EFAR - Escola de Farmácia

URI permanente desta comunidadehttp://www.hml.repositorio.ufop.br/handle/123456789/451

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O curso de Farmácia em Ouro Preto foi criado em 1839, sendo a mais antiga Escola de Farmácia da América Latina.

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Kinetics of cell migration to the dermis and hypodermis in dogs vaccinated with antigenic compounds of Leishmania braziliensis plus saponin.
(2008) Souza, Juliana Vitoriano de; Reis, Alexandre Barbosa; Moreira, Nádia das Dores; Giunchetti, Rodolfo Cordeiro; Oliveira, Rodrigo Corrêa de; Carneiro, Cláudia Martins
The search for new immunobiologicals against canine visceral leishmaniasis (CVL) has intensified in the last decade. However, it still remains to be elucidated that mechanisms of the innate immune response in situ after immunization (a.i.). The aim of this study was to investigate the kinetics of cell migration in the skin dogs with distinct antigenic compounds of the LBSap vaccine. Our major findings indicated that saponin adjuvant alone or combined with Leishmania braziliensis antigen induced strong local acute inflammatory reaction. However, these reactions not progressed to ulcerated lesions. Overall, the cell profile found in Sap and LBSap was composed of neutrophils, lymphocytes and eosinophils. There was also increased production of iNOS in Sap and LBSap groups. Thus, we can conclude that dogs immunized by LBSap and the saponin adjuvant elicited a potential innate-immune activations status compatible with effective control of the resistance to infection by Leishmania and contributing to a better understanding of the innate-immunity events induced by the LBSap vaccine.
Systemic and compartmentalized immune response in canine visceral leishmaniasis.
(2009) Reis, Alexandre Barbosa; Martins Filho, Olindo Assis; Carvalho, Andréa Teixeira de; Giunchetti, Rodolfo Cordeiro; Carneiro, Cláudia Martins; Mayrink, Wilson; Tafuri, Washington Luiz; Oliveira, Rodrigo Corrêa de
Human visceral leishmaniasis (VL) and canine visceral leishmaniasis (CVL) are the most important emerging diseases with high prevalence in Latin American countries and are mainly caused by Leishmania (L.) chagasi (Syn = L. infantum). CVL has a great impact on Brazilian public health because domestic dogs are the most important VL peri-domicile reservoirs in both urban and peri-urban areas. Our findings highlight the complexity of cellular immunological events related to the natural infection from dogs by L. chagasi, additionally correlating major peripheral blood phenotypic markers with clinical status and tissues parasite density. Our main results demonstrated that lower frequency ofcirculating B cells and monocytes are important markers of severe CVL, whereas increased levels of CD8+ lymphocytes appear to be the major phenotypic feature of asymptomatic disease. Determination of the isotypes patterns during CVL demonstrated thatasymptomatic dogs and those with low parasitism are associated with an increase of IgG1, while the symptomatic dogs and those with high parasitism are associated with an increase of IgG, IgG2, IgM, IgA and IgE immunoglobulins. Pioneer findings obtained by our group showed a correlation between clinical status of CVL with degree of tissue parasite density. This data demonstrated that asymptomatic dogs presented low parasitism while symptomatic dogs are associated with high parasite load in various tissues such as skin, bone marrow and spleen. We have also investigated the association between tissue parasitism and CVL clinical forms. Regardless of clinical status, skin and spleen are the major sites of high parasite density during ongoing CVL. Furthermore, we demonstrated that bone marrow and spleen parasite density are the most reliable parasitological markers to decode the clinical status of CVL. In this article, we have reviewed some aspectsof the histopathological and immunological events occurring in natural and experimentalL. chagasi/L. infantum infection, pointing out the main L. chagasi-parasitized tissue. Wehave discussed the importance of the association between parasite density, immunological/ histopathological aspects and clinical status of the CVL, their current applications, challenges for the future and potential opportunities in CVL research.
Effective immunotherapy against canine visceral leishmaniasis with the FML-vaccine.
(2004) Borja Cabrera, Gulnara Patricia; Mendes, Amanda Cruz; Souza, Edilma Paraguai de; Okada, Lilian Y. Hashimoto; Trivellato, Fernando Antonio de Assis; Kawasaki, Jarbas Kiyoshi Alves; Costa, Andreia Cerqueira; Reis, Alexandre Barbosa; Genaro, Odair; Batista, Leopoldina Maria Melo; Palatnik, Marcos; Souza, Clarisa Beatriz Palatnik de
The potential effect of the fucose mannose ligand (FML)-vaccine on immunotherapy of canine visceral leishmaniasis was assayed on five mongrel dogs experimentally infected withLeishmania donovani and on 21 Leishmania chagasi naturally infected dogs when seropositive to FML but completely asymptomatic. The clinical signs of the experimentally infected, symptomatic dogs only disappeared after the complete vaccination. Protection was obtained in 3/5 animals that remained asymptomatic, IDR positive and parasite free, 1 year after infection. Furthermore, the asymptomatic, FML-vaccine treated dogs showed stable anti-FML IgG1 levels, increasing IgG2 levels and 79–95% of positive DTH response, during the whole experiment. Twenty-two months after complete vaccination, no obits due to visceral leishmaniasis were recorded and 90% of these dogs were still asymptomatic, healthy and parasite free. On the other hand, 37% (17/46 dogs) kala-azar obits were recorded in a control group that received no treatment during the same period, and that was FML-seropositive and asymtpomatic at the beginning of the assay. Our results indicate that the FML-vaccine was effective in the immunotherapy against visceral leishmaniasis of asymptomatic infected dogs. Normal proportions of CD4 and CD21 lymphocytes were detected in PBMC by FACS analysis, in dogs submitted to immunotherapy, suggesting their non-infectious condition. All animals showed as well significantly increased percents of CD8 lymphocytes as expected for Quillajasaponin (QuilA) vaccine treatments.
Evaluation of enzyme-linked immunosorbent assay using crude Leishmania and recombinant antigens as a diagnostic marker for canine visceral leishmaniasis.
(2005) Rosário, Eliza Yoshie do; Genaro, Odair; Silva, João Carlos França da; Costa, Roberto Teodoro da; Mayrink, Wilson; Reis, Alexandre Barbosa; Carneiro, Mariângela
The performances of ELISA assays with different antigen preparations, such as Leishmania amazonensis or L. chagasi lysates and the recombinant antigens rK-39 and rK-26, were compared using sera or eluates from dried blood collected on filter paper to detect anti-Leishmania antibodies in dogs from a visceral leishmaniasis-endemic area in Brazil. Of 115 IFAT-reactive dogs at 1:40 titre, 106 (92.2%) were positive in parasitological exams (skin and/or spleen). These animals were compared to healthy animals (n = 25), negative for IFAT at a titre of 1:40 and parasitological exams. The sensitivities of crude and recombinant antigens were similar and remarkably high for both sera and eluates (97-100%). Specificity was higher than 96% for sera and eluates for different antigens, except for L. chagasi antigen using eluates (88%). Concordance values among the tests were higher either for sera or eluates (J = 0.95-1.00). High concordances were observed between sera and eluates tested with different antigens (kappa = 0.93-0.97). Crude and recombinant antigens identified different clinical phases of canine leishmaniasis. These results show that eluates could be used in canine surveys to identify L. chagasi infection. Recombinant antigens added little when compared to crude antigen in identifying positive dogs. Cross-reactivity with other diseases whose distribution often overlaps VL-endemic areas is a limitation of crude antigen use however.
Isotype patterns of immunoglobulins : hallmarks for clinical status and tissue parasite density in brazilian dogs naturally infected by Leishmania (Leishmania) chagasi.
(2006) Reis, Alexandre Barbosa; Carvalho, Andréa Teixeira de; Vale, André Macedo; Marques, Marcos José; Giunchetti, Rodolfo Cordeiro; Mayrink, Wilson; Guerra, Luanda Liboreiro; Andrade, Renata Aline de; Oliveira, Rodrigo Corrêa de; Martins Filho, Olindo Assis
The role of anti-leishmanial immune response underlying the susceptibility/resistance during canine visceral leishmaniasis (CVL) has been recognized throughout ex vivo and in vitro investigations. Recently, we demonstrated that immunoglobulin levels (Igs), as well as the parasite load are relevant hallmarks of distinct clinical status of CVL. To further characterize and upgrade the background on this issue, herein, we have evaluated, inLeishmania ( Leishmania ) chagasinaturally infected dogs, the relationship between tissue parasitism (skin, bone marrow, spleen, liver and lymph node), the CVL clinical status (asymptomatic (AD), with no suggestive signs of the disease; oligosymptomatic (OD), with maximum three clinical signs—opaque bristles; localized alopecia and moderate loss of weight; symptomatic (SD), serologically positive with severe clinical signs of visceral leishmaniasis), and the humoral immunological profile of anti-Leishmania immunoglobulins (IgG, IgG1, IgG2, IgM, IgA and IgE). Our major statistically significant findings revealed distinct patterns of tissue parasite density within L. chagasi-infected dogs despite their clinical status, pointing out the spleen and skin as the most relevant sites of high parasitism during ongoing CVL. Parasite density of bone marrow and spleen were the most reliable parasitological markers to decode the clinical status of CVL. Moreover, the parasite density of bone marrow better correlates with most anti- Leishmania Igs reactivity. Additionally, a prognostic hallmark for canine visceral leishmaniasis was found, highlighting strong correlation between IgG1 and asymptomatic disease, but with IgA, IgE and IgG2 displaying better association with symptomatic disease. The new aspects of this study highlighted pioneer findings that correlated the degree of tissue parasite density (low (LP), medium (MP) and high (HP) parasitism) with distinct patterns of anti- Leishmania Igs reactivity. In this scope, our data re-enforce the anti- Leishmania IgG but with IgA reactivity as the better marker for overall tissue parasitism. The association between clinical status, Ig profile and the tissue parasitism support a novel investigation on the impact of humoral immune response and susceptibility/resistance mechanism during ongoing CVL
Parasite density and impaired biochemical/hematological status are associated with severe clinical aspects of canine visceral leishmaniasis.
(2006) Reis, Alexandre Barbosa; Martins Filho, Olindo Assis; Carvalho, Andréa Teixeira de; Carvalho, Maria das Graças; Mayrink, Wilson; Silva, João Carlos França da; Giunchetti, Rodolfo Cordeiro; Genaro, Odair; Oliveira, Rodrigo Corrêa de
We have performed a detailed investigation in 40 dogs naturally infected withLeishmania infantum ( syn. chagasi), subdivided into three groups: asymptomatic (AD = 12), oligosymptomatic (OD = 12) and symptomatic (SD = 16), based on their clinical features. Twenty non-infected dogs (CD) were included as control group. Serological analysis, performed by IFAT and ELISA, demonstrated higher antibodies titers in SD in comparison to the AD. A positive correlation was found between parasite density in the spleen and skin smears as well as the bone marrow parasitism with clinical status of the infection. We observed that the progression of the disease from asymptomatic to symptomatic clinical form was accompanied by intense parasitism in the bone marrow. It is likely that this led to the impaired biochemical/hematological status observed. Finally, we believe that the follow-up of these parameters could be a relevant approach to be used as markers during therapeutic and vaccine evaluations.
Immunotherapy against experimental canine visceral leishmaniasis with the saponin enriched-Leishmune ® vaccine.
(2007) Santos, Fernanda Nunes; Borja Cabrera, Gulnara Patricia; Myashiro, L. M.; Grechi, Juliana; Reis, Alexandre Barbosa; Moreira, Márcio Antônio Batistela; Martins Filho, Olindo Assis; Luvizotto, Maria Cecília Rui; Menz, Ingrid; Pessôa, L. M.; Gonçalves, Pablo Rodrigues; Palatnik, Marcos; Souza, Clarisa Beatriz Palatnik de
In order to assess the immunotherapeutic potential on canine visceral leishmaniasis of the Leishmune ® vaccine, formulated with an increased adjuvant concentration (1 mg of saponin rather than 0.5 mg), 24 mongrel dogs were infected with Leishmania (L.) chagasi . The enriched-Leishmune ® vaccine was injected on month 6, 7 and 8 after infection, when animals were seropositive and symptomatic. The control group were injected with a saline solution. Leishmune ® -treated dogs showed significantly higher levels of anti-FML IgG antibodies (ANOVA; p < 0.0001), a higher and stable IgG2 and a decreasing IgG1 response, pointing to a TH1 T cell mediated response. The vaccine had the following effects: it led to more positive delayed type hypersensitivity reactions against Leishmania lysate in vaccinated dogs (75%) than in controls (50%), to a decreased average of CD4+ Leishmania -specific lymphocytes in saline controls (32.13%) that fell outside the 95% confidence interval of the vaccinees (41.62%, CI95% 43.93–49.80) and an increased average of the clinical scores from the saline controls (17.83) that falls outside the 95% confidence interval for the Leishmune ® immunotherapy-treated dogs (15.75, CI95% 13.97–17.53). All dogs that received the vaccine were clustered, and showed lower clinical scores and normal CD4+ counts, whereas 42% of the untreated dogs showed very diminished CD4+ and higher clinical score. The increase in clinical signs of the saline treated group was correlated with an increase in anti-FML antibodies (p < 0.0001), the parasitological evidence ( p = 0.038) and a decrease inLeishmania -specific CD4+ lymphocyte proportions (p = 0.035). These results confirm the immunotherapeutic potential of the enriched-Leishmune ® vaccine. The vaccine reduced the clinical symptoms and evidence of parasite, modulating the outcome of the infection and the dog’s potential infectiosity to phlebotomines. The enriched-Leishmune ® vaccine was subjected to a safety analysis and found to be well tolerated and safe
Immunogenicity of a killed Leishmania vaccine with saponin adjuvant in dogs.
(2007) Giunchetti, Rodolfo Cordeiro; Oliveira, Rodrigo Corrêa de; Martins Filho, Olindo Assis; Carvalho, Andréa Teixeira de; Roatt, Bruno Mendes; Soares, Rodrigo Dian de Oliveira Aguiar; Souza, Juliana Vitoriano de; Moreira, Nádia das Dores; Malaquias, Luiz Cosme Cotta; Castro, Luciana Lisboa Mota e; Lana, Marta de; Reis, Alexandre Barbosa
Cellular and humoral immune responses of dogs to a candidate vaccine, composed of Leishmania braziliensis promastigote protein plus saponin as adjuvant, have been investigated as a pre-requisite to understanding the mechanisms of immunogenicity against canine visceral leishmaniasis (CVL). The candidate vaccine elicited strong antigenicity related to the increases of anti- Leishmania IgG isotypes, together with higher levels of lymphocytes, particularly of circulating CD8 + T-lymphocytes and Leishmania chagasi antigen-specific CD8 + T-lymphocytes. As indicated by the intense cell proliferation and increased nitric oxide production during in vitro stimulation by L. chagasisoluble antigens, the candidate vaccine elicited an immune activation status potentially compatible with effective control of the etiological agent of CVL.
Clinical value of anti-Leishmania (Leishmania) chagasi IgG titers detected by flow cytometry to distinguish infected from vaccinated dogs.
(2007) Andrade, Renata Aline de; Reis, Alexandre Barbosa; Gontijo, Célia Maria Ferreira; Braga, Lidiane Bento; Rocha, Roberta Dias Rodrigues; Araújo, Márcio Sobreira Silva; Vianna, Leonardo Rocha; Martins Filho, Olindo Assis
Leishmune® vaccination covers a broader number of endemic areas of canine visceral leishmaniasis (CVL) and therefore the development of new serological devices able to discriminate CVL from Leishmune® vaccinees becomes an urgent need considering the post-vaccine seroconversion detected throughout conventional methodologies. Herein, we have described the establishment of a flow cytometry based methodology to detect anti-fixedL. ( L. ) chagasipromastigotes antibodies (FC-AFPA-IgG, FC-AFPA-IgG1 and FC-AFPA-IgG2) in sera samples from Leishmania ( Leishmania ) chagasiinfected dogs and Leishmune® vaccinees. The results of FC-AFPA were reported along the sera titration curve (1:128–1:524,288), as percentage-of-positive-fluorescent-parasite (PPFP). The use of PPFP = 20% as a cut-off edge to segregate negative and positive results at sera dilution 1:2048 revealed outstanding performance indexes that elect FC-AFPA-IgG and IgG2 (both detected by polyclonal FITC-labeled second step reagent) applicable to the serological diagnosis of CVL, with 100% of specificity for both IgG and IgG2 and 97 and 93% of sensitivity, respectively. Moreover, FC-AFPA-IgG, applied at sera dilution 1:2048, also appeared as a useful tool to discriminate L. chagasiinfected dogs from Leishmune® vaccinees, with 76% of specificity. Outstanding likelihood indexes further support the performance of FC-AFPA-IgG for exclusion diagnosis of CVL in Leishmune® vaccinees. Analysis of FC-AFPA-IgG at sera dilution 1:8192 revealed the most outstanding indexes, demonstrating that besides the ability of PPFP 20% to exclude the diagnosis of CVL, a PPFP values higher 80%, mostly observed for infected dogs (INF) have a minimal change to come from a non-infected animal (NI) or Leishmune®.
Despite Leishvaccine and Leishmune ® trigger distinct immune profiles, their ability to activate phagocytes and CD8 + T-cells support their high-quality immunogenic potential against canine visceral leishmaniasis.
(2008) Araújo, Márcio Sobreira Silva; Andrade, Renata Aline de; Vianna, Leonardo Rocha; Mayrink, Wilson; Reis, Alexandre Barbosa; Avelar, Renato Sathler; Carvalho, Andréa Teixeira de; Andrade, Mariléia Chaves; Melo, Maria Norma; Martins Filho, Olindo Assis
Phenotypic features of peripheral blood leukocytes have been investigated as a prerequisite to characterize the protective immunity attributed to both Leishvaccine and Leishmune ® . Our results showed that either those vaccine were accompanied by distinct profiles on innate immune compartment. While Leishvaccine promoted early changes in phenotypic fea-tures of neutrophils and eosinophils with late involvement of monocytes, Leishmune ® induced early and persistent activation of neutrophils and monocytes, without changes on eosinophil activation status. Regarding the adaptive immunity, Leishvaccine sponsored a mixed profile, associated with phenotypic changes of T and B-lymphocytes. Major phenotypic changes in CD4 + T-cells with transient activation of CD8 + T-cell, besides decreased frequency of B-cell expressing

EFAR - Escola de Farmácia

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