In Vivo Antileishmanial Efficacy of Miltefosine AgainstLeishmania(Leishmania)amazonensis

Leishmania protozoans are the causal agents of neglected diseases that represent an important public health issue worldwide. The growing occurrence of drug-resistant strains of Leishmania and severe side effects of available treatments represent an important challenge for the leishmaniases treatment. We have previously reported the leishmanicidal activity of phylloseptin-1 (PSN-1), a peptide found in the skin secretion of Phyllomedusa azurea (=Pithecopus azureus), against Leishmania amazonensis promastigotes. However, its impact on the amastigote form of L. amazonensis and its impact on infected macrophages are unknown. In this work, we evaluated the effects of PSN-1 on amastigotes of L. amazonensis inside macrophages infected in vitro. We assessed the production of hydrogen peroxide and nitric oxide, as well as the levels of inflammatory and immunomodulatory markers (TGF-β, TNF-α and IL-12), in infected and non-infected macrophages treated with PSN-1. Treatment with PSN-1 decreased the number of infected cells and the number of ingested amastigotes per cell when compared with the untreated cells. At 32 µM (64 µg/mL), PSN-1 reduced hydrogen peroxide levels in both infected and uninfected macrophages, whereas it had little effect on NO production or TGF-β release. The effect of PSN-1 on IL-12 and TNF-α secretion depended on its concentration, but, in general, their levels tended to increase as PSN-1 concentration increased. Further in vitro and in vivo studies are needed to clarify the mechanisms of action of PSN-1 and its interaction with the immune system aiming to develop pharmacological applications.

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Antigen-Responsive CD4+ T Cells from C3H Mice Chronically Infected with Leishmania amazonensis Are Impaired in the Transition to an Effector Phenotype

Infection and Immunity ◽

10.1128/iai.74.3.1547-1554.2006 ◽

2006 ◽

Vol 74 (3) ◽

pp. 1547-1554 ◽

Cited By ~ 18

Author(s):

Amanda E. Ramer ◽

Yannick F. Vanloubbeeck ◽

Douglas E. Jones

Keyword(s):

T Cells ◽

T Cell ◽

Cell Population ◽

T Cell Response ◽

Leishmania Amazonensis ◽

Effector Memory ◽

Intrinsic Defect ◽

Antigen Stimulation ◽

Ifn Γ

ABSTRACT C3HeB/FeJ mice challenged with Leishmania major develop a polarized Th1 response and subsequently heal, whereas Leishmania amazonensis challenge leads to chronic lesions with high parasite loads at 10 weeks postinfection. In this study, a comparison of draining lymph node cells from L. amazonensis- and L. major-infected mice at 10 weeks postinfection showed equivalent percentages of effector/memory phenotype CD44hi CD4+ T cells producing interleukin-2 (IL-2) and proliferating after antigen stimulation. However, these cells isolated from L. amazonensis-infected mice were not skewed toward either a Th1 or Th2 phenotype in vivo, as evidenced by their unbiased Th1/Th2 transcription factor mRNA profile. In vivo antigen stimulation with added IL-12 failed to enhance gamma interferon (IFN-γ) production of CD4+ T cells from L. amazonensis-infected mice. Antigen stimulation of CD4+ T cells from L. amazonensis-infected mice in vitro in the presence of IL-12 resulted in production of only 10 to 15% of the IFN-γ produced by T cells from L. major-infected mice under identical conditions. These results suggest that the CD4+ T-cell response during chronic L. amazonensis infection is limited during the transition from an early activated CD4+ T-cell population to an effector cell population and demonstrate that these T cells have an intrinsic defect beyond the presence or absence of IL-12 during antigen stimulation.

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The natural absence of RPA1N domain did not impair Leishmania amazonensis RPA-1 participation in DNA damage response and telomere protection

Parasitology ◽

10.1017/s0031182012002028 ◽

2013 ◽

Vol 140 (4) ◽

pp. 547-559 ◽

Cited By ~ 11

Author(s):

RITA DE CÁSSIA VIVEIROS DA SILVEIRA ◽

MARCELO SANTOS DA SILVA ◽

VINÍCIUS SANTANA NUNES ◽

ARINA MARINA PEREZ ◽

MARIA ISABEL NOGUEIRA CANO

Keyword(s):

S Phase ◽

Leishmania Amazonensis ◽

Nuclear Accumulation ◽

Dna Double Strand Breaks ◽

Telomeric Dna ◽

Strand Breaks ◽

Cycle Arrest ◽

Telomere Protection ◽

Damage Response

SUMMARYWe have previously shown that the subunit 1 of Leishmania amazonensis RPA (LaRPA-1) alone binds the G-rich telomeric strand and is structurally different from other RPA-1. It is analogous to telomere end-binding proteins described in model eukaryotes whose homologues were not identified in the protozoan´s genome. Here we show that LaRPA-1 is involved with damage response and telomere protection although it lacks the RPA1N domain involved with the binding with multiple checkpoint proteins. We induced DNA double-strand breaks (DSBs) in Leishmania using phleomycin. Damage was confirmed by TUNEL-positive nuclei and triggered a G1/S cell cycle arrest that was accompanied by nuclear accumulation of LaRPA-1 and RAD51 in the S phase of hydroxyurea-synchronized parasites. DSBs also increased the levels of RAD51 in non-synchronized parasites and of LaRPA-1 and RAD51 in the S phase of synchronized cells. More LaRPA-1 appeared immunoprecipitating telomeres in vivo and associated in a complex containing RAD51, although this interaction needs more investigation. RAD51 apparently co-localized with few telomeric clusters but it did not immunoprecipitate telomeric DNA. These findings suggest that LaRPA-1 and RAD51 work together in response to DNA DSBs and at telomeres, upon damage, LaRPA-1 works probably to prevent loss of single-stranded DNA and to assume a capping function.

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Caracterização de proteinas que se associam in vivo com a simples-fita telomerica de Leishmania amazonensis

10.47749/t/unicamp.2007.416456 ◽

2007 ◽

Author(s):

Jair Lage de Siqueira Neto

Keyword(s):

Leishmania Amazonensis

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A Leishmania amazonensis ZIP family iron transporter is essential for parasite replication within macrophage phagolysosomes

Journal of Experimental Medicine ◽

10.1084/jem.20060559 ◽

2006 ◽

Vol 203 (10) ◽

pp. 2363-2375 ◽

Cited By ~ 105

Author(s):

Chau Huynh ◽

David L. Sacks ◽

Norma W. Andrews

Keyword(s):

Essential Role ◽

Iron Acquisition ◽

Axenic Culture ◽

Leishmania Amazonensis ◽

Intracellular Iron ◽

Iron Deprivation ◽

Iron Transporter ◽

Zip Family ◽

Intracellular Expression

Infection of mammalian hosts with Leishmania amazonensis depends on the remarkable ability of these parasites to replicate within macrophage phagolysosomes. A critical adaptation for survival in this harsh environment is an efficient mechanism for gaining access to iron. In this study, we identify and characterize LIT1, a novel L. amazonensis membrane protein with extensive similarity to IRT1, a ZIP family ferrous iron transporter from Arabidopsis thaliana. The ability of LIT1 to promote iron transport was demonstrated after expression in yeast and in L. amazonensis LIT1-null amastigotes. Endogenous LIT1 was only detectable in amastigotes replicating intracellularly, and its intracellular expression was accelerated under conditions predicted to result in iron deprivation. Although L. amazonensis lacking LIT1 grew normally in axenic culture and had no defects differentiating into infective forms, replication within macrophages was abolished. Consistent with an essential role for LIT1 in intracellular growth as amastigotes, Δlit1 parasites were avirulent. After inoculation into highly susceptible mice, no lesions were detected, even after extensive periods of time. Despite the absence of pathology, viable Δlit1 parasites were recovered from the original sites of inoculation, indicating that L. amazonensis can persist in vivo independently of the ability to grow in macrophages. Our findings highlight the essential role played by intracellular iron acquisition in Leishmania virulence and identify this pathway as a promising target for therapeutic intervention.

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High-through identification of T cell-specific phage-exposed mimotopes using PBMCs from tegumentary leishmaniasis patients and their use as vaccine candidates against Leishmania amazonensis infection

Parasitology ◽

10.1017/s0031182018001403 ◽

2018 ◽

Vol 146 (3) ◽

pp. 322-332 ◽

Cited By ~ 4

Author(s):

Gerusa B. Carvalho ◽

Lourena E. Costa ◽

Daniela P. Lage ◽

Fernanda F. Ramos ◽

Thaís T. O. Santos ◽

...

Keyword(s):

Healthy Subjects ◽

Mononuclear Cells ◽

Parasite Load ◽

Leishmania Amazonensis ◽

Interleukin 4 ◽

Peripheral Blood Mononuclear ◽

Th1 Response ◽

Tegumentary Leishmaniasis

AbstractIn the current study, phage-exposed mimotopes as targets against tegumentary leishmaniasis (TL) were selected by means of bio-panning cycles employing sera of TL patients and healthy subjects, besides the immune stimulation of peripheral blood mononuclear cells (PBMCs) collected from untreated and treated TL patients and healthy subjects. The clones were evaluated regarding their specific interferon-γ (IFN-γ) and interleukin-4 (IL-4) production in the in vitro cultures, and selectivity and specificity values were calculated, and those presenting the best results were selected for the in vivo experiments. Two clones, namely A4 and A8, were identified and used in immunization protocols from BALB/c mice to protect against Leishmania amazonensis infection. Results showed a polarized Th1 response generated after vaccination, being based on significantly higher levels of IFN-γ, IL-2, IL-12, tumour necrosis factor-α (TNF-α) and granulocyte-macrophage colony-stimulating factor (GM-CSF); which were associated with lower production of specific IL-4, IL-10 and immunoglobulin G1 (IgG1) antibodies. Vaccinated mice presented significant reductions in the parasite load in the infected tissue and distinct organs, when compared with controls. In conclusion, we presented a strategy to identify new mimotopes able to induce Th1 response in PBMCs from TL patients and healthy subjects, and that were successfully used to protect against L. amazonensis infection.

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Improvement of In Vitro and In Vivo Antileishmanial Activities of 2′,6′-Dihydroxy-4′-Methoxychalcone by Entrapment in Poly(d,l-Lactide) Nanoparticles

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.43.7.1776 ◽

1999 ◽

Vol 43 (7) ◽

pp. 1776-1778 ◽

Cited By ~ 41

Author(s):

Eduardo Caio Torres-Santos ◽

José M. Rodrigues ◽

Davyson L. Moreira ◽

Maria Auxiliadora C. Kaplan ◽

Bartira Rossi-Bergmann

Keyword(s):

Polymeric Nanoparticles ◽

Leishmania Amazonensis ◽

Antileishmanial Activity ◽

Antileishmanial Activities ◽

Piper Aduncum

ABSTRACT The inhibition of intracellular Leishmania amazonensisgrowth by 2′,6′-dihydroxy-4′-methoxychalcone (DMC) isolated fromPiper aduncum was further enhanced after encapsulation of DMC in polymeric nanoparticles. Encapsulated DMC also showed increased antileishmanial activity in infected BALB/c mice, as evidenced by significantly smaller lesions and fewer parasites in the lesions.

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