ferreira, Lydia Masako [UNIFESP]
Tipo
Tese de doutorado
Título da Revista
ISSN da Revista
Título de Volume
Resumo
Objetivos: Comparar o fenótipo e capacidade estimulatória in vitro de populações de
células dendríticas (DC) de ratos; testar estas populações celulares na sobrevida de
retalhos cutâneos alógenos (RcuA); avaliar os mecanismos pelos quais as DC
condicionadas com rapamicina e pulsadas com lisado de hospedeiro, em associação a
uma curta administração de drogas imunossupressoras e soro anti-linfocitário atuam na
sobrevida de RCuA. Métodos: As DC foram condicionadas com rapamicina (DC
GM+Rapa) ou IL-4 (DC controle), sendo pulsadas ou não com aloantígenos de doador. O
fenótipo e função das células foram comparados in vitro. Testou-se o potencial destas DC
de aumentar a sobrevida de RCuA e comparou-se a hiporesponsividade doadoraespecífica
dos receptores de RCuA. Avaliou-se e quantificou-se a presença das células T
reguladoras Foxp3+CD4+ (Tregs) nos retalhos transplantados e nos órgãos linfóides
secundários com citometria de fluxo e imunofluorescência. Resultados: As DC
GM+Rapa expressaram níveis menores de MHCII e CD40 e apresentaram menor
capacidade para estimular células T em reação mista de linfócitos. Ao estímulo com LPS,
as DC GM+Rapa produziram níveis significantemente menores de IL-6 e IL-10.
Somente as DC GM+Rapa pulsadas com aloantígenos aumentaram significantemente a
sobrevida dos retalhos cutâneos alógenos (>180 dias). Células T periféricas em
receptores com sobrevida prolongada de RcuA mostraram hiporesponsividade doadorespecífica.
Quantidades significantemente maiores de Tregs foram encontradas nos
retalhos com sobrevida prolongada e nos órgãos linfóides secundários destes receptores.
Conclusões: As DC GM+Rapa apresentaram fenótipo mais imaturo e menor capacidade
de estimulação de células T do que a população controle. A administração de DC
GM+Rapa pulsadas com aloantígenos prolongou significantemente a sobrevida dos
RCuA, sendo este fato associado à expansão de Tregs.
Introduction: Clinically applicable protocols capable of promoting operational tolerance to vascularized skin allografts (VSA) would contribute towards a more widespread use of composite tissue allografts (CTA). This study compared tolerogeneic properties of different host bone-marrow (BM)-derived dendritic cells (DC) populations in vitro and their potential to prolong survival of VSA across a full MHC mismatch in rats. Methods: DC were generated either with GM-CSF, rapamycin and donor-alloAg-pulsing (GM+Rapa DCp) or GM-CSF and IL-4, with (control DCp) or without alloAg-pulsing (control DC). Phenotype and function were assessed in vitro. We then tested the potential of those DC, combined with transient cyclosporine and anti-lymphocyte serum treatment to mediate prolonged survival of VSA. Graft survival and donor-specific hyporesponsiveness were compared. Presence of Foxp3+CD4+T cells (Tregs) in VSA and secondary lymphoid organs was assessed and quantified by FACS and immunofluorescence. Results: These myeloid DC populations demonstrated LPS stimulation-resistant immature phenotypes. GM+Rapa DC displayed significantly lower T-cell stimulatory capacity and produced less proinflammatory cytokines upon TLR activation. When combined with transient immunosuppresion, GM+Rapa DCp was the only population to significantly prolong VSA survival (>180days), in a donor-specific hyporesponsive milieu. Significantly higher amounts of Tregs were detected in long-term surviving VSA and secondary lymphoid organs of these recipients. Conclusion: Among the DC populations studied herein, GM+Rapa DCp showed the most tolerogeneic profile in vitro and when employed in a clinically applicable protocol in vivo led to operational tolerance of VSA, an unprecedented finding in literature. Furthermore, this is the first report to correlate use of rapamycintreated DC with Tregs expansion in vivo.
Introduction: Clinically applicable protocols capable of promoting operational tolerance to vascularized skin allografts (VSA) would contribute towards a more widespread use of composite tissue allografts (CTA). This study compared tolerogeneic properties of different host bone-marrow (BM)-derived dendritic cells (DC) populations in vitro and their potential to prolong survival of VSA across a full MHC mismatch in rats. Methods: DC were generated either with GM-CSF, rapamycin and donor-alloAg-pulsing (GM+Rapa DCp) or GM-CSF and IL-4, with (control DCp) or without alloAg-pulsing (control DC). Phenotype and function were assessed in vitro. We then tested the potential of those DC, combined with transient cyclosporine and anti-lymphocyte serum treatment to mediate prolonged survival of VSA. Graft survival and donor-specific hyporesponsiveness were compared. Presence of Foxp3+CD4+T cells (Tregs) in VSA and secondary lymphoid organs was assessed and quantified by FACS and immunofluorescence. Results: These myeloid DC populations demonstrated LPS stimulation-resistant immature phenotypes. GM+Rapa DC displayed significantly lower T-cell stimulatory capacity and produced less proinflammatory cytokines upon TLR activation. When combined with transient immunosuppresion, GM+Rapa DCp was the only population to significantly prolong VSA survival (>180days), in a donor-specific hyporesponsive milieu. Significantly higher amounts of Tregs were detected in long-term surviving VSA and secondary lymphoid organs of these recipients. Conclusion: Among the DC populations studied herein, GM+Rapa DCp showed the most tolerogeneic profile in vitro and when employed in a clinically applicable protocol in vivo led to operational tolerance of VSA, an unprecedented finding in literature. Furthermore, this is the first report to correlate use of rapamycintreated DC with Tregs expansion in vivo.
Descrição
Citação
HORIBE, Elaine Kawano Horibe. Células dendríticas condicionadas com rapamicina e pulsadas com aloantígenos na sobrevida de transplantes cutâneos alógenos microcirúrgicos em ratos. 2007. 110 f.
Tese (Doutorado em Ciências) – Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, 2007.