Infectividade e patogenicidade de diferentes isolados de Leishmania (Leishmania) chagasi obtidos de c?es naturalmente infectados.

Programa de P?s-Gradua??o em Ci?ncias Biol?gicas. N?cleo de Pesquisas em Ci?ncias Biol?gicas, Pr?-Reitoria de Pesquisa de P?s Gradua??o, Universidade Federal de Ouro Preto.No Brasil, a Leishmaniose Visceral (LV), que tem como agente etiol?gico a Leishmania chagasi (sinon?mia Leishmania infantum), vem sendo amplamente distribu?da. O c?o se destaca como reservat?rio dom?stico deste parasito, favorecendo sua transmiss?o no ambiente urbano. Considerando que a transmiss?o da L. chagasi ocorre necessariamente com a participa??o dos c?es no ambiente urbano, e que muitos deles podem ter sido tratados ou estar em tratamento contra esta infec??o, sem induzir cura parasitol?gica, ? poss?vel especular que em ambientes dom?sticos possam estar circulando cepas com caracter?sticas de resist?ncia ao tratamento, bem como alta infectividade e patogenicidade. Neste contexto, torna-se fundamental a caracteriza??o de isolados de L. chagasi, oriundos de c?es naturalmente infectados, bem como o estudo do comportamento in vitro e in vivo destes isolados para identifica??o de padr?es de infectividade e patogenicidade. Dentre os resultados obtidos neste estudo, foi poss?vel confirmar, pela PCR-RFLP, que todos os isolados s?o da esp?cie L. chagasi. De forma interessante, o cultivo dos diferentes isolados em linhagem de macr?fago canino (DH82) evidenciou perfis distintos de infectividade e carga parasit?ria. Esta abordagem permitiu identificar os isolados com comportamentos polares in vitro, representados por infectividade e carga parasit?ria elevada (isolado 616) ou reduzida (isolado 614). Ap?s este estudo inicial in vitro, foi realizado uma an?lise do perfil de infec??o em hamsters, utilizando estes isolados, bem como a cepa padr?o PP75 e a cepa selvagem OP46. Em um segundo experimento com estes isolados e com as cepas padr?o (PP75 e OP46), os animais de cada um dos grupos foram submetidos a uma interven??o com a administra??o de Glucantime? em dose subterap?utica. De forma interessante, a taxa de sobrevida nos animais infectados com o isolado 616 foi de 100%, enquanto que para os animais infectados com o isolado 614 (com ou sem interven??o medicamentosa), bem como com o isolado 616 (nos animais que receberam Glucantime?) foi em torno de 50%. De um modo em geral, o isolado 614 induziu: (i) maiores altera??es clinicopatol?gicas, (ii) indicativo de altera??o hep?tica pelas an?lises bioqu?micas, (iii) intenso infiltrado inflamat?rio no f?gado, (iv) eleva??o da carga parasit?ria avaliada por qPCR no ba?o e no f?gado. Estes resultados mostraram que o isolado que foi menos infectivo (614) nos ensaios in vitro foi o mais infectivo e patog?nico em hamsters. Por outro lado, na presen?a de infec??o com o isolado 616 e com administra??o de Glucantime?, o perfil da infec??o mostrou maior gravidade, alterando, portanto, a evolu??o da infec??o. Em conjunto, os dados obtidos pela an?lise in vitro em macr?fagos caninos da linhagem DH82, a partir de isolados de L. chagasi apresentando perfis polares de infectividade, n?o refletiram o mesmo padr?o nos ensaios in vivo utilizando hamsters. Al?m disto, foi observado que o tratamento n?o efetivo da leishmaniose visceral experimental pode induzir altera??o no curso da infec??o, tornando-a mais agressiva.In Brazil, Visceral Leishmaniasis (VL) is caused by Leishmania chagasi (synonymy Leishmania infantum), has been widely distributed. The dog is considered the domestic reservoir of the parasite, favoring its transmission in the urban environment. Whereas the transmission of L. chagasi necessarily occurs with dog?s participation in the urban area, and that many of them may be in treatment for this infection without inducing parasitological cure. Considering this scenario, is possible to speculate that parasite transmission would include strains with characteristics as high infectivity, pathogenicity and resistance to treatment. In this context, it is fundamental to identify L. chagasi strains, derived from L. chagasi-naturally infected dogs including in vitro and in vivo analysis for identification of infectivity and pathogenicity patterns. The data considering all analyzed strains were confirmed by PCR-RFLP as L. chagasi specie. Interestingly, the in vitro approach using canine macrophage lineage (DH82) showed distinct infectivity and parasite burden profiles. This analysis allowed us to identify strains with in vitro polar patterns, represented by high (strain 616) or low (strain 614) infectivity and parasite load. After this initial in vitro study, it was carried out an in vivo analysis of the infection profile using hamsters and the strains 614, 616, in addition to PP75 and wild type OP46 (standard strains). In a second experiment with these strains, the hamsters of each group were subjected to an intervention by the administration of Glucantime?. Interestingly, the survival rate of the infected animals using 616 strains was 100%, whereas for strain 614 (with or without Glucantime? treatment), as well as to 616 strains (after Glucantime? treatment) was around 50%. The major results using 614 strain in hamsters demonstrated: (i) higher clinicopathological changes, (ii) indicative of liver disorders according to biochemical analysis, (iii) intense liver inflammation, (iv) increased parasite burden assessed by qPCR in both spleen and liver. These results showed that the less infective strain (614) in the in vitro assays was the most pathogenic and infective in hamsters. Moreover, the infection with 616 strain following by Glucantime? treatment, the infection profile showed greater severity by changing the infection outcome. Taken together, the in vitro data analysis using canine macrophages (DH82 lineage) from L. chagasi strains presenting polar infectivity profiles did not reflect the same outcome by in vivo analysis using hamsters. Furthermore, it was observed that the non-effective treatment of experimental visceral leishmaniasis might induce changes in the infection outcome making it more pathogenic

Perfil de citocinas, linfócitos T e níveis de óxido nítrico em cães imunizados com as vacinas LBSap e LBSapSal e submetidos ao desafio experimental com Leishmania (Leishmania) chagasi e saliva de Lutzomyia longipalpis.

Os cães representam o reservatório doméstico mais importante de L. chagasi (sin. L. infantum), e uma vacina contra leishmaniose viscer al canina (LVC) pode ser uma importante ferramenta para o controle da leishmanio se visceral humana (LV). Além disso, para a obtenção de uma vacina efetiva, é fundamental o entendimento de eventos relacionados à imunogenicidade após a vacinação e desafio co m o agente etiológico para que seja definida a resposta imune associada à proteção contra a infecção por Leishmania. Assim, neste trabalho foi verificada a imunogenicidade e proteção conferida pela vacinação de cães contra LVC com os imunobiológicos LBSap e LBSapSal. Desta forma, trinta e cinco cães sem raça definida foram distribuídos em sete grupos experimentais, entre os quais: (i) grupo controle C (n = 5) que recebeu 1 mL de salina estéril a 0,9%; (ii) grupo LB (n = 5) que recebeu 600 μg de proteína de Leishmania braziliensis em 1 mL de salina estéril a 0,9%; (iii) grupo Sap (n = 5) que recebeu 1 mg de saponina em 1 mL de salina estéril a 0,9%; (iv) grupo LBSap (n = 5) que recebeu 600 μg de proteína de L. braziliensis associado a 1 mg de saponina em 1 mL de salina estéril a 0,9%; (v) grupo Sal (n = 5) que recebeu extrato de glândula salivar de Lutzomyia longipalpis (SGE) equivalente ao conteúdo de 5 ácinos da glândula salivar em 1 mL de salina estéril a 0,9%; (vi) grupo LBSal (n = 5) que recebeu 600 μg de promastigotas de L. braziliensis associado ao SGE em 1 mL de salina estéril a 0,9%; (vii) grupo LBSapSal (n = 5) que recebeu 600 μg de promastigotas de L. braziliensis associado a 1 mg de saponina e ao SGE em 1 mL de salina estéril a 0,9%. Cada animal recebeu três aplicações subcutâneas no flanco esquerdo em intervalos de quatro semanas. Posteriormente, os cães foram infectados por via intradérmica com 1x10 7 promastigotas de Leishmania (Leishmania) chagasi na presença de saliva de L. longipalpis, sendo avaliados 90 e 885 dias pós desafio. Os níveis de óxido nítrico (NO) e citocinas (IL-4, IL-10, TGF-, TNF-, IL-12, IFN-) foram avaliados em sobrenadantes de cultura de células mononucleares do sangue periférico (CMSP) após estímulo com antígeno solúvel vacinal (VSA) ou antígeno solúvel de L. chagasi (SLcA). O grupo LBSap apresentou altos níveis de IL-4 (T3 e T90dpd), IL-10 (885dpd) e baixos níveis de TGF- (885dpd) após estímulo com antígenos de Leishmania. Além disso, o grupo LBSapSal apresentou baixos níveis de IL-4 (90dpd) e TGF- (90 e 885dpd) após estímulo com antígeno de Leishmania. Adicionalment e, altos níveis de IL-12 foram observados nos grupos LBSap e LBSapSal (T3, 90dpd e 885dpd). Da mesma forma, aumento dos níveis de IFN- foram observados nos grupos LBSap (T3, 90dpd e 885dpd) e LBSapSal (T3, 90dpd e 885dpd). O perfil de citocinas sugere que as vacinas LBSap e LBSapSal podem conferir imunidade protetora contra a infecção por Leishmania. Os resultados obtidos a partir da análise dos níveis de NO confirmaram a hipótese de que as estas vacinas induzem um perfil de resistência contra a infecção por Leishmania. Os principais resultados do presente estudo apontam para uma forte imunogenicidade induzida pela vacinação de cães com LBSap e LBSapSal e submetidos ao desafio experimental com L. chagasi, indicando uma ação destes imunobiológicos compatível com controle do parasito em cães. Novos estudos de avaliação da carga parasitária em medula óssea através da PCR em tempo real podem fornecer importantes informações para um melhor entendimento da eficácia das vacinas LBSap e LBSapSal.Dogs represent the most important domestic reservoirs of L. chagasi (syn. L. infantum), and a vaccine against canine visceral leishmaniasis (CVL) would be an important tool in the control of human visceral leishmaniasis (VL) by decreasing dramatically the infect ion pressure of L. chagasi/L. infantum. Moreover, to obtain a successful vaccine, the understanding of immunogenicity mechanisms involved after vaccination and challenge is crucial to define the protective responses associated against Leishmania infection. Thus, in this work it was investigated the immunogenicity and protective effect of LBSap and LBSapSal vaccination in dogs. In this sense, thirty five mongrel dogs were treated within seven experimental groups as follow: (i) control group C (n = 5) received 1 mL of sterile 0.9% saline; (ii) LB group (n = 5) received 600 µg of Leishmania braziliensis promastigote protein in 1 mL sterile 0.9% saline; (iii) Sap group (n = 5) received 1 mg of saponin in 1 mL sterile 0.9% saline; (iv) LBSap group (n = 5) received 600 µg of L. braziliensis promastigote protein and 1 mg of saponin in 1 mL sterile 0.9% saline; (v) Sal group (n = 5) received sand fly gland extract (SGE) prepared from 5 acini of salivary glands of Lutzomyia longipalpis in 1 mL sterile 0.9% saline; (vi) LBSal group (n = 5) received 600 µg of L. braziliensis promastigote protein plus SGE in 1 mL sterile 0.9% saline; and (vii) the LBSapSal group (n = 5) received 600 µg of L. braziliensis promastigote protein plus 1 mg of saponin together with SGE in 1 mL sterile 0.9% saline. Each animal received three subcutaneous injections in the right flank at intervals of 4 weeks. Afterwards, the dogs were intradermally infected with 1×10 7 late-log-phase promastigotes of Leishmania (Leishmania) chagasi in the presence of sand fly saliva of L. longipalpis and follow-up at 90 and 885 days post challenge (dpc). Nitric oxide (NO) levels (determined as nitrite) and cytokines (IL-4, IL-10, TGF-, TNF-, IL-12, IFN-) were evaluated in culture supernatants of peripheral blood mononuclear cells (PBMC) stimulated by vaccine soluble antigen (VSA) or soluble Leishmania chagasi antigen (SLcA). LBSap group displayed higher levels of IL-4 (T3 and 90dpi), IL-10 (885dpi) and lower levels of TGF- (885dpi) following Leishmania stimulation. Moreover, LBSapSal group presented lower levels of IL-4 (90dpi) and TGF- (90 and 885dpi) after Leishmania stimulation. In addition, higher levels of IL-12 were observed in LBSap and LBSapSal groups (T3, 90 and 885dpi). Likewise, increased levels of IFN- were related to LBSap (T3, 90 and 885dpi) and LBSapSal (T3, 90 and 885dpi) groups. The cytokines profile suggest that LBSap and LBSapSal vaccine have potential protective immunity against Leishmania infection. The results obtained from the analysis of NO levels confirmed the hypothesis that these vaccines induce a potential resistance profile against Leishmania infection. In conclusion, the major findings in the present study point to a strong immunogenicity elicited when dogs were vaccinated with LBSap or LBSapSal and submitted to experimental challenge indicating a compatible action of this immunobiological with the effective control of the parasitism in dogs. Further studies on bone marrow parasitism using real time PCR may provide important information that will lead to a better understanding on LBSap and LBSapSal efficacies

Post-Ganglionic Horner’s Syndrome: An Unusual Presentation of Non-Hodgkin Lymphoma

In this paper, we present the rare case of a patient with cervical lymphadenopathy diagnosed as a T-cell-rich B-cell non-Hodgkin lymphoma that manifested Horner’s syndrome due to a post-ganglionic sympathetic neuron lesion caused by the tumor

Paracoccidioidomycosis in patients with Lymphoma and review of published literature

This paper describes four new cases of lymphomas, two Hodgkin lymphomas and two non-Hodgkin lymphomas in patients with paracoccidioidomycosis. All had mycosis diagnosed before lymphomas with Paracoccidioides brasiliensis demonstrated in several lymph nodes, as seen in the disseminated form of the disease. When lymphoma was diagnosed, one patient was under regular paracoccidioidomycosis treatment and in clinic-serological remission for this disease, another was under regular treatment but with clinic-serological mycosis activity, one had abandoned paracoccidioidomycosis treatment 6 years earlier, and the other had not yet received any kind of antifungal drugs. Three patients received treatment for lymphomas with one remaining in remission until now, one achieving tumor remission which relapsed years later, and one having only residual lymphoma in bone marrow for a decade but clinically well. All three experienced paracoccidioidomycosis clinical remission, however, serology became negative just in one. Similar previously described cases were reviewed: five Hodgkin lymphomas, three non-Hodgkin lymphomas, and one described only as "lymphoma" without specifying type; a summary of their findings is presented. Finally, there is also a brief discussion on the possible pathophysiological mechanisms involved in the concomitance of these two disorders.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Guidelines for morphological diagnosis of myelodysplastic syndromes

As síndromes mielodisplásicas são reconhecidas como doenças que se originam nas células-tronco da medula óssea e que requerem avaliação sistemática e criteriosa de sangue periférico e medula óssea para seu correto diagnóstico. O objetivo deste relato é estabelecer os critérios morfológicos (cito-histológicos) como parâmetros para o diagnóstico de SMD em amostras de sangue periférico e medula óssea, com especial direcionamento aos hematologistas e patologistas clínicos que exercem a hematologia laboratorial na sua rotina de trabalho. Os principais achados morfológicos são listados no final deste relato, na forma de check-list, objetivando a sistematização sobre estes achados.Myelodysplastic syndromes require both thorougly and systematic blood smear and bone marrow examinations. The main goal of this report is to establish criteria of the morphological ( cyto-histological) features, as parameters for the diagnosis of myelodysplastic syndromes ( MDS) from peripheral blood smears and bone marrow samples, with especial address to hematology and pathology practitioners. The main features are listed ( checklist) at the end of this report, in order to synthesize them

Chemical Composition and Antioxidant Activity of Essential Oils from Cinnamodendron dinisii Schwacke and Siparuna guianensis Aublet

The objectives of this study were to chemically characterize and evaluate the antioxidant activity of essential oils Cinnamodendron dinisii Schwacke (pepper) and Siparuna guianensis Aublet (negramina). The essential oil was isolated by hydrodistillation using a Clevenger modified apparatus, and the identification and quantification of constituents, through GC/MS and GC-FID analysis. The antioxidant activity was evaluated using β-carotene/linoleic acid system and the DPPH radical sequestering method. In chromatographic analysis, the majority constituents found in the essential oil of C. dinisii were bicyclic monoterpenes, α-pinene (35.41%), β-pinene (17.81%), sabinene (12.01%) and sesquiterpene bicyclogermacrene (7.59%). In the essential oil of the fresh leaves of Siparuna guianensis Aublet, acyclic monoterpene, β-myrcene (13.14%), and sesquiterpenes, germacrene-D (8.68%) and bicyclogermacrene (16.71%) were identified. The antioxidant activity was low by the β-carotene/linoleic acid test and was not evidenced by the DPPH test, for both oils evaluated

Analysis using canine peripheral blood for establishing in vitro conditions for monocyte differentiation into macrophages for Leishmania chagasi infection and T-cell subset purification.

Canine visceral leishmaniasis (CVL) is a parasitic disease endemic in many countries, anddogs present as the major natural reservoir of the parasite, Leishmania chagasi (syn. L.infantum). Biomarkers in the canine immune system is an important technique in thecourse of developing vaccines and treatment strategies against CVL. New methodologiesfor studying the immune response of dogs during Leishmania infection and after receivingvaccines and treatments against CVL would be useful. In this context, we used peripheralblood mononuclear cells (PBMCs) from healthy dogs to evaluate procedures related to (i)establishment of in vitro conditions of monocytes differentiated into macrophages infectedwith L. chagasi and (ii) purification procedures of T-cell subsets (CD4+and CD8+) usingmicrobeads. Our data demonstrated that after 5 days of differentiation, macrophages wereable to induce significant phagocytic and microbicidal activity after L. chagasi infectionand also showed increased frequency of parasitism and a higher parasite load. Although N-acetyl- _-d-glucosaminidase (NAG) levels presented similar levels of macrophage cultureand L. chagasi infection, a progressive decrease in myeloperoxidase (MPO) levels was ahallmark over 5 days of culture. High purity levels (>90%) of CD4 and CD8 T cells wereobtained on a magnetic separation column. We concluded that monocytes differentiatedinto macrophages at 5 days and displayed an intermediate frequency of parasitism andparasite load 72 h after L. chagasi infection. Furthermore, the purification system usingcanine T-lymphocyte subsets obtained after 5 days of monocyte differentiation provedefficient for CD4 or CD8 T-cell purification (≥90%). The in vitro analysis using L. chagasi-infected macrophages and purified T cells presented a prospective methodology that couldbe incorporated in CVL vaccine and treatment studies that aim to analyze the microbicidalpotential induced by specific CD4+and/or CD8+T cells

Multicomponent LBSap vaccine displays immunological and parasitological profiles similar to those of Leish-Tec® and Leishmune® vaccines against visceral leishmaniasis

Submitted by Nuzia Santos ([email protected]) on 2017-07-06T18:40:38Z No. of bitstreams: 1 Mendonça_Ludmila_Multicomponent LBSap_2016 .pdf: 12533394 bytes, checksum: a405111aabb97116bbbbf1a5403174c5 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2017-07-06T18:48:28Z (GMT) No. of bitstreams: 1 Mendonça_Ludmila_Multicomponent LBSap_2016 .pdf: 12533394 bytes, checksum: a405111aabb97116bbbbf1a5403174c5 (MD5)Made available in DSpace on 2017-07-06T18:48:28Z (GMT). No. of bitstreams: 1 Mendonça_Ludmila_Multicomponent LBSap_2016 .pdf: 12533394 bytes, checksum: a405111aabb97116bbbbf1a5403174c5 (MD5) Previous issue date: 2016Universidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, Brazil/Universidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas. Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, Brazil/Universidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas. Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas. Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas. Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Ouro Preto. Programa de Pós-Graduação de Ciências Farmacêuticas. Laboratório de Pesquisas Clínicas. Ouro Preto, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Biomarcadores de Diagnóstico e Monitoração. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, BrazilLaboratório de Genômica de Parasitos. Belo Horizonte, MG, BrazilUniversidade Federal de Ouro Preto. Programa de Pós-Graduação de Ciências Farmacêuticas. Laboratório de Pesquisas Clínicas. Ouro Preto, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Biomarcadores de Diagnóstico e Monitoração. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Imunologia Celular e Molecular. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, BrazilUniversidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas. Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, BrazilBackground: In past years, many researchers have sought canine visceral leishmaniasis (CVL) prevention through the characterization of Leishmania antigens as vaccine candidates. Despite these efforts, there is still no efficient vaccine for CVL control. Methods: In the present study, we performed a pre-clinical vaccine trial using BALB/c mice to compare the effects of the multicomponent LBSap vaccine with those of Leish-Tec® and Leishmune®. Blood was collected to determine the frequency of peripheral blood cells and to evaluate hematologic and immunophenotypic parameters. Liver and spleen samples were collected for parasitological quantification, and spleen samples were used to access the cytokine profile. Results: When measuring total IgG and IgG1 anti-Leishmania levels after the third vaccination and L. infantum challenge, it was evident that all vaccines were able to induce humoral immune response. Regarding the innate immune response, increased levels of NK CD3-CD49+ cells were the hallmark of all vaccinated groups, whereas only the Leish-Tec® group displayed a high frequency of CD14+ monocytes after L. infantum challenge. Moreover, CD3+CD4+ T cells were the main circulating lymphocytes induced after L. infantum challenge with all evaluated vaccines. Importantly, after L. infantum challenge, splenocytes from the Leishmune® vaccine produced high levels of IL-2, whereas a prominent type 1 immune response was the hallmark of the LBSap vaccine, which presented high levels of IL-2, IL-6, TNF-α, and IFN-γ. The efficacy analysis using real-time polymerase chain reaction demonstrated a reduction in the parasitism in the spleen (Leishmune®: 64 %; LBSap: 42 %; and Leish-Tec®: 36 %) and liver (Leishmune®: 71 %; LBSap: 62 %; and Leish-Tec®: 48 %). Conclusions: The dataset led to the conclusion that the LBSap vaccination was able to induce immune and efficacy profiles comparable with those of commercial vaccines, thus demonstrating its potential as a promising vaccine candidate for visceral leishmaniasis control

Cytokine and nitric oxide patterns in dogs immunized with LBSap vaccine, before and after experimental challenge with Leishmania chagasi plus saliva of Lutzomyia longipalpis.

In the studies presented here, dogs were vaccinated against Leishmania (Leishmania) cha-gasi challenge infection using a preparation of Leishmania braziliensis promastigote proteinsand saponin as adjuvant (LBSap). Vaccination with LBSap induced a prominent type 1immune response that was characterized by increased levels of interleukin (IL-) 12 andinterferon gamma (IFN- _) production by peripheral blood mononuclear cells (PBMC) uponstimulation with soluble vaccine antigen. Importantly, results showed that this type ofresponsiveness was sustained after challenge infection; at day 90 and 885 after L. chagasichallenge infection, PBMCs from LBSap vaccinated dogs produced more IL-12, IFN- _ andconcomitant nitric oxide (NO) when stimulated with Leishmania antigens as comparedto PBMCs from respective control groups (saponin, LB- treated, or non-treated controldogs). Moreover, transforming growth factor (TGF)- _ decreased in the supernatant of SLcA-stimulated PBMCs in the LBSap group at 90 days. Bone marrow parasitological analysisrevealed decreased frequency of parasitism in the presence of vaccine antigen. It is con-cluded that vaccination of dogs with LBSap vaccine induced a long-lasting type 1 immuneresponse against L. chagasi challenge infection

Impact of LbSapSal Vaccine in Canine Immunological and Parasitological Features before and after Leishmania chagasi-Challenge

Submitted by Nuzia Santos ([email protected]) on 2017-03-02T17:14:04Z No. of bitstreams: 1 ve_Resende_Lucilene_Impact of LbSapSal_CPqRR_2016.pdf: 7982893 bytes, checksum: b3d67b0ad26288c3a2b36cb1f1a24ab5 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2017-03-02T17:21:02Z (GMT) No. of bitstreams: 1 ve_Resende_Lucilene_Impact of LbSapSal_CPqRR_2016.pdf: 7982893 bytes, checksum: b3d67b0ad26288c3a2b36cb1f1a24ab5 (MD5)Made available in DSpace on 2017-03-02T17:21:03Z (GMT). No. of bitstreams: 1 ve_Resende_Lucilene_Impact of LbSapSal_CPqRR_2016.pdf: 7982893 bytes, checksum: b3d67b0ad26288c3a2b36cb1f1a24ab5 (MD5) Previous issue date: 2016Fundação de Amparo a Pesquisa do Estado de Minas Gerais. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, Brazil/Universidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas/NUPEB Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas/NUPEB Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas/NUPEB Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas/NUPEB Laboratório de Imunopatologia. Ouro Preto, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Biomarcadores de Diagnóstico e Monitoração. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Imunologia Celular e Molecular. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Biomarcadores de Diagnóstico e Monitoração. Belo Horizonte, MG, BrazilFundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Biomarcadores de Diagnóstico e Monitoração. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Parasitologia. Laboratório de Imunologia e Genômica de Parasitos. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Parasitologia. Laboratório de Fisiologia de Insetos Hematófagos. Belo Horizonte, MG, BrazilUniversidade Federal de Ouro Preto. Instituto de Ciências Exatas e Biológicas. Núcleo de Pesquisas em Ciências Biológicas/NUPEB Laboratório de Imunopatologia. Ouro Preto, MG, Brazil/Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Imunologia Celular e Molecular. Belo Horizonte, MG, BrazilUniversidade Federal de Minas Gerais. Departamento de Morfologia. Laboratório de Biologia das Interações Celulares. Belo Horizonte, MG, Brazil/Fundação Oswaldo Cruz. Centro de Pesquisa René Rachou. Laboratório de Biomarcadores de Diagnóstico e Monitoração. Belo Horizonte, MG, BrazilDogs represent the most important domestic reservoir of L. chagasi (syn. L. infantum). A vaccine against canine visceral leishmaniasis (CVL) would be an important tool for decreasing the anxiety related to possible L. chagasi infection and for controlling human visceral leishmaniasis (VL). Because the sand fly salivary proteins are potent immunogens obligatorily co-deposited during transmission of Leishmania parasites, their inclusion in an anti-Leishmania vaccine has been investigated in past decades. We investigated the immunogenicity of the "LbSapSal" vaccine (L. braziliensis antigens, saponin as adjuvant, and Lutzomyia longipalpis salivary gland extract) in dogs at baseline (T0), during the post-vaccination protocol (T3rd) and after early (T90) and late (T885) times following L. chagasi-challenge. Our major data indicated that immunization with "LbSapSal" is able to induce biomarkers characterized by enhanced amounts of type I (tumor necrosis factor [TNF]-α, interleukin [IL]-12, interferon [IFN]-γ) cytokines and reduction in type II cytokines (IL-4 and TGF-β), even after experimental challenge. The establishment of a prominent pro-inflammatory immune response after "LbSapSal" immunization supported the increased levels of nitric oxide production, favoring a reduction in spleen parasitism (78.9%) and indicating long-lasting protection against L. chagasi infection. In conclusion, these results confirmed the hypothesis that the "LbSapSal" vaccination is a potential tool to control the Leishmania chagasi infection