Immune and inflammatory responses to Leishmania amazonensis isolated from different clinical forms of human leishmaniasis in CBA mice

Leishmania amazonensis causes different diseases depending on the host and parasitic virulence factors. In this study, CBA mice were infected with L. amazonensis isolates from patients with localized (Ba125), diffuse cutaneous (Ba276) or visceral leishmaniasis (Ba109). Mice infected with Ba125 and Ba276 progressed rapidly and lesions displayed an infiltrate rich in parasitized macrophages and were necrotic and ulcerated. Ba109 induced smaller lesions and a mixed inflammatory infiltrate without necrosis or ulceration. Ba109 induced an insidious disease with lower parasite load in CBA mice, similar to human disease. Levels of IFN-γ, IL-4 and IL-10 did not differ among the groups. Because all groups were unable to control the infection, expression of IL-4 associated with low production of IFN-γ in the early phase of infection may account for susceptibility, but others factors may contribute to the differences observed in inflammatory responses and infection progression. Evaluation of some parasitic virulence factors revealed that Ba276 exhibits higher ecto-ADPase and 5'-nucleotidase activities compared to the Ba109 and Ba125 strains. Both Ba276 and Ba125 had higher arginase activity in comparison to Ba109. Finally, these data suggest that the differences in enzyme activities among parasites can account for differences in host inflammatory responses and infection progression

Expressão de RNAm de quimiocinas e citocinas na resposta imuno-inflamatória in situ de camundongos CBA infectados com diferentes especies de Leishmania

Submitted by Repositório Arca ([email protected]) on 2019-07-17T17:05:43Z No. of bitstreams: 1 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2019-07-29T16:11:41Z (GMT) No. of bitstreams: 2 Marcus Welby Borges 2000.pdf: 47801152 bytes, checksum: fd36c4593288c3503f7150f58d9a7ee4 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Made available in DSpace on 2019-07-29T16:11:41Z (GMT). No. of bitstreams: 2 Marcus Welby Borges 2000.pdf: 47801152 bytes, checksum: fd36c4593288c3503f7150f58d9a7ee4 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2000PAPES - 0250.250.354, CNPq - 523005/96-2, PRONEX.Universidade Federal da Bahia. Faculdade de Medicina. Salvador, BA, Brasil / Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, Brasil.Camundongos CBA são resistentes a infecção com L. major (Lm) e altamente susceptíveis a L. amazonensis (La). A resistência à infecção por Lm é mediada por uma resposta imune do tipo Th1 enquanto a susceptibilidade a La está associada com uma resposta imune do tipo Th2. As lesões de camundongos CBA infectados com L. major apresentam um infiltrado mononuclear misto, composto por poucos macrófagos infectados, granulomas e um aumento progressivo do número de linfócitos na lesão. Camundongos CBA infectados com L. amazonensis apresentam uma reação tecidual caracterizada pela presença de um infiltrado monomórfico, com macrófagos altamente parasitados e poucos linfócitos nas lesões desses animais. Considerando-se o papel de quimiocinas nos eventos relacionados com o recrutamento de leucócitos para os sítios inflamatórios, F-tés analisou-se se as diferenças existentes na composição do infiltrado de lesões de camundongos infectados com Lm ou La correlacionam-se com uma expressão diferenciada dessas moléculas. Para isso, infectou-se camundongos CBA com 5x106 promastigotas de L. major (Lm), L. amazonensis (La) e avaliou-se a expressão de RNAm para CXC quimiocinas (KC, MIG e CRG-2), CC quimiocinas (JE, MCP-5 e RANTES) e algumas citocinas (TNF-a e IL-4) nas lesões desses animais através da técnica de RT-PCR semi-quantitativa. Os resultados apresentados demonstraram que a expressão de RNAm para as quimiocinas MIG e MCP-5 foi significativamente maior nas lesões de camundongos do grupo Lm nos diferentes períodos após infecção. Camundongos infectados com La apresentaram uma baixa expressão de MIG, CRG-2, KC e TNF-a no primeiro dia após a infecção e um aumento na expressão de IL-4 no 40° dia. Esses dados demonstram uma expressão diferenciada de RNAm para quimiocinas e citocinas em camundongos CBA infectados com Lm e La que pode estar relacionada com os diferentes perfis de infiltrado inflamatório e resposta imuno-celular envolvidos com resistência ou susceptibilidade.CBA mice are resistant to L.major {Lm ) and are higlily susceptible to L. amazonensis {La) infection. Resistance and susceptibility in this model are associated with a Th1 and a Th2 type-immune response, respectively. Resistance to Lm is mediated by a Th1 type -immune response, whereas susceptibility to La is associated with a Th2 type-immune response. Lesions of Lm infected CBA mice present a mixed mononuclear inflammatory cell infiltrate composed of few infected macrophages, granulomas and a progressive increase in the number of lymphocytes. Incontrast, CBA mice infected with La show tissular reaction constituted by a monomorphic infiltrate of heavily parasitized macrophages and scanty lymphocytes. Given the role of chemokines in the recruitment of leukocytes, we decided to analyze whether differences in inflammatory infiltration observed in the CBA infected with La or Lm could be due to a differential expression of chemokine genes. CBA Mice were infected in the hind footpad with 5x10® promastigotes of La or Lm and a semi quantitative RT-PCR was used to measure the mRNA expression of CXC chemokines (KC, MIG and CRG-2), CC chemokines (JE/MCP-1, MCP-5 and RANTES) and some cytokines (TNF-a and IL-4). Our results show that the RNAm expression of MIG and MCP-5 was significantly increased in mice infected with Lm in the different times after infection. Mice infected with La showed a reduced expression of MIG, CRG-2, KC and TNF-a by the first day post-infection and an increase in the expression of IL-4 forty days after infection. Taken together, the results present here show a differential expression of chemokine mRNA in CBA mice infected with Lm as compared to those infected with La. These differences may be implicated in the different inflammatory cell infiltrate and cellular immune response that correlate with resistance or susceptibility

Avaliação dos perfis de resposta de macrófagos peritoneais de balb/c infectados in vitro com Leishmania amazonensis ou Leishmania braziliensis

Submitted by Repositório Arca ([email protected]) on 2019-09-04T17:23:44Z No. of bitstreams: 1 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2019-09-12T13:04:32Z (GMT) No. of bitstreams: 2 Marcus Welby Borges Avaliação...2008.pdf: 2589450 bytes, checksum: b0e5a7e4a0b656ef5c65412429ccbb5b (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5)Made available in DSpace on 2019-09-12T13:04:32Z (GMT). No. of bitstreams: 2 Marcus Welby Borges Avaliação...2008.pdf: 2589450 bytes, checksum: b0e5a7e4a0b656ef5c65412429ccbb5b (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2008Universidade Federal da Bahia. Faculdade de Medicina. Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, Brasil.Macrófagos (MФs) são células do sistema imunológico que desempenham importante papel na defesa contra Leishmania. Neste trabalho, foram investigados alguns fatores e mecanismos que podem estar envolvidos na determinação dos perfis de resposta de MФs peritoneais de BALB/c infectados in vitro com L. amazonensis ou L. braziliensis. Observou-se que IFN-y não foi capaz de reduzir a infecção causada por ambas as espécies de Leishmania. A sinergia entre IFN-y e TNF-a leva a uma diminuição do percentual de infecção dos MФs por La e Lb no tempo de 72h. Tanto o pré-tratamento com IFN-y quanto com IFN-y e TNF-a não alterou a viabilidade de L. amazonensis isoladas de MФs em relação ao grupo que não sofreu pré-tratamento. No entanto, L. braziliensis se mostrou susceptível a ação dessas citocinas, já que o pré-tratamento dos MФs com IFN-y foi capaz de diminuir o número de parasitos viáveis isolados. A sinergia entre IFN-y e TNF-a se mostrou bastante efetiva nesse grupo uma vez que o número de Lb isoladas a partir dos MФs infectados mostrou-se bastante reduzido. Embora não tenha sido detectada produção de NO nos grupos de MФs infectados com La e Lb, o bloqueio da produção dessa molécula, tanto in vitro quanto in vivo, é capaz de aumentar a infecção por Lb, mas não altera a infecção por La. A avaliação da produção de IL-12, revelou que MФs infectados com Lb produzem maior quantidade dessa citocina e apresentam uma maior expressão de RNAm de TNF-a nas primeiras 6h após infecção que os infectados por La. Por outro lado, a infecção com La induz uma maior produção de TGF-J3, maior expressão de RNAm de IL-10 e menor expressão de RNAm de quimiocinas e receptores dessas moléculas em relação ao grupo infectado com Lb. A investigação da atividade de arginase de La e Lb demonstrou que promastigotas de La apresentam uma elevada atividade de arginase, apresentando-se aproximadamente 50 a 100 vezes maior que nos promastigotas de Lb. Esses dados correlacionam-se com a maior carga parasitária e maior sobrevivência desse parasito no interior dos MФs, enquanto que a menor atividade dessa enzima em Lb pode estar relacionada com a baixa carga parasitária, alta susceptibilidade aos mecanismos microbicidas dos MФs e uma conseqüente diminuição na sobrevivência dessa espécie de Leishmania. Esses dados demonstram que La e Lb induzem perfis diferentes de resposta à infecção, tanto in vitro quanto in vivo, e sugerem que fatores produzidos pelo parasito, a exemplo de arginase, podem estar envolvidos no desenvolvimento desses perfis diferenciados de resposta.Macrophages (M(j)s) are cells that play an important role in the control of infection by Leishmania. In this work, we investigated some factors and mechanisms that could be related to the different responses of BALB/c peritoneal infected in vitro with L. amazonensis or L braziliensis. We observed that IFN-y alone was not able to reduce the infection by both Leishmania species. However, association of IFN-y and TNF-a resulted in a decrease of the percentual of M(j)S infected by Lb 72 h after infection, but not by La. Pre-treatment with IFN-y or IFN-y plus TNF-a did not change the viability of L. amazonensis compared to non-treated group. Nevertheless, L braziliensis was susceptible to the action of IFN-y because pre-treatment with this cytokine was sufficient to reduce the number of viable Lb isolated from M(|)S. The synergy of IFN-y and TNF-a was highly effective in this group and resolved the infection. We were not able to detect NO production by Ms infected with La or Lb. However, NO inhibition, in vitro and in vivo, increased Lb infection and did not change La infection. Lb infected M(j)S produced more IL-12 than La infected ones. Furthermore, M(|)S infected with Lb expressed more mRNA to TNF-a, chemokines and chemokine receptors than M(j)S infected with La. On the other hand. La induced a high production of TGF-p and IL-10 by M(j)S in relation to Lb. Arginase activity of parasites was evaluated and showed that La promastigotes had an arginase activity 50 to 100 times higher than Lb promastigotes. These data correlated to high parasite burden and survival of La in the M(j)S. On the contrary, low arginase activity in Lb is related to a low parasite burden and high susceptibility of the Lb to the M<j)S killing mechanisms. These data demonstrate that La and Lb induce different responses to infection, in vitro and in vivo, and suggest that factors produced by parasite, such as arginase, could be involved in the development of these profiles

Leishmania amazonensis: participation of regulatory T and B cells in the in vitro priming (PIV) of CBA/J spleen cells susceptible response.

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2014-06-02T17:38:10Z No. of bitstreams: 1 Veras PST Leishmania amazonensis....pdf: 304340 bytes, checksum: 057e83846ed49fde782d133e5d48ac0a (MD5)Made available in DSpace on 2014-06-02T17:38:10Z (GMT). No. of bitstreams: 1 Veras PST Leishmania amazonensis....pdf: 304340 bytes, checksum: 057e83846ed49fde782d133e5d48ac0a (MD5) Previous issue date: 2006Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Bio-intervenção. Salvador, BA, Brasil / Escola Bahiana de Medicina e Saúde Pública. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Bio-intervenção. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Bio-intervenção. Salvador, BA, Brasil / Instituto de Biologia da Universidade Federal da Bahia. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Imunologia Celular, Auto-imunidade e Doença de Chagas Experimental. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Imunologia Celular, Auto-imunidade e Doença de Chagas Experimental. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Bio-intervenção. Salvador, BA, Brasil / Faculdade de Medicina da Universidade Federal da Bahia. Salvador, BA, BraslCBA/J mice are resistant to Leishmania major and susceptible to Leishmania amazonensis. Early events determine infection outcome. Until now, PIV (in vitro priming) immune response to L. amazonensis has not been assessed. Herein, we have shown that compared to L. major, L. amazonensis induced higher parasite burden associated to similar IL-4, IFN- , and TNF- mRNA expressions and IFN- and IL-10 levels. Although similar amounts of IL-10 were detected, the frequency of intracellular IL-10 positive B cells was enhanced in spleen cells stimulated with anti-CD3/anti-CD28, or anti-CD3/anti-CD28 and L. amazonensis, compared to L. major-stimulation. Interestingly, IL-10- producing B cells were reduced in response to anti-CD3/anti-CD28 stimulation combined with L. major compared to the other groups. L. amazonensis may favor T regulatory cell development, since 40% of all the CD4+CD25+ were CD25high cells. These data suggest that in PIV, susceptibility to L. amazonensis is not related to Th cell polarization, but to the presence and activity of regulatory T and B cells

Immune and inflammatory responses to Leishmania amazonensis isolated from different clinical forms of human leishmaniasis in CBA mice

Leishmania amazonensis causes different diseases depending on the host and parasitic virulence factors. In this study, CBA mice were infected with L. amazonensis isolates from patients with localized (Ba125), diffuse cutaneous (Ba276) or visceral leishmaniasis (Ba109). Mice infected with Ba125 and Ba276 progressed rapidly and lesions displayed an infiltrate rich in parasitized macrophages and were necrotic and ulcerated. Ba109 induced smaller lesions and a mixed inflammatory infiltrate without necrosis or ulceration. Ba109 induced an insidious disease with lower parasite load in CBA mice, similar to human disease. Levels of IFN-γ, IL-4 and IL-10 did not differ among the groups. Because all groups were unable to control the infection, expression of IL-4 associated with low production of IFN-γ in the early phase of infection may account for susceptibility, but others factors may contribute to the differences observed in inflammatory responses and infection progression. Evaluation of some parasitic virulence factors revealed that Ba276 exhibits higher ecto-ADPase and 5&apos;-nucleotidase activities compared to the Ba109 and Ba125 strains. Both Ba276 and Ba125 had higher arginase activity in comparison to Ba109. Finally, these data suggest that the differences in enzyme activities among parasites can account for differences in host inflammatory responses and infection progression