Efficacy of a diarylheptanoid derivative against Leishmania amazonensis

The activity of several diarylheptanoid derivatives (curcuminoids) was previously evaluated against Leishmania amazonensis   promastigotes and among them the most active compound was the [1-(4-methoxy-phenyl)-7-(3,4-methoxy-4-hydroxy-phenyl)-1,6-heptadien-3, 5-dione]. This derivative was chosen to be assayed in vivo in a treatment trial. For these experiments, the curcuminoid compound was used in a concentration equivalent to the IC50/24 h, obtained from the previous study. Balb/c mice were inoculated subcutaneously in the footpad with L. amazonensis infective promastigotes and 4 weeks after the inoculation, the animals were treated with different schemes, varying from 1 to 3 doses. In all the experiments, Pentamidine Isethionate was used as reference drug under the same experimental conditions. The results showed that one dose was not enough to heal the lesion, however, with 2 and 3 doses the efficiency of the assayed compound was clear. On the other hand, treatment with Pentamidine Isethionate using the three different schemes was not satisfactory when compared to the curcuminoid derivative

Kinetoplastid membrane protein-11 as a vaccine candidate and a virulence factor in Leishmania

Submitted by sandra infurna ([email protected]) on 2016-04-14T14:57:40Z No. of bitstreams: 1 sergio2_mendonça_etal_IOC_2015.pdf: 164599 bytes, checksum: 2fb52f9528782857eccf99e933467e50 (MD5)Approved for entry into archive by sandra infurna ([email protected]) on 2016-04-14T15:10:57Z (GMT) No. of bitstreams: 1 sergio2_mendonça_etal_IOC_2015.pdf: 164599 bytes, checksum: 2fb52f9528782857eccf99e933467e50 (MD5)Made available in DSpace on 2016-04-14T15:10:57Z (GMT). No. of bitstreams: 1 sergio2_mendonça_etal_IOC_2015.pdf: 164599 bytes, checksum: 2fb52f9528782857eccf99e933467e50 (MD5) Previous issue date: 2015Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunoparasitologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunoparasitologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto de Tecnologia em Imunobiológicos. Laboratório de Tecnologia Imunológica. Rio de Janeiro, RJ, Brasil.Kinetoplastid membrane protein-11 (KMP-11), a protein present in all kinetoplastid protozoa, is considered a potential candidate for a leishmaniasis vaccine. In Leishmania amazonensis, KMP-11 is expressed in promastigotes and amastigotes. In both stages, the protein was found in association with membrane structures at the cell surface, flagellar pocket, and intracellular vesicles. More importantly, its surface expression is higher in amastigotes than in promastigotes and increases during metacyclogenesis. The increased expression of KMP-11 in metacyclic promastigotes, and especially in amastigotes, indicates a role for this molecule in the parasite relationship with the mammalian host. In this connection, we have shown that addition of KMP-11 exacerbates L. amazonensis infection in peritoneal macrophages from BALB/c mice by increasing interleukin (IL)-10 secretion and arginase activity while reducing nitric oxide production. The doses of KMP-11, the IL-10 levels, and the intracellular amastigote loads were strongly, positively, and significantly correlated. The increase in parasite load induced by KMP-11 was inhibited by anti-KMP-11 or anti-IL-10-neutralizing antibodies, but not by isotype controls. The neutralizing antibodies, but not the isotype controls, were also able to significantly decrease the parasite load in macrophages cultured without the addition of KMP-11, demonstrating that KMP-11-induced exacerbation of the infection is not dependent on the addition of exogenous KMP-11 and that the protein naturally expressed by the parasite is able to promote it. All these data indicate that KMP-11 acts as a virulence factor in L. amazonensis infection

Exploring the Association of Surface Plasmon Resonance with Recombinant MHC:Ig Hybrid Protein as a Tool for Detecting T Lymphocytes in Mice Infected with Leishmania (Leishmania) amazonensis

Submitted by Sandra Infurna ([email protected]) on 2017-07-06T15:09:57Z No. of bitstreams: 1 franklin_silva_etal_IOC_2017.pdf: 2955041 bytes, checksum: 6c9e52b90361d7bf859cd01c83269b34 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2017-07-06T15:29:06Z (GMT) No. of bitstreams: 1 franklin_silva_etal_IOC_2017.pdf: 2955041 bytes, checksum: 6c9e52b90361d7bf859cd01c83269b34 (MD5)Made available in DSpace on 2017-07-06T15:29:06Z (GMT). No. of bitstreams: 1 franklin_silva_etal_IOC_2017.pdf: 2955041 bytes, checksum: 6c9e52b90361d7bf859cd01c83269b34 (MD5) Previous issue date: 2017Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil / Universidade Federal do Rio Grande do Norte. Centro de Ciências da Saúde. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Laboratório de Imunologia Clínica. Natal, RN, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. BrasilFundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunoparasitologia. Rio de Janeiro, RJ. Brasil.Universidade Federal do Rio Grande do Norte. Centro de Ciências da Saúde. Faculdade de Farmácia. Departamento de Análises Clínicas e Toxicológicas. Laboratório de Imunologia Clínica. Natal, RN, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.A surface plasmon resonance- (SPR-) based recognition method applying H-2 L(d):Ig/peptides complexes for ex vivo monitoring cellular immune responses during murine infection with Leishmania (Leishmania) amazonensis is described. Lymphocytes from lesion-draining popliteal lymph nodes were captured on a carboxylated sensor chip surface previously functionalized with H-2 L(d):Ig (DimerX) protein bound to synthetic peptides derived from the COOH-terminal region of cysteine proteinase B of L. (L.) amazonensis. In computational analysis, these peptides presented values of kinetic constants favorable to form complexes with H-2 L(d) at neutral pH, with a Gibbs free energy ΔG° < 0. The assayed DimerX:peptide complexes presented the property of attaching to distinct T lymphocytes subsets, obtained from experimentally infected BALB/c mice, in each week of infection, thus indicating a temporal variation in specific T lymphocytes populations, each directed to a different COOH-terminal region-derived peptide. The experimental design proposed herein is an innovative approach for cellular immunology studies of a neglected disease, providing a useful tool for the analysis of specific T lymphocytes subsets

Serine Proteinases in Leishmania (Viannia) braziliensis Promastigotes Have Distinct Subcellular Distributions and Expression

Submitted by Sandra Infurna ([email protected]) on 2019-09-10T15:40:28Z No. of bitstreams: 1 RaquelSOuza_LuziaCortes_etal_IOC_2019.pdf: 3159744 bytes, checksum: 66d8b1d671d06bcd20df2ff676c76c9f (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2019-09-10T15:47:42Z (GMT) No. of bitstreams: 1 RaquelSOuza_LuziaCortes_etal_IOC_2019.pdf: 3159744 bytes, checksum: 66d8b1d671d06bcd20df2ff676c76c9f (MD5)Made available in DSpace on 2019-09-10T15:47:43Z (GMT). No. of bitstreams: 1 RaquelSOuza_LuziaCortes_etal_IOC_2019.pdf: 3159744 bytes, checksum: 66d8b1d671d06bcd20df2ff676c76c9f (MD5) Previous issue date: 2019Fundação Oswaldo Cruz, Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz, Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz, Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunoparasitologia. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz, Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ, Brasil / Fundação Oswaldo Cruz. Centro de Desenvolvimento Tecnológico em Saúde. Rio de Janeiro, RJ, Brasil.Serine proteinases in Leishmania (Viannia) braziliensis promastigotes were assessed in this work. This study included the investigation of the enzymatic activity of subcellular fractions obtained from benzamidine affinity chromatography, reverse transcription polymerase chain reactions, and in silico assays of subcellular localization of subtilisin. Promastigote serine proteinases showed gelatinolytic activity with molecular masses of 43 kDa to 170 kDa in the cytosolic fraction and 67 kDa to 170 kDa in the membranous fraction. Serine proteinase activities were detected using N-benzyloxycarbonyl-l-phenylalanyl-l-arginine 7-amino-4-methylcoumarin (Z-FR-AMC) and N-succinyl-l-alanine-l-phenylalanine-l-lysine 7-amino-4-methylcoumarin (Suc-AFK-AMC) as substrates in the cytosolic fraction (Z-FR-AMC = 392 ± 30 µmol.min-1 mg of protein-1 and Suc-AFK-AMC = 252 ± 20 µmol.min-1 mg of protein-1) and in the membranous fraction (Z-FR-AMC = 53 ± 5 µmol.min-1 mg of protein-1 and Suc-AFK-AMC = 63.6 ± 6.5 µmol.min-1 mg of protein-1). Enzyme specificity was shown by inhibition with aprotinin (19% to 80% inhibition) and phenylmethanesulfonyl fluoride (3% to 69%), depending on the subcellular fraction and substrate. The expression of subtilisin (LbrM.13.0860 and LbrM.28.2570) and tryparedoxin peroxidase (LbrM.15.1080) genes was observed by the detection of RNA transcripts 200 bp, 162 bp, and 166 bp long, respectively. Subsequent in silico assays showed LbrM.13.0860 can be located in the cytosol and LbrM.28.2570 in the membrane of the parasite. Data obtained here show the subcellular distribution and expression of serine proteinases, including the subtilisin-like serine proteinases in L. (V.) braziliensis promastigotes

Effect of mesoionic 4-phenyl-5-(cinnamoyl)-1,3,4-thiadiazolium-2-phenylamine chloride derivative salts on the activities of the nitric oxide synthase and arginase of Leishmania amazonensis

Submitted by Sandra Infurna ([email protected]) on 2019-04-11T10:37:23Z No. of bitstreams: 1 LeonorL_Leon_etal_IOC_2008.pdf: 442059 bytes, checksum: 3a58dab3266d52431ed8e7c5b3d66307 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2019-04-11T10:45:58Z (GMT) No. of bitstreams: 1 LeonorL_Leon_etal_IOC_2008.pdf: 442059 bytes, checksum: 3a58dab3266d52431ed8e7c5b3d66307 (MD5)Made available in DSpace on 2019-04-11T10:45:58Z (GMT). No. of bitstreams: 1 LeonorL_Leon_etal_IOC_2008.pdf: 442059 bytes, checksum: 3a58dab3266d52431ed8e7c5b3d66307 (MD5) Previous issue date: 2008Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Imunologia. Rio de Janeiro, RJ. Brasil.Universidade Federal Rural do Rio de Janeiro. Departamento de Química. Seropédica, RJ, Brasil.Universidade Federal Rural do Rio de Janeiro. Departamento de Química. Seropédica, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Imunologia. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Imunologia. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Imunologia. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Imunologia. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Departamento de Imunologia. Rio de Janeiro, RJ. Brasil.L-arginine is involved in the production of both nitric oxide (NO), mediated by nitric oxide synthase (NOS) and L-ornithine, by arginase activity. It is generally accepted that NO regulation occurs mainly at the transcriptional level of NOS. In a previous work we purported that there is evidence that Leishmania sp. can produce NO from L-arginine. An arginase activity in its gene sequence has also been reported in Leishmania parasites. In a search for intracellular targets as potential antileishmanicidal agents, such as the L-arginine metabolism, we used 1,3,4-thiadiazolium mesoionic compounds, that have been demonstrated to be cytotoxic to the Leishmania amazonensis, when compared to Pentamidine isethionate as a reference drug. Parasites were assayed in absence/presence of 4'- and 3'-methoxy mesoionic derivatives in order to verify the effect on NO production and arginase activity in L. amazonensis. The results indicated that the drugs reduce from 70 to 90% of the NO production by the parasite and act on a soluble nitric oxide synthase purified from L. amazonensis promastigotes and axenic amastigotes

Kinetoplastid membrane protein-11 is present in promastigotes and amastigotes of Leishmania amazonensis and its surface expression increases during metacyclogenesis

Kinetoplastid membrane protein-11 (KMP-11), a protein present in all kinetoplastid protozoa, is considered a potential candidate for a leishmaniasis vaccine. A suitable leishmaniasis vaccine candidate molecule must be expressed in amastigotes, the infective stage for mammals. However, the expression of KMP-11 in Leishmania amastigotes has been a subject of controversy. We evaluated the expression of this molecule in logarithmic and stationary growth phase promastigotes, as well as in amastigotes, of Leishmania amazonensis by immunoblotting, flow cytometry and immunocytochemistry, using a monoclonal antibody against KMP-11. We found that KMP-11 is present in promastigotes and amastigotes. In both stages, the protein was found in association with membrane structures (at the cell surface, flagellar pocket and intracellular vesicles). More importantly, its surface expression is higher in amastigotes than in promastigotes and increases during metacyclogenesis. The increased expression of KMP-11 in metacyclic promastigotes, and especially in amastigotes, indicates a role for this molecule in the parasite relationship with the mammalian host. The presence of this molecule in amastigotes is consistent with the previously demonstrated immunoprotective capacity of vaccine prototypes based on the KMP-11-coding gene and the presence of humoral and cellular immune responses to KMP-11 in Leishmania-infected humans and animals

Evidence for Tissue Toxicity in BALB/c Exposed to a Long-Term Treatment with Oxiranes Compared to Meglumine Antimoniate

Submitted by Sandra Infurna ([email protected]) on 2017-11-30T15:03:03Z No. of bitstreams: 1 luizfilipe_oliveira_etal_IOC_2017.pdf: 13582294 bytes, checksum: 884ee7892f0c8033aa9f9268a38130c7 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2017-11-30T15:19:55Z (GMT) No. of bitstreams: 1 luizfilipe_oliveira_etal_IOC_2017.pdf: 13582294 bytes, checksum: 884ee7892f0c8033aa9f9268a38130c7 (MD5)Made available in DSpace on 2017-11-30T15:19:55Z (GMT). No. of bitstreams: 1 luizfilipe_oliveira_etal_IOC_2017.pdf: 13582294 bytes, checksum: 884ee7892f0c8033aa9f9268a38130c7 (MD5) Previous issue date: 2017Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunoparasitologia. Rio de Janeiro, RJ. Brasil.Universidade do Estado do Rio de Janeiro. Instituto de Biologia Roberto Alcântara Gomes. Laboratório de Ultraestrutura e Biologia Tecidual. Rio de Janeiro, RJ, Brasil.Fundação Oswaldo Cruz. Instituto de Tecnologia em Imunobiológicos. Laboratório de Tecnologia Virológica,.Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto de Tecnologia em Imunobiológicos. Laboratório de Tecnologia Virológica,.Rio de Janeiro, RJ. Brasil.Universidade Federal Fluminense. Instituto de Biologia. Niterói, RJ, Brasil.Universidade Federal Fluminense. Instituto de Química. Departamento de Química Orgânica. Niterói, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório Interdisciplinar de Pesquisas Médicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Biologia Molecular e Doenças Endêmicas. Rio de Janeiro, RJ. Brasil.Leishmaniasis remains a serious public health problem in developing countries without effective control, whether by vaccination or chemotherapy. Part of the failure of leishmaniasis control is due to the lack of new less toxic and more effective drugs able to eliminate both the lesions and the parasite. Oxiranes derived from naphthoquinones now being assayed are promising drugs for the treatment of this group of diseases. The predicted pharmacokinetic properties and toxicological profiles of epoxy-α-lapachone and epoxymethoxy-lawsone have now been compared to those of meglumine antimoniate, and histological changes induced by these drugs in noninfected BALB/c mice tissues are described. Effects of these compounds on liver, kidney, lung, heart, and cerebral tissues of healthy mice were examined. The data presented show that both these oxiranes and meglumine antimoniate induce changes in all BALB/c mice tissues, with the lung, heart, and brain being the most affected. Epoxymethoxy-lawsone was the most toxic to lung tissue, while most severe damage was caused in the heart by epoxy-α-lapachone. Meglumine antimoniate caused mild-to-moderate changes in heart and lung tissues

Leishmania (Viannia) braziliensis: Influence of successive in vitro cultivation on the expression of promastigote proteinases

Cysteine proteinases are an important virulence factor in Leishmania parasites. In this study we analyzed the cysteine proteinase expression of infective Leishmania (Viannia) braziliensis promastigotes, examining the expression induced by successive in vitro passages in culture. We observed that this parasite presents a decrease in its virulence over BALB/c macrophages, after successive passages in culture, but still they present proteinase activity, being capable of hydrolyzing the substrate pGlu-Phe-Leu-p Nitroanilide at pH 7.0. This proteinase activity also decreases in the course of the successive passages. Additionally, the decrease in the amount of CPB proteins following successive passages of promastigotes was verified by immunoblotting assays, using an anti-CPB antiserum. Real-time PCR assays were performed to assess the relative cpb expression when compared to a housekeeping gene in promastigote cDNA preparations from the first, fourth and seventh passages. Interestingly, the data indicate a relative increase in cpb gene transcripts as the promastigotes were maintained under in vitro culture: 2.2 times higher for fourth and 2.7 times higher for seventh passages when compared to the first passage. Thus, the information gathered here shows that the expression of cysteine proteinases is modified during in vitro cultivation of L. (V.) braziliensis promastigotes