Two biochemically distinct lipophosphoglycans from Leishmania braziliensis and Leishmania infantum trigger different innate immune responses in murine macrophages

BACKGROUND: The dominant, cell surface lipophosphoglycan (LPG) of Leishmania is a multifunctional molecule involved in the interaction with vertebrate and invertebrate hosts. Although the role of LPG on infection has been extensively studied, it is not known if LPG interspecies variations contribute to the different immunopathologies of leishmaniases. To investigate the issue of interspecies polymorphisms, two Leishmania species from the New World that express structural variations of side chains of LPG repeat units were examined. In this context, the procyclic form of L. braziliensis LPG (strain M2903), is devoid of side chains, while the L. infantum LPG (strain BH46) has up to three glucoses residues in the repeat units. METHODS: Mice peritoneal macrophages from Balb/c, C57BL/6 and knock-out (TLR2 -/-, TLR4 -/-) were primed with IFN-γ and stimulated with purified LPG from both species. Nitric oxide and cytokine production, MAPKs (ERK, p38 and JNK) and NF-kB activation were evaluated. RESULTS: Macrophages stimulated with L. braziliensis LPG, had a higher TNF-α, IL-1β, IL-6 and NO production than those stimulated with that of L. infantum. Furthermore, the LPGs from the two species resulted in differential kinetics of signaling via MAPK activation. L. infantum LPG exhibited a gradual activation profile, whereas L. braziliensis LPG showed a sharp but transient activation. L. braziliensis LPG was able to activate NF-kB. CONCLUSION: These data suggest that two biochemically distinct LPGs were able to differentially modulate macrophage functions

Glycoinositolphospholipids from Leishmania braziliensis and L. infantum: Modulation of Innate Immune System and Variations in Carbohydrate Structure

The essential role of the lipophosphoglycan (LPG) of Leishmania in innate immune response has been extensively reported. However, information about the role of the LPG-related glycoinositolphospholipids (GIPLs) is limited, especially with respect to the New World species of Leishmania. GIPLs are low molecular weight molecules covering the parasite surface and are similar to LPG in sharing a common lipid backbone and a glycan motif containing up to 7 sugars. Critical aspects of their structure and functions are still obscure in the interaction with the vertebrate host. In this study, we evaluated the role of those molecules in two medically important South American species Leishmania infantum and L. braziliensis, causative agents of visceral (VL) and cutaneous Leishmaniasis (CL), respectively. GIPLs derived from both species did not induce NO or TNF-α production by non-primed murine macrophages. Additionally, primed macrophages from mice (BALB/c, C57BL/6, TLR2−/− and TLR4−/−) exposed to GIPLs from both species, with exception to TNF-α, did not produce any of the cytokines analyzed (IL1-β, IL-2, IL-4, IL-5, IL-10, IL-12p40, IFN-γ) or p38 activation. GIPLs induced the production of TNF-α and NO by C57BL/6 mice, primarily via TLR4. Pre incubation of macrophages with GIPLs reduced significantly the amount of NO and IL-12 in the presence of IFN-γ or lipopolysaccharide (LPS), which was more pronounced with L. braziliensis GIPLs. This inhibition was reversed after PI-specific phospholipase C treatment. A structural analysis of the GIPLs showed that L. infantum has manose rich GIPLs, suggestive of type I and Hybrid GIPLs while L. braziliensis has galactose rich GIPLs, suggestive of Type II GIPLs. In conclusion, there are major differences in the structure and composition of GIPLs from L. braziliensis and L. infantum. Also, GIPLs are important inhibitory molecules during the interaction with macrophages

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear un derstanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5–7 vast areas of the tropics remain understudied.8–11 In the American tropics, Amazonia stands out as the world’s most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepre sented in biodiversity databases.13–15 To worsen this situation, human-induced modifications16,17 may elim inate pieces of the Amazon’s biodiversity puzzle before we can use them to understand how ecological com munities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple or ganism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region’s vulnerability to environmental change. 15%–18% of the most ne glected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lostinfo:eu-repo/semantics/publishedVersio

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear understanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5,6,7 vast areas of the tropics remain understudied.8,9,10,11 In the American tropics, Amazonia stands out as the world's most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepresented in biodiversity databases.13,14,15 To worsen this situation, human-induced modifications16,17 may eliminate pieces of the Amazon's biodiversity puzzle before we can use them to understand how ecological communities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple organism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region's vulnerability to environmental change. 15%–18% of the most neglected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lost

Glicoconjugados (GIPLs e LPGs) de Leishmania braziliensis e L. infantum: Modulação do Sistema Imune Inato e variações na estrutura de carboidratos

Submitted by Nuzia Santos ([email protected]) on 2013-06-03T12:50:25Z No. of bitstreams: 1 Tese-Rafael Assis.pdf: 16998488 bytes, checksum: 138dc6a874eb36a1c66935f3dfab1da4 (MD5)Made available in DSpace on 2013-06-03T12:50:25Z (GMT). No. of bitstreams: 1 Tese-Rafael Assis.pdf: 16998488 bytes, checksum: 138dc6a874eb36a1c66935f3dfab1da4 (MD5) Previous issue date: 2012Fundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Belo Horizonte, MG, Brasil.Os glicoconjugados de Leishmaniatem sido extensivamente estudados, mas ainda pouco se sabe sobre o quanto polimorfismos intra e interespecíficos contribuem com o desenvolvimento das diferentes imunopatologias das leishmanioses. Por este motivo, duas espécies de importância epidemiológica foram examinadas, L. braziliensise L. infantum, agentes causadores das leishmanioses cutânea e visceral, respectivamente. O LPG de L. braziliensisnão possui cadeias laterais enquanto o LPG de L. infantumcarrega em sua estrutura oligômeros de até três β-glicoses como cadeias laterais. Por outro lado a estrutura dos GIPLs destas espécies era desconhecida e foi objeto de estudo deste trabalho. A análise estrutural dos GIPLs mostrou que L. infantumpossui GIPLs pequenos e ricos em manose, sugerindo predominância de GIPLs do tipoI e híbridos enquanto L. braziliensis apresenta GIPLs grandes e ricos em galactose, sugestivo do tipo II. Para analisar o papel destas moléculas na interação com o hospedeiro, macrófagosperitoneais murinos foram tratados com LPG ou GIPLs e a produção de nitrito, citocinas, bem como a ativação de MAPKs foram avaliados. De forma geral, macrófagos estimulados com LPG de L. braziliensis, demonstraram uma produção maior de TNF-α, IL-1β, IL-6 e NO do que os estimulados com LPG de L. infantum, adicionalmente, células tratadas com LPG de ambas as espécies mostraram uma resposta pro inflamatória mais proeminente. Além disto, os GIPLsmostraram a capacidade de inibir a produção de IL-12 e NO em macrófagos estimulados com IFN-γe LPS. Finalmente, os glicoconjugados destas duas espécies resultaram em uma cinética diferencial na ativação de MAPKs. O LPG de L. braziliensis mostrou uma ativação transiente enquanto o de L. infantum uma ativação gradual. Os GIPLs de ambas espécies falharam em ativar MAPKsLeismania LPG has being extensively studied but little is known about in what extent interspecies variations contribute to the differentimmunopathologies of leishmaniases. Similarly, for GIPLs, a relatively neglected molecule, little is known about its polymorphisms among Leishmania species and its role during infection. To address this issue, two epidemiologically important South American species of Leishmania were examined, L. braziliensis and L. infantum, causative agents of cutaneous and visceral leishmaniasis, respectively. The LPG from these two species differ in structure, being L. braziliensis LPG devoid of side chains while L. infantum LPG carries one to three β-glucoses on its side chains. In the other hand the structure of the GIPLs from these two species is still unknown and was addressed in this present work. A structural analysis of the GIPLs showed that L. infantum has small, mannose rich GIPLs, suggestive of type I and Hybrid GIPLs while L. braziliensis has larger and galactose rich GIPLs, suggestive of Type II GIPLs. To address the role of these molecules upon macrophage invasion, mouse peritoneal macrophages were treatedwith either LPG or GIPLs and Nitrite and cytokine production, as well as MAPKs activation were evaluated. Overall, macrophages stimulated with L. braziliensis LPG, had a higher TNF-α, IL-1β, IL-6 e NO production than those stimulated with that of L. infantum, also, IFN-γprimed macrophages stimulated with LPG, had a higher production of NO and TNF-αthan GIPLs stimulated cells. Additionally, GIPLs showed the capacity of inhibit IL-12 and NO production in IFN-γand LPS stimulated macrophages. Furthermore, the glycoconjugates from the two species resulted in differential kinetics of signaling via MAPK activation. L. infantum LPG exhibited a gradual activation profile, whereas L. braziliensisLPG showed a sharp but transient activation, while GIPLs failed to activate MAPK

The Influence of B Cell Depletion Therapy on Naturally Acquired Immunity to Streptococcus pneumoniae

The anti-CD20 antibody Rituximab to deplete CD20+ B cells is an effective treatment for rheumatoid arthritis and B cell malignancies, but is associated with an increased incidence of respiratory infections. Using mouse models we have investigated the consequences of B cell depletion on natural and acquired humoral immunity to Streptococcus pneumoniae. B cell depletion of naïve C57Bl/6 mice reduced natural IgM recognition of S. pneumoniae, but did not increase susceptibility to S. pneumoniae pneumonia. ELISA and flow cytometry assays demonstrated significantly reduced IgG and IgM recognition of S. pneumoniae in sera from mice treated with B cell depletion prior to S. pneumoniae nasopharyngeal colonization compared to untreated mice. Colonization induced antibody responses to protein rather than capsular antigen, and when measured using a protein array B cell depletion prior to colonization reduced serum levels of IgG to several protein antigens. However, B cell depleted S. pneumoniae colonized mice were still partially protected against both lung infection and septicemia when challenged with S. pneumoniae after reconstitution of their B cells. These data indicate that although B cell depletion markedly impairs antibody recognition of S. pneumoniae in colonized mice, some protective immunity is maintained, perhaps mediated by cellular immunity

Preclinical Development of Virulence Attenuated Strains Able to Enhance Protective Immunity Against Pneumococcal Infection.

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Leishmania enriettii: biochemical characterisation of lipophosphoglycans (LPGs) and glycoinositolphospholipids (GIPLs) and infectivity to Cavia porcellus

Background: Leishmania enriettii is a species non-infectious to man, whose reservoir is the guinea pig Cavia porcellus. Many aspects of the parasite-host interaction in this model are unknown, especially those involving parasite surface molecules. While lipophosphoglycans (LPGs) and glycoinositolphospholipids (GIPLs) of Leishmania species from the Old and New World have already been described, glycoconjugates of L. enriettii and their importance are still unknown.Methods: Mice peritoneal macrophages from C57BL/6 and knock-out (TLR2 -/-, TLR4 -/-) were primed with IFN-gamma and stimulated with purified LPG and GIPLs from both species. Nitric oxide and cytokine production were performed. MAPKs (p38 and JNK) and NF-kB activation were evaluated in J774.1 macrophages and CHO cells, respectively.Results: LPGs were extracted, purified and analysed by western-blot, showing that LPG from L88 strain was longer than that of Cobaia strain. LPGs and GIPLs were depolymerised and their sugar content was determined. LPGs from both strains did not present side chains, having the common disaccharide Gal(beta 1,4) Man(alpha 1)-PO4. the GIPL from L88 strain presented galactose in its structure, suggestive of type II GIPL. On the other hand, the GIPL of Cobaia strain presented an abundance of glucose, a characteristic not previously observed. Mice peritoneal macrophages from C57BL/6 and knock-outs (TLR2 -/- and TLR4 -/-) were primed with IFN-gamma and stimulated with glycoconjugates and live parasites. No activation of NO or cytokines was observed with live parasites. On the other hand, LPGs and GIPLs were able to activate the production of NO, IL-6, IL-12 and TNF-alpha preferably via TRL2. However, in CHO cells, only GIPLs were able to activate TRL2 and TRL4. in vivo studies using male guinea pigs (Cavia porcellus) showed that only strain L88 was able to develop more severe ulcerated lesions especially in the presence of salivary gland extract (SGE).Conclusion: the two L. enriettii strains exhibited polymorphisms in their LPGs and GIPLs and those features may be related to a more pro-inflammatory profile in the L88 strain