Click
here to close Hello! We notice that
you are using Internet Explorer, which is not supported by Echinobase
and may cause the site to display incorrectly. We suggest using a
current version of Chrome,
FireFox,
or Safari.
Front Microbiol
2018 Jan 01;9:3056. doi: 10.3389/fmicb.2018.03056.
Show Gene links
Show Anatomy links
Extracellular Vesicles Released by Leishmania (Leishmania) amazonensis Promote Disease Progression and Induce the Production of Different Cytokines in Macrophages and B-1 Cells.
Barbosa FMC
,
Dupin TV
,
Toledo MDS
,
Reis NFDC
,
Ribeiro K
,
Cronemberger-Andrade A
,
Rugani JN
,
De Lorenzo BHP
,
Novaes E Brito RR
,
Soares RP
,
Torrecilhas AC
,
Xander P
.
???displayArticle.abstract???
The extracellular vesicles (EVs) released by Leishmania can contribute to the establishment of infection and host immunomodulation. In this study, we characterized the shedding of EVs from Leishmania (Leishmania) amazonensis promastigotes. This species is the causative agent of cutaneous leishmaniasis, and its role during interactions with bone marrow-derived macrophages (BMDMs) and peritoneal B-1 cells was evaluated. Leishmania amazonensis promastigotes cultivated in vitro at different times and temperatures spontaneously released EVs. EVs were purified using size-exclusion chromatography (SEC) and quantitated by nanoparticle tracking analysis (NTA). NTA revealed that the average size of the EVs was approximately 180 nm, with concentrations ranging from 1.8 × 108 to 2.4 × 109 vesicles/mL. In addition, the presence of LPG and GP63 were detected in EVs obtained at different temperatures. Naïve BMDMs stimulated with EVs exhibited increased IL-10 and IL-6 expression. However, incubating B-1 cells with parasite EVs did not stimulate IL-10 expression but led to an increase in the expression of IL-6 and TNFα. After 7 weeks post-infection, animals infected with L. amazonensis promastigotes in the presence of parasite EVs had significant higher parasite load and a polarization to Th2 response, as compared to the group infected with the parasite alone. This work demonstrated that EVs isolated from L. amazonensis promastigotes were able to stimulate macrophages and B-1 cells to express different types of cytokines. Moreover, the immunomodulatory properties of EVs probably contributed to an increase in parasite burden in mice. These findings suggest that the functionality of L. amazonensis EVs on immune system favor of parasite survival and disease progression.
FIGURE 1. Leishmania amazonensis promastigotes spontaneously release EVs from their surface after incubation at 26, 34, and 37°C for (A–C) 1, (D–F) 2, (G–I) 4, and (J–L) 24 h. Parasites are seen by SEM at different magnifications. The arrows indicate EVs on the surface of the parasites. Scale bars = 1 μm.
FIGURE 4. Purification of EVs released by L. amazonensis by SEC. Titration of the L. amazonensis EV-specific antibody response in BALB/c mice. Blood samples were collected before immunization (non-immunized mice), 15 days after the first immunization and 15 days after the second immunization. The level of IgG was measured in (A) animals immunized with EVs prepared after 4 h of parasite incubation at 26°C and in (B) animals immunized with EVs prepared after 4 h of parasite incubation at 37°C. Analyses were performed using ELISA with twofold serially diluted sera. The result is depicted as the mean optical density (OD) at 492 nm of five mice per group. (C) SEC. The promastigotes were cultured for 4 h at 26, 34, or 37°C. The presence of EVs was measured in each fraction by ELISA with anti-EV polyclonal antibodies. Seventy samples were collected, and each fraction was analyzed by ELISA. The graph shows the overlap of the measurements performed for the L. amazonensis EVs released at 26, 34, and 37°C. (D) Size profile of L. amazonensis EVs obtained by SEC. The particles were analyzed by NTA.
FIGURE 5. Enzyme-linked immunoassay (ELISA) and dot blotting analysis of EVs released by L. amazonensis promastigotes. The parasites were cultured at 26, 34, or 37°C for 4 h. The presence of gp63 and LPG were evaluated by specific mAbs. (A) Dot blotting of EVs (4 μg per sample) probed with the mAb anti-gp63. (B) The detection of LPG was performed by ELISA with mAb CA7AE.
FIGURE 8. Development of infection in BALB/c mice experimentally infected with L. amazonensis promastigotes in the presence or absence of EVs from the parasite (L. ama+EVs and L. ama, respectively). BALB/c mice were infected in the footpad with 1 × 106
L. amazonensis promastigotes in the presence or absence of 1 × 107 EVs. (A) The lesion size was evaluated weekly for 7 weeks. The graph shows the measurement of the paw size and each point is representative of the mean of the measurements. (B) The parasite burden was evaluated by limiting dilution in the footpads (n = 5). Bars denote the average of 5 measurements, and error bars denote the SD. Student’s t-test ∗P < 0.05. (C) Histopathological findings in lesions after 7 weeks post-infection of BALB/c mice infected in the absence of EVs and (D) mice infected in the presence of EVs. The paws were removed, fixed, and 2 tissue samples from each animal were cut into 5 μm sections for staining with hematoxylin and eosin. The data are representative of 3 independent experiments.
Arcanjo,
B-1 cells modulate the murine macrophage response to Leishmania major infection.
2017, Pubmed
Arcanjo,
B-1 cells modulate the murine macrophage response to Leishmania major infection.
2017,
Pubmed
Arcanjo,
The PGE2/IL-10 Axis Determines Susceptibility of B-1 Cell-Derived Phagocytes (B-1CDP) to Leishmania major Infection.
2015,
Pubmed
Atayde,
Exosome Secretion by the Parasitic Protozoan Leishmania within the Sand Fly Midgut.
2015,
Pubmed
Baumgarth,
A Hard(y) Look at B-1 Cell Development and Function.
2017,
Pubmed
Belkaid,
The role of interleukin (IL)-10 in the persistence of Leishmania major in the skin after healing and the therapeutic potential of anti-IL-10 receptor antibody for sterile cure.
2001,
Pubmed
Biswas,
Interleukin-6 induces monocyte chemotactic protein-1 in peripheral blood mononuclear cells and in the U937 cell line.
1998,
Pubmed
Bouvy,
Transfer of multidrug resistance among acute myeloid leukemia cells via extracellular vesicles and their microRNA cargo.
2017,
Pubmed
Bustin,
The MIQE guidelines: minimum information for publication of quantitative real-time PCR experiments.
2009,
Pubmed
Böing,
Single-step isolation of extracellular vesicles by size-exclusion chromatography.
2014,
Pubmed
Campos,
Extracellular Vesicles: Role in Inflammatory Responses and Potential Uses in Vaccination in Cancer and Infectious Diseases.
2015,
Pubmed
,
Echinobase
Cantacessi,
The past, present, and future of Leishmania genomics and transcriptomics.
2015,
Pubmed
Castellano,
Potential Use of Interleukin-10 Blockade as a Therapeutic Strategy in Human Cutaneous Leishmaniasis.
2015,
Pubmed
Coakley,
Extracellular Vesicles from a Helminth Parasite Suppress Macrophage Activation and Constitute an Effective Vaccine for Protective Immunity.
2017,
Pubmed
Deng,
Neurons Export Extracellular Vesicles Enriched in Cysteine String Protein and Misfolded Protein Cargo.
2017,
Pubmed
Garcia,
Culture-independent species typing of neotropical Leishmania for clinical validation of a PCR-based assay targeting heat shock protein 70 genes.
2004,
Pubmed
Geherin,
IL-10+ Innate-like B Cells Are Part of the Skin Immune System and Require α4β1 Integrin To Migrate between the Peritoneum and Inflamed Skin.
2016,
Pubmed
Geraldo,
In vivo and in vitro phagocytosis of Leishmania (Leishmania) amazonensis promastigotes by B-1 cells.
2016,
Pubmed
,
Echinobase
Gonzaga,
B-1 cells contribute to susceptibility in experimental infection with Leishmania (Leishmania) chagasi.
2015,
Pubmed
,
Echinobase
Gonzaga,
Evaluation of Experimental Infection with L. ( L.) Amazonensis in X-Linked Immunodeficient Mice.
2017,
Pubmed
Hassani,
Temperature-induced protein secretion by Leishmania mexicana modulates macrophage signalling and function.
2011,
Pubmed
Hassani,
Absence of metalloprotease GP63 alters the protein content of Leishmania exosomes.
2014,
Pubmed
Kalra,
Focus on Extracellular Vesicles: Introducing the Next Small Big Thing.
2016,
Pubmed
Kane,
The role of IL-10 in promoting disease progression in leishmaniasis.
2001,
Pubmed
Kowal,
Biogenesis and secretion of exosomes.
2014,
Pubmed
Lambertz,
Secreted virulence factors and immune evasion in visceral leishmaniasis.
2012,
Pubmed
Lima,
A simple method for quantifying Leishmania in tissues of infected animals.
1997,
Pubmed
Liu,
The early interaction of Leishmania with macrophages and dendritic cells and its influence on the host immune response.
2012,
Pubmed
Martin-Jaular,
Exosomes from Plasmodium yoelii-infected reticulocytes protect mice from lethal infections.
2011,
Pubmed
Martinez,
Chronic infection by Leishmania amazonensis mediated through MAPK ERK mechanisms.
2014,
Pubmed
Matte,
Leishmania-induced cellular recruitment during the early inflammatory response: modulation of proinflammatory mediators.
2002,
Pubmed
Meyer-Bahlburg,
Differential impact of Toll-like receptor signaling on distinct B cell subpopulations.
2012,
Pubmed
Nagase,
Despite increased CD4+Foxp3+ cells within the infection site, BALB/c IL-4 receptor-deficient mice reveal CD4+Foxp3-negative T cells as a source of IL-10 in Leishmania major susceptibility.
2007,
Pubmed
Nogueira,
Vesicles from different Trypanosoma cruzi strains trigger differential innate and chronic immune responses.
2015,
Pubmed
,
Echinobase
Nogueira,
Lipophosphoglycans from Leishmania amazonensis Strains Display Immunomodulatory Properties via TLR4 and Do Not Affect Sand Fly Infection.
2016,
Pubmed
Paranaiba,
An overview on Leishmania (Mundinia) enriettii: biology, immunopathology, LRV and extracellular vesicles during the host-parasite interaction.
2018,
Pubmed
,
Echinobase
Popi,
Interleukin-10 secreted by B-1 cells modulates the phagocytic activity of murine macrophages in vitro.
2004,
Pubmed
Popi,
B-1 cells facilitate Paracoccidioides brasiliensis infection in mice via IL-10 secretion.
2008,
Pubmed
Popi,
An Overview of B-1 Cells as Antigen-Presenting Cells.
2016,
Pubmed
Read,
Nuclear transportation of exogenous epidermal growth factor receptor and androgen receptor via extracellular vesicles.
2017,
Pubmed
Regev-Rudzki,
Cell-cell communication between malaria-infected red blood cells via exosome-like vesicles.
2013,
Pubmed
Rocha,
An alternative in vitro drug screening test using Leishmania amazonensis transfected with red fluorescent protein.
2013,
Pubmed
Sacks,
Re-examination of the immunosuppressive mechanisms mediating non-cure of Leishmania infection in mice.
2004,
Pubmed
Sampaio,
Extracellular vesicles from early stage Plasmodium falciparum-infected red blood cells contain PfEMP1 and induce transcriptional changes in human monocytes.
2018,
Pubmed
Savage,
Blimp-1-dependent and -independent natural antibody production by B-1 and B-1-derived plasma cells.
2017,
Pubmed
Schmittgen,
Analyzing real-time PCR data by the comparative C(T) method.
2008,
Pubmed
Schönian,
PCR diagnosis and characterization of Leishmania in local and imported clinical samples.
2003,
Pubmed
Silveira,
Clinical and immunopathological spectrum of American cutaneous leishmaniasis with special reference to the disease in Amazonian Brazil: a review.
2004,
Pubmed
Silveira,
Immunopathogenic competences of Leishmania (V.) braziliensis and L. (L.) amazonensis in American cutaneous leishmaniasis.
2009,
Pubmed
Silverman,
Proteomic analysis of the secretome of Leishmania donovani.
2008,
Pubmed
Silverman,
An exosome-based secretion pathway is responsible for protein export from Leishmania and communication with macrophages.
2010,
Pubmed
Silverman,
Leishmania exosomes modulate innate and adaptive immune responses through effects on monocytes and dendritic cells.
2010,
Pubmed
Smallwood,
Extracellular vesicles released by CD40/IL-4-stimulated CLL cells confer altered functional properties to CD4+ T cells.
2016,
Pubmed
Soares,
Leishmania braziliensis: a novel mechanism in the lipophosphoglycan regulation during metacyclogenesis.
2005,
Pubmed
Soares,
Highlights of the São Paulo ISEV workshop on extracellular vesicles in cross-kingdom communication.
2017,
Pubmed
Soong,
Subversion and Utilization of Host Innate Defense by Leishmania amazonensis.
2012,
Pubmed
Spandidos,
PrimerBank: a resource of human and mouse PCR primer pairs for gene expression detection and quantification.
2010,
Pubmed
Synowsky,
The major histocompatibility complex class I immunopeptidome of extracellular vesicles.
2017,
Pubmed
Szempruch,
Extracellular Vesicles from Trypanosoma brucei Mediate Virulence Factor Transfer and Cause Host Anemia.
2016,
Pubmed
Théry,
Isolation and characterization of exosomes from cell culture supernatants and biological fluids.
2006,
Pubmed
Tkach,
Qualitative differences in T-cell activation by dendritic cell-derived extracellular vesicle subtypes.
2017,
Pubmed
Tkach,
Why the need and how to approach the functional diversity of extracellular vesicles.
2018,
Pubmed
Tolson,
The immunochemical structure and surface arrangement of Leishmania donovani lipophosphoglycan determined using monoclonal antibodies.
1989,
Pubmed
Torrecilhas,
Vesicles as carriers of virulence factors in parasitic protozoan diseases.
2012,
Pubmed
,
Echinobase
Trocoli Torrecilhas,
Trypanosoma cruzi: parasite shed vesicles increase heart parasitism and generate an intense inflammatory response.
2009,
Pubmed
Zamboni,
Nitric oxide partially controls Coxiella burnetii phase II infection in mouse primary macrophages.
2003,
Pubmed
Zhou,
Retinal progenitor cells release extracellular vesicles containing developmental transcription factors, microRNA and membrane proteins.
2018,
Pubmed
de Assis,
Glycoconjugates in New World species of Leishmania: polymorphisms in lipophosphoglycan and glycoinositolphospholipids and interaction with hosts.
2012,
Pubmed
e Brito,
Role of distinct immune components in the radiation-induced abrogation of systemic lupus erythematosus development in mice.
2007,
Pubmed
