Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp.
Ano de defesa: | 2013 |
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Autor(a) principal: | |
Orientador(a): | |
Banca de defesa: | , , , , |
Tipo de documento: | Tese |
Tipo de acesso: | Acesso aberto |
Idioma: | por |
Instituição de defesa: |
Universidade Federal de Goiás
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Programa de Pós-Graduação: |
Programa de Pós-graduação em Medicina Tropical e Saúde Publica (IPTSP)
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Departamento: |
Instituto de Patologia Tropical e Saúde Pública - IPTSP (RG)
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País: |
Brasil
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Palavras-chave em Português: | |
Palavras-chave em Inglês: | |
Área do conhecimento CNPq: | |
Link de acesso: | http://repositorio.bc.ufg.br/tede/handle/tede/5958 |
Resumo: | American Tegumentary Leishmaniasis (ATL) is a disease caused by protozoa Leishmania. In Brazil, the most prevalent species is L. (Viannia) braziliensis. Several cytokines and receptors are involved in immunopathogenesis of ATL, however, the role of interleukin 32 (IL-32) was not investigated in this disease. Besides, toll-like receptors (TLR) were poorly evaluated in Leishmania infection, especially when it is caused by L. (V.) braziliensis. The aim of the present study was to evaluate IL-32, TNF and IL-10 expression in ATL lesions; the induction of IL-32 by L. (V.) braziliensis amastigotes in human peripheral blood mononuclear cells (PBMC) cultures as well as the involvement of TLR4 in monocyte/macrophage response to L. (V.) brazilienis amastigotes. Biopsies fragments from cutaneous and mucosal lesion and healthy tissues were used to investigate the subgenus of the parasites by PCR-RFLP assay; expression of IL-32, TNF and IL-10 was assayed by immunohistochemistry and expression of IL-32 isoforms , TNF and IL-10 was analysed by qRT-PCR. The PBMC were cultured with L. (V.) braziliensis amastigotes in the absence or presence of IFN and IL-32 induction was assayed by qRT-PCR; and TNF and IL-10, by ELISA. TLR4 was neutralized by monoclonal antibodies and lipopolysaccharide (LPS) was used as TLR4 agonist. The expression of TLR4 in monocyte/macrophages was evaluated by flow cytometry. Thirty five patients were evaluated, 23 with cutaneous leishmaniasis (CL) and 12 with mucosal leihsmaniasis (ML). All parasite positive samples contained L. (Viannia) sp. The expression of IL-32 (protein and mRNA) was similar in CL and ML lesions but was higher than in health tissues. Only IL-32 was detected. The proteins TNF and IL-10 were detected in similar levels in CL and ML lesions, but TNF mRNA was present in higher levels in ML (4.069x) than in CL lesions (141 x, p < 0.05). L. (V.) braziliensis amastigotes induced IL-32, TNF and IL-10 in IFN pre-treated PBMC. The production of TNF and IL-10 was TLR4 dependent and treatment of PBMC with LPS further increased the production of TNF induced by amastigotes (p < 0.05). However, LPS did not altere the IL-10 production. Treatment with IFN enhanced the percentage of TLR4+ monocyte/macrophage (p < 0.05), which was decreased following incubation with amastigotes (p = 0.06). The results showed that IL-32 is produced during L. (Viannia) infection and TLR4 mediates L. (V.) braziliensis amastigote-induced TNF and IL-10 production in human PBMC. Moreover, the data suggest that amastigotes can lead to TLR4 internalization what can allow parasite to evade of innate immune response. This study indicates that IL-32 and TLR4 are important players in human infection caused by L. (Viannia), especially L. (V.) braziliensis. Whether TLR4 is also important to IL-32 production by human monocytes/macrophages deserves further investigation. |
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Dias, Fátima Ribeirohttp://lattes.cnpq.br/5741031258926403Dias, Fátima RibeiroJunqueira, Maria Imaculada Muniz BarbozaShio, Marina TiemiMarques, Mara RubiaLino Júnior, Ruy de Souzahttp://lattes.cnpq.br/1772950544649876Galdino Júnior, Hélio2016-08-18T12:50:37Z2013-07-26Galdino Júnior, H. Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp.. 2013.122 f. Tese (Doutorado em Medicina Tropical e Saúde Publica) - Universidade Federal de Goiás, Goiânia, 2016.http://repositorio.bc.ufg.br/tede/handle/tede/5958American Tegumentary Leishmaniasis (ATL) is a disease caused by protozoa Leishmania. In Brazil, the most prevalent species is L. (Viannia) braziliensis. Several cytokines and receptors are involved in immunopathogenesis of ATL, however, the role of interleukin 32 (IL-32) was not investigated in this disease. Besides, toll-like receptors (TLR) were poorly evaluated in Leishmania infection, especially when it is caused by L. (V.) braziliensis. The aim of the present study was to evaluate IL-32, TNF and IL-10 expression in ATL lesions; the induction of IL-32 by L. (V.) braziliensis amastigotes in human peripheral blood mononuclear cells (PBMC) cultures as well as the involvement of TLR4 in monocyte/macrophage response to L. (V.) brazilienis amastigotes. Biopsies fragments from cutaneous and mucosal lesion and healthy tissues were used to investigate the subgenus of the parasites by PCR-RFLP assay; expression of IL-32, TNF and IL-10 was assayed by immunohistochemistry and expression of IL-32 isoforms , TNF and IL-10 was analysed by qRT-PCR. The PBMC were cultured with L. (V.) braziliensis amastigotes in the absence or presence of IFN and IL-32 induction was assayed by qRT-PCR; and TNF and IL-10, by ELISA. TLR4 was neutralized by monoclonal antibodies and lipopolysaccharide (LPS) was used as TLR4 agonist. The expression of TLR4 in monocyte/macrophages was evaluated by flow cytometry. Thirty five patients were evaluated, 23 with cutaneous leishmaniasis (CL) and 12 with mucosal leihsmaniasis (ML). All parasite positive samples contained L. (Viannia) sp. The expression of IL-32 (protein and mRNA) was similar in CL and ML lesions but was higher than in health tissues. Only IL-32 was detected. The proteins TNF and IL-10 were detected in similar levels in CL and ML lesions, but TNF mRNA was present in higher levels in ML (4.069x) than in CL lesions (141 x, p < 0.05). L. (V.) braziliensis amastigotes induced IL-32, TNF and IL-10 in IFN pre-treated PBMC. The production of TNF and IL-10 was TLR4 dependent and treatment of PBMC with LPS further increased the production of TNF induced by amastigotes (p < 0.05). However, LPS did not altere the IL-10 production. Treatment with IFN enhanced the percentage of TLR4+ monocyte/macrophage (p < 0.05), which was decreased following incubation with amastigotes (p = 0.06). The results showed that IL-32 is produced during L. (Viannia) infection and TLR4 mediates L. (V.) braziliensis amastigote-induced TNF and IL-10 production in human PBMC. Moreover, the data suggest that amastigotes can lead to TLR4 internalization what can allow parasite to evade of innate immune response. This study indicates that IL-32 and TLR4 are important players in human infection caused by L. (Viannia), especially L. (V.) braziliensis. Whether TLR4 is also important to IL-32 production by human monocytes/macrophages deserves further investigation.A leishmaniose Tegumentar Americana (LTA) é uma doença infecto-parasitária, causada, no Brasil, principalmente pela espécie Leishmania (Viannia) braziliensis. Na imunopatogenia da LTA participam várias citocinas e receptores. A interleucina 32 (IL-32) é uma citocina pro-inflamatória, cuja participação na LTA ainda não foi investigada. O papel dos receptores similares a Toll (TLR) na LTA tem sido pouco investigado, especialmente considerando infecção por L. (V.) braziliensis. O objetivo deste estudo foi avaliar a expressão da IL-32, do TNF e da IL-10 nas lesões de pacientes com LTA, a indução de IL-32 por formas amastigotas de L. (V.) braziliensis em células mononucleares (CMN) humanas, bem como avaliar a participação do TLR4 na infecção de monócitos/macrófagos humanos com formas amastigotas de L. (V.) brazilienis. Fragmentos de lesões cutâneas ou mucosas de pacientes com LTA e tecidos sadios foram obtidos, sendo usados para a determinação do subgênero de Leishmania, usando a PCR-RFLP; para análise de IL-32, TNF e IL-10 por imunoistoquímica; e para análise da expressão das isoformas da IL-32, do TNF e da IL-10, por qRT-PCR. As CMN foram cultivadas com amastigotas de L. (V.) braziliensis, na ausência ou presença de IFN e a indução de IL-32 foi avaliada por qRT-PCR e TNF e IL-10, por ELISA. Para neutralização de TLR4 foram usados anticorpos monoclonais e lipopolissacarídeo (LPS), como agonista de TLR4. A expressão de TLR4 nos monócitos/macrófagos presentes nas CMN foi avaliada por citometria de fluxo. Foram analisados 35 pacientes sendo 23 leishmaniose cutânea (LC) e 12 com leishmaniose mucosa (LM). Todas as amostras PCR positivas continham parasitos pertencentes ao subgênero Viannia. Nas lesões de pacientes com LC e LM foi detectada uma maior expressão tanto da proteína quanto do mRNA da IL-32 do que nos tecidos controles, porém níveis similares foram encontrados nas duas formas clínicas. Somente a isoforma IL-32 foi detectada. As proteínas TNF e a IL-10 foram detectadas nas lesões, em níveis similares na LC e na LM, porém, o mRNA do TNF estava em níveis mais elevados nas lesões de LM (4.069 vezes) do que nas lesões de LC (141 vezes, p < 0,05). As formas amastigotas de L. (V.) braziliensis indiziram a produção de IL-32 TNF e de IL-10 nas CMN pré-tratadas com IFN. A produção de TNF e IL-10 foi dependente de TLR4 e o tratamento das CMN com LPS aumentou a produção do TNF induzido pelas amastigotas (p < 0,05), mas não alterou a produção da IL-10. A incubação com IFN aumentou significantemente a porcentagem de monócitos/macrófagos TLR4+ (p < 0,05), a qual foi diminuída pelas formas amastigotas (p = 0,06). Os resultados mostram que a IL-32 é produzida durante a infecção humana causada por L. (Viannia) e evidenciam a participação de TLR4 na indução de TNF e IL-10 em CMN humanas por formas amastigotas de L. (V.) braziliensis. Ainda, os dados sugerem que as formas amastigotas causam a internalização dos TLR4, o que pode ser um mecanismo de evasão da resposta imune. Portanto, a IL-32 e o TLR4 parecem importantes componentes na infecção humana causada por L. (Viannia), especialmente L. (V.) braziliensis, podendo contribuir para a imunopatogenia ou para o controle da infecção.Submitted by Erika Demachki (erikademachki@gmail.com) on 2016-08-17T20:24:56Z No. of bitstreams: 2 Tese - Hélio Galdino Júnior - 2013.pdf: 3450540 bytes, checksum: 01adc52cca765449b5eed357351fb5ee (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5)Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2016-08-18T12:50:37Z (GMT) No. of bitstreams: 2 Tese - Hélio Galdino Júnior - 2013.pdf: 3450540 bytes, checksum: 01adc52cca765449b5eed357351fb5ee (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5)Made available in DSpace on 2016-08-18T12:50:37Z (GMT). No. of bitstreams: 2 Tese - Hélio Galdino Júnior - 2013.pdf: 3450540 bytes, checksum: 01adc52cca765449b5eed357351fb5ee (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2013-07-26application/pdfporUniversidade Federal de GoiásPrograma de Pós-graduação em Medicina Tropical e Saúde Publica (IPTSP)UFGBrasilInstituto de Patologia Tropical e Saúde Pública - IPTSP (RG)http://creativecommons.org/licenses/by-nc-nd/4.0/info:eu-repo/semantics/openAccessLeishmanioseIL-32TLR4Leishmania(Viania) spLeishmaniasisCIENCIAS BIOLOGICAS::IMUNOLOGIAAvaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp.Evaluation of the participation of cytokines (interleukin-32, interleukin-10, tumor necrosis factor) and Toll-Like receptors (TLR 4) in infection by Leishmania (V.) sp.info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/doctoralThesis6085308344741430434600600600-77690114445645562885989919188376747614reponame:Repositório Institucional da UFGinstname:Universidade Federal de Goiás (UFG)instacron:UFGLICENSElicense.txtlicense.txttext/plain; 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dc.title.por.fl_str_mv |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. |
dc.title.alternative.eng.fl_str_mv |
Evaluation of the participation of cytokines (interleukin-32, interleukin-10, tumor necrosis factor) and Toll-Like receptors (TLR 4) in infection by Leishmania (V.) sp. |
title |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. |
spellingShingle |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. Galdino Júnior, Hélio Leishmaniose IL-32 TLR4 Leishmania(Viania) sp Leishmaniasis CIENCIAS BIOLOGICAS::IMUNOLOGIA |
title_short |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. |
title_full |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. |
title_fullStr |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. |
title_full_unstemmed |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. |
title_sort |
Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp. |
author |
Galdino Júnior, Hélio |
author_facet |
Galdino Júnior, Hélio |
author_role |
author |
dc.contributor.advisor1.fl_str_mv |
Dias, Fátima Ribeiro |
dc.contributor.advisor1Lattes.fl_str_mv |
http://lattes.cnpq.br/5741031258926403 |
dc.contributor.referee1.fl_str_mv |
Dias, Fátima Ribeiro |
dc.contributor.referee2.fl_str_mv |
Junqueira, Maria Imaculada Muniz Barboza |
dc.contributor.referee3.fl_str_mv |
Shio, Marina Tiemi |
dc.contributor.referee4.fl_str_mv |
Marques, Mara Rubia |
dc.contributor.referee5.fl_str_mv |
Lino Júnior, Ruy de Souza |
dc.contributor.authorLattes.fl_str_mv |
http://lattes.cnpq.br/1772950544649876 |
dc.contributor.author.fl_str_mv |
Galdino Júnior, Hélio |
contributor_str_mv |
Dias, Fátima Ribeiro Dias, Fátima Ribeiro Junqueira, Maria Imaculada Muniz Barboza Shio, Marina Tiemi Marques, Mara Rubia Lino Júnior, Ruy de Souza |
dc.subject.por.fl_str_mv |
Leishmaniose IL-32 TLR4 Leishmania(Viania) sp |
topic |
Leishmaniose IL-32 TLR4 Leishmania(Viania) sp Leishmaniasis CIENCIAS BIOLOGICAS::IMUNOLOGIA |
dc.subject.eng.fl_str_mv |
Leishmaniasis |
dc.subject.cnpq.fl_str_mv |
CIENCIAS BIOLOGICAS::IMUNOLOGIA |
description |
American Tegumentary Leishmaniasis (ATL) is a disease caused by protozoa Leishmania. In Brazil, the most prevalent species is L. (Viannia) braziliensis. Several cytokines and receptors are involved in immunopathogenesis of ATL, however, the role of interleukin 32 (IL-32) was not investigated in this disease. Besides, toll-like receptors (TLR) were poorly evaluated in Leishmania infection, especially when it is caused by L. (V.) braziliensis. The aim of the present study was to evaluate IL-32, TNF and IL-10 expression in ATL lesions; the induction of IL-32 by L. (V.) braziliensis amastigotes in human peripheral blood mononuclear cells (PBMC) cultures as well as the involvement of TLR4 in monocyte/macrophage response to L. (V.) brazilienis amastigotes. Biopsies fragments from cutaneous and mucosal lesion and healthy tissues were used to investigate the subgenus of the parasites by PCR-RFLP assay; expression of IL-32, TNF and IL-10 was assayed by immunohistochemistry and expression of IL-32 isoforms , TNF and IL-10 was analysed by qRT-PCR. The PBMC were cultured with L. (V.) braziliensis amastigotes in the absence or presence of IFN and IL-32 induction was assayed by qRT-PCR; and TNF and IL-10, by ELISA. TLR4 was neutralized by monoclonal antibodies and lipopolysaccharide (LPS) was used as TLR4 agonist. The expression of TLR4 in monocyte/macrophages was evaluated by flow cytometry. Thirty five patients were evaluated, 23 with cutaneous leishmaniasis (CL) and 12 with mucosal leihsmaniasis (ML). All parasite positive samples contained L. (Viannia) sp. The expression of IL-32 (protein and mRNA) was similar in CL and ML lesions but was higher than in health tissues. Only IL-32 was detected. The proteins TNF and IL-10 were detected in similar levels in CL and ML lesions, but TNF mRNA was present in higher levels in ML (4.069x) than in CL lesions (141 x, p < 0.05). L. (V.) braziliensis amastigotes induced IL-32, TNF and IL-10 in IFN pre-treated PBMC. The production of TNF and IL-10 was TLR4 dependent and treatment of PBMC with LPS further increased the production of TNF induced by amastigotes (p < 0.05). However, LPS did not altere the IL-10 production. Treatment with IFN enhanced the percentage of TLR4+ monocyte/macrophage (p < 0.05), which was decreased following incubation with amastigotes (p = 0.06). The results showed that IL-32 is produced during L. (Viannia) infection and TLR4 mediates L. (V.) braziliensis amastigote-induced TNF and IL-10 production in human PBMC. Moreover, the data suggest that amastigotes can lead to TLR4 internalization what can allow parasite to evade of innate immune response. This study indicates that IL-32 and TLR4 are important players in human infection caused by L. (Viannia), especially L. (V.) braziliensis. Whether TLR4 is also important to IL-32 production by human monocytes/macrophages deserves further investigation. |
publishDate |
2013 |
dc.date.issued.fl_str_mv |
2013-07-26 |
dc.date.accessioned.fl_str_mv |
2016-08-18T12:50:37Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/doctoralThesis |
format |
doctoralThesis |
status_str |
publishedVersion |
dc.identifier.citation.fl_str_mv |
Galdino Júnior, H. Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp.. 2013.122 f. Tese (Doutorado em Medicina Tropical e Saúde Publica) - Universidade Federal de Goiás, Goiânia, 2016. |
dc.identifier.uri.fl_str_mv |
http://repositorio.bc.ufg.br/tede/handle/tede/5958 |
identifier_str_mv |
Galdino Júnior, H. Avaliação da participação de citocinas (interleucina 32, interleucina 10, fator de necrose tumoral) e receptores similares a Toll (TLR 4) na infecção por Leishmania (Viannia) sp.. 2013.122 f. Tese (Doutorado em Medicina Tropical e Saúde Publica) - Universidade Federal de Goiás, Goiânia, 2016. |
url |
http://repositorio.bc.ufg.br/tede/handle/tede/5958 |
dc.language.iso.fl_str_mv |
por |
language |
por |
dc.relation.program.fl_str_mv |
6085308344741430434 |
dc.relation.confidence.fl_str_mv |
600 600 600 |
dc.relation.department.fl_str_mv |
-7769011444564556288 |
dc.relation.cnpq.fl_str_mv |
5989919188376747614 |
dc.rights.driver.fl_str_mv |
http://creativecommons.org/licenses/by-nc-nd/4.0/ info:eu-repo/semantics/openAccess |
rights_invalid_str_mv |
http://creativecommons.org/licenses/by-nc-nd/4.0/ |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Universidade Federal de Goiás |
dc.publisher.program.fl_str_mv |
Programa de Pós-graduação em Medicina Tropical e Saúde Publica (IPTSP) |
dc.publisher.initials.fl_str_mv |
UFG |
dc.publisher.country.fl_str_mv |
Brasil |
dc.publisher.department.fl_str_mv |
Instituto de Patologia Tropical e Saúde Pública - IPTSP (RG) |
publisher.none.fl_str_mv |
Universidade Federal de Goiás |
dc.source.none.fl_str_mv |
reponame:Repositório Institucional da UFG instname:Universidade Federal de Goiás (UFG) instacron:UFG |
instname_str |
Universidade Federal de Goiás (UFG) |
instacron_str |
UFG |
institution |
UFG |
reponame_str |
Repositório Institucional da UFG |
collection |
Repositório Institucional da UFG |
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Repositório Institucional da UFG - Universidade Federal de Goiás (UFG) |
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tasesdissertacoes.bc@ufg.br |
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