Lutzomyia longipalpis Saliva Triggers Lipid Body Formation and Prostaglandin E2 Production in Murine Macrophages

After the injection of saliva into the host's skin by sand flies, a transient erythematous reaction is observed, which is related to an influx of inflammatory cells and the release of various molecules that actively facilitate the blood meal. It is important to understand the specific mechanisms by which sand fly saliva manipulates the host's inflammatory responses. Herein, we report that saliva from Lutzomyia (L.) longipalpis, a widespread Leishmania vector, induces early production of eicosanoids. Intense formation of intracellular organelles called lipid bodies (LBs) was noted within those cells that migrated to the site of saliva injection. In vitro and ex vivo, sand fly saliva was able to induce LB formation and PGE2 release by macrophages. Interestingly, PGE2 production induced by L. longipalpis saliva was dependent on intracellular mechanisms involving phosphorylation of signaling proteins such as PKC-α and ERK-1/2 and subsequent activation of cyclooxygenase-2. Thus, this study provides new insights into the pharmacological properties of sand fly saliva and opens new opportunities for intervening with the induction of the host's inflammatory pathways by L. longipalpis bites

Using Recombinant Proteins from Lutzomyia longipalpis Saliva to Estimate Human Vector Exposure in Visceral Leishmaniasis Endemic Areas

During the blood meal, female sand flies (insects that transmit the parasite Leishmania) inject saliva containing a large variety of molecules with different pharmacological activities that facilitate the acquisition of blood. These molecules can induce the production of anti-saliva antibodies, which can then be used as markers for insect (vector) biting or exposure. Epidemiological studies using sand fly salivary gland sonicate as antigens are hampered by the difficulty of obtaining large amounts of salivary glands. In the present study, we have investigated the use of two salivary recombinant proteins from the sand fly Lutzomyia longipalpis, considered the main vector of visceral leishmaniasis, as an alternative method for screening of exposure to the sand fly. We primarily tested the suitability of using the recombinant proteins to estimate positive anti-saliva ELISA test in small sets of serum samples. Further, we validated the assay in a large sample of 1,077 individuals from an epidemiological survey in a second area endemic for visceral leishmaniasis. Our findings indicate that these proteins represent a promising epidemiological tool that can aid in implementing control measures against leishmaniasis

Interaction of lipophorin with Rhodnius prolixus oocytes: biochemical properties and the importance of blood feeding

Lipophorin (Lp) is the main haemolymphatic lipoprotein in insects and transports lipids between different organs. In adult females, lipophorin delivers lipids to growing oocytes. In this study, the interaction of this lipoprotein with the ovaries of Rhodnius prolixus was characterised using an oocyte membrane preparation and purified radiolabelled Lp (125I-Lp). Lp-specific binding to the oocyte membrane reached equilibrium after 40-60 min and when 125I-Lp was incubated with increasing amounts of membrane protein, corresponding increases in Lp binding were observed. The specific binding of Lp to the membrane preparation was a saturable process, with a Kdof 7.1 ± 0.9 x 10-8M and a maximal binding capacity of 430 ± 40 ng 125I-Lp/µg of membrane protein. The binding was calcium independent and pH sensitive, reaching its maximum at pH 5.2-5.7. Suramin inhibited the binding interaction between Lp and the oocyte membranes, which was completely abolished at 0.5 mM suramin. The oocyte membrane preparation from R. prolixus also showed binding to Lp from Manduca sexta. When Lp was fluorescently labelled and injected into vitellogenic females, the level of Lp-oocyte binding was much higher in females that were fed whole blood than in those fed blood plasma

Leishmania infantum lipophosphoglycan induced-Prostaglandin E2production in association with PPAR-γ expression via activation of Toll like receptors-1 and 2

Submitted by Sandra Infurna ([email protected]) on 2018-02-20T14:30:30Z No. of bitstreams: 1 alan_carneiro_etal_IOC_2017.pdf: 1888939 bytes, checksum: b9e4323376f30ec53fccc102bfc178bc (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2018-02-20T14:45:35Z (GMT) No. of bitstreams: 1 alan_carneiro_etal_IOC_2017.pdf: 1888939 bytes, checksum: b9e4323376f30ec53fccc102bfc178bc (MD5)Made available in DSpace on 2018-02-20T14:45:35Z (GMT). No. of bitstreams: 1 alan_carneiro_etal_IOC_2017.pdf: 1888939 bytes, checksum: b9e4323376f30ec53fccc102bfc178bc (MD5) Previous issue date: 2017Fundação Oswaldo Cruz. Instituto Gonçalo Muniz. Salvador, BA, Brasil / Universidade Federal do Oeste da Bahia. Centro de Ciências Biológicas e da Saúde. Barreiras, BA, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Muniz. Salvador, BA, Brasil / Universidade Federal do Oeste da Bahia. Centro de Ciências Biológicas e da Saúde. Barreiras, BA, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Muniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Salvador, BA, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunofarmacologia. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto René Rachou. Belo Horizonte, MG, BrasilFundação Oswaldo Cruz. Instituto Gonçalo Muniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Salvador, BA, Brasil.Fundação Oswaldo Cruz. Instituto René Rachou. Belo Horizonte, MG, Brasil.Fundação Oswaldo Cruz. Instituto Gonçalo Muniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Salvador, BA, Brasil.Universidade Federal do Rio de Janeiro. NUPEM. Campus Macaé. macaé, RJ, Brasil.Institut National de la Recherche Scientifique. Institut Armand-Frappier. Laval, Canada.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunofarmacologia. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto René Rachou. Belo Horizonte, MG, BrasilFundação Oswaldo Cruz. Instituto Gonçalo Muniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Salvador, BA, Brasil.Lipophosphoglycan (LPG) is a key virulence factor expressed on the surfaces of Leishmania promastigotes. Although LPG is known to activate macrophages, the underlying mechanisms resulting in the production of prostaglandin E2(PGE2) via signaling pathways remain unknown. Here, the inflammatory response arising from stimulation by Leishmania infantum LPG and/or its lipid and glycan motifs was evaluated with regard to PGE2induction. Intact LPG, but not its glycan and lipid moieties, induced a range of proinflammatory responses, including PGE2and nitric oxide (NO) release, increased lipid droplet formation, and iNOS and COX2 expression. LPG also induced ERK-1/2 and JNK phosphorylation in macrophages, in addition to the release of PGE2, MCP-1, IL-6, TNF-α and IL-12p70, but not IL-10. Pharmacological inhibition of ERK1/2 and PKC affected PGE2and cytokine production. Moreover, treatment with rosiglitazone, an agonist of peroxisome proliferator-activated receptor gamma (PPAR-γ), also modulated the release of PGE2and other proinflammatory mediators. Finally, we determined that LPG-induced PPAR-γ signaling occurred via TLR1/2. Taken together, these results reinforce the role played by L. infantum-derived LPG in the proinflammatory response seen in Leishmania infection

Searching genes encoding Leishmania antigens for diagnosis and protection

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2014-03-12T19:47:16Z No. of bitstreams: 1 Soto M Searching genes....pdf: 1180937 bytes, checksum: eceb54efdc1f94600c17649db8d84baf (MD5)Made available in DSpace on 2014-03-12T19:47:16Z (GMT). No. of bitstreams: 1 Soto M Searching genes....pdf: 1180937 bytes, checksum: eceb54efdc1f94600c17649db8d84baf (MD5) Previous issue date: 2009Universidad Autónoma de Madrid. Facultad de Ciencias. Centro de Biología Molecular Severo Ochoa. Departamento de Biología Molecular. Madrid, SpainUniversidad Autónoma de Madrid. Facultad de Ciencias. Centro de Biología Molecular Severo Ochoa. Departamento de Biología Molecular. Madrid, SpainUniversidad Autónoma de Madrid. Facultad de Ciencias. Centro de Biología Molecular Severo Ochoa. Departamento de Biología Molecular. Madrid, SpainHospital Ramón y Cajal. Departamento de Bioquímica-Investigación. Madrid, SpainFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilUniversidad Autónoma de Madrid. Facultad de Ciencias. Centro de Biología Molecular Severo Ochoa. Departamento de Biología Molecular. Madrid, SpainUniversidad Autónoma de Madrid. Facultad de Ciencias. Centro de Biología Molecular Severo Ochoa. Departamento de Biología Molecular. Madrid, SpainUniversidad Autónoma de Madrid. Facultad de Ciencias. Centro de Biología Molecular Severo Ochoa. Departamento de Biología Molecular. Madrid, SpainFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilInstituto de Salud Carlos III. Unidad de Inmunología Viral. Centro Nacional de Microbiología. Madrid, SpainLeishmaniases are a wide spectrum of parasitic diseases caused by the infection of different species of the genus Leishmania. Currently, these diseases are one of the most neglected diseases threatening 350 million people in different countries around the world. Thus, these diseases require better screening, diagnostics and treatment. An effective vaccine, that is not currently available, would be the best way to confront leishmaniases. In the past 20 years the molecular characterization of Leishmania genes encoding parasite antigens has been carried out. In this review we summarize the most common strategies employed for the isolation and characterization of genes encoding Leishmania antigens. To provide a collective view, we also discuss the results related with diagnosis and protection based on different recombinant DNA-derived Leishmania products

Leukocyte influx into the peritoneal cavity of C57BL/6 mice in response to <i>L. longipalpis</i> SGS.

<p>Mice were injected i.p. with endotoxin-free saline or SGS (0.5 pair/cavity). One (A), 3 (B) and 6 (C) hours after stimulation, cells were harvested by peritoneal lavage and differential leukocyte counts were performed on Diff-quick stained cytospin preparations. The data are the means and SEM from an experiment representative of three independent experiments. Groups were compared using Student's <i>t</i> test at each time point. *, <i>p</i><0.05 and ***, <i>p</i><0.001.</p

<i>L. longipalpis</i> SGS induces PGE₂ production via COX-2.

<p>A, Dose-response of PGE₂ production induced by SGS in peritoneal macrophages. B, Macrophages were pre-treated for 1 hour with the COX-2 inhibitor N-398 before incubation with SGS (1.5 pair/well). Twenty-four hours after stimulation, PGE₂ was measured in the supernatant. The data are the means and SEM from a representative experiment of three independent experiments. **, <i>p</i><0.01 and ^#, <i>p</i><0.05.</p

Effect of <i>L. longipalpis</i> SGS on lipid body formation in peritoneal macrophages <i>in vitro</i>.

<p>Representative image of peritoneal macrophages untreated (A) or stimulated with SGS (1.5 pair/well) (B) for 24 hours. Dose-response (C) and kinetics (D) of lipid body formation induced by SGS in peritoneal macrophages. **, <i>p</i><0.01 and ***, <i>p</i><0.001 compared with unstimulated cells.</p

<i>L. longipalpis</i> SGS induces PKC-α and ERK phosphorylation.

<p>Peritoneal macrophages were incubated in the absence (control) or presence of SGS (1.5 pair/mL) for 40 min. The cells were lysed and immunoblotted using polyclonal anti-ERK-1/2 (A) or anti-PKC-α (B) antibodies. The membranes was discharged and immunoblotted using polyclonal anti- phospho-ERK-1/2 (A) or anti- phosphor-PKC-α (B) antibodies. Quantification of phosphorylated-ERK-1/2 (C) and phosphorylated-PKCα (D) was determined by densitometry. The data show the fold increase in the phosphorylated/unphosphorylated kinase ratio of the SGS group relative to the control group. P-, phosphorylated.</p

ERK and PKC kinase inhibitors abrogate PGE₂ production induced by <i>L. longipalpis</i> SGS.

<p>Peritoneal macrophages were pre-treated for 1 hour with BIS I (A) or PD98059 (B) before incubation with SGS (1.5 pair/well). Twenty-four hours after stimulation, PGE₂ was measured in the supernatant. The data are the mean and SEM from an experiment representative of three independent experiments. ***, <i>p</i><0.001; ^##, <i>p</i><0.01 and ^###, <i>p</i><0.001. PD98059, ERK inhibitor; BIS-I, PKC inhibitor.</p