105 research outputs found
Histological study of cell migration in the dermis of hamsters after immunisation with two different vaccines against visceral leishmaniasis.
Vaccine candidates, including live and/or killed parasites, Leishmania -purified fractions, defined recombinant antigens and antigen-encoding DNA-plasmids have been proposed to use as vaccine anti- Leishmania . More recently, the hamsters have been used to pre selection of antigens candidate to apply in further experiments using canine model. In this report we evaluated the kinetics of cell migration in dermal inflammatory infiltrate, circulating leukocytes and the presence of nitric oxide (NO)/induced nitric oxide synthase during the early (1?24 h) and late (48?168 h) periods following inoculation of hamsters with antigenic components of anti-canine visceral leishmaniasis vaccines Leishmune 1 and Leishmania braziliensis antigen (LB) with and without saponin (Sap) adjuvant. Our results show that LB caused an early reduction of lymphocytes in the dermis while Sap and LBSap triggered a late recruitment, suggesting the role of the adjuvant in the traffic of antigen-presenting cells and the induction of lymphocyte migration. In that manner our results suggest that the kinetics of cell migration on hamster model may be of value in the selection of vaccine antigens prior the tests in dogs particularly in respect of the toxicity of the preparations
Immunotherapy and immunochemotherapy in visceral leishmaniasis : promising treatments for this neglected disease.
Leishmaniasis has several clinical forms: self-healing or chronic cutaneous leishmaniasis
or post-kala-azar dermal leishmaniasis; mucosal leishmaniasis; visceral leishmaniasis (VL),
which is fatal if left untreated.The epidemiology and clinical features of VL vary greatly due
to the interaction of multiple factors including parasite strains, vectors, host genetics, and
the environment. Human immunodeficiency virus infection augments the severity of VL
increasing the risk of developing active disease by 100?2320 times. An effective vaccine
for humans is not yet available. Resistance to chemotherapy is a growing problem in many
regions, and the costs associated with drug identification and development, make commercial
production for leishmaniasis, unattractive.The toxicity of currently drugs, their long
treatment course, and limited efficacy are significant concerns. For cutaneous disease,
many studies have shown promising results with immunotherapy/immunochemotherapy,
aimed to modulate and activate the immune response to obtain a therapeutic cure. Nowadays,
the focus of many groups centers on treating canine VL by using vaccines and
immunomodulators with or without chemotherapy. In human disease, the use of cytokines
like interferon-g associated with pentavalent antimonials demonstrated promising results
in patients that did not respond to conventional treatment. In mice, immunomodulation
based on monoclonal antibodies to remove endogenous immunosuppressive cytokines
(interleukin-10) or block their receptors, antigen-pulsed syngeneic dendritic cells, or biological
products like Pam3Cys (TLR ligand) has already been shown as a prospective treatment
of the disease. This review addresses VL treatment, particularly immunotherapy and/or
immunochemotherapy as an alternative to conventional drug treatment in experimental
models, canine VL, and human disease
Impact of dose and surface features on plasmatic and liver concentrations of biodegradable polymeric nanocapsules.
The effect of polymeric nanocapsule dose on plasmatic and liver concentrations 20 min after intravenous
administration in mice was evaluated. Nanocapsules were prepared with different polymers, namely, poly(D,Llactide)
(PLA), polyethylene glycol-block-poly(D,L-lactide) (PLA-PEG), and PLA with chitosan (PLA-Cs) and
compared with a nanoemulsion. These formulations were labelled with a phthalocyanine dye for fluorescent
detection. The nanostructures had narrow size distributions upon separation by asymmetric flow field flow
fractionation with static and dynamic light scattering detection, with average hydrodynamic diameters in the
130?300 nm range, negative zeta potentials, except PLA-Cs nanocapsules, which had a positive zeta potential.
Flow cytometry revealed uptake mostly by monocytes and neutrophils in mice and human blood. PLA
nanocapsules and the nanoemulsion showed dose-dependent plasma concentrations, where the percentage of
plasmatic fluorescence increased with increasing administered dose. In contrast, PLA-PEG nanocapsules led to a
dose-independent plasmatic profile. PLA-Cs nanocapsules showed the lowest plasmatic and liver levels of
fluorescence at all administered doses and significant intravenous toxicity in mice. This work demonstrates the
importance of considering the nanocarrier dose when evaluating pharmacokinetic and biodistribution data and
emphasizes the role of surface features in determining the plasmatic and liver concentrations of a poorly soluble
lipophilic encapsulated compound
Comparative genomics of canine-isolated Leishmania (Leishmania) amazonensis from an endemic focus of visceral leishmaniasis in Governador Valadares, southeastern Brazil.
Leishmaniasis is a highly diverse group of diseases caused by kinetoplastid of the genus Leishmania.
These parasites are taxonomically diverse, with human pathogenic species separated into two
subgenera according to their development site inside the alimentary tract of the sand fly insect
vector. The disease encompasses a variable spectrum of clinical manifestations with tegumentary or
visceral symptoms. Among the causative species in Brazil, Leishmania (Leishmania) amazonensis is an
important etiological agent of human cutaneous leishmaniasis that accounts for more than 8% of all
cases in endemic regions. L. (L.) amazonensis is generally found in the north and northeast regions of
Brazil. Here, we report the first isolation of L. (L.) amazonensis from dogs with clinical manifestations
of visceral leishmaniasis in Governador Valadares, an endemic focus in the southeastern Brazilian State
of Minas Gerais where L. (L.) infantum is also endemic. These isolates were characterized in terms of
SNPs, chromosome and gene copy number variations, confirming that they are closely related to a
previously sequenced isolate obtained in 1973 from the typical Northern range of this species. The
results presented in this article will increase our knowledge of L. (L.) amazonensis-specific adaptations
to infection, parasite survival and the transmission of this Amazonian species in a new endemic area of
Brazil
Synthetic peptides elicit strong cellular immunity in Visceral Leishmaniasis natural reservoir and contribute to long-lasting polyfunctional T-cells in BALB/c mice.
Reverse vaccinology or immunoinformatics is a computational methodology which integrates data from in silico epitope prediction, associated to other important information as, for example, the predicted subcellular location of the proteins used in the design of the context of vaccine development. This approach has the potential to search for new targets for vaccine development in the predicted proteome of pathogenic organisms. To date, there is no effective vaccine employed in vaccination campaigns against visceral leishmaniasis (VL). For the first time, herein, an in silico, in vitro, and in vivo peptide screening was performed, and immunogenic peptides were selected to constitute VL peptide-based vaccines. Firstly, the screening of in silico potential peptides using dogs naturally infected by L. infantum was conducted and the peptides with the best performance were selected. The mentioned peptides were used to compose Cockt-1 (cocktail 1) and Cockt-2 (cocktail 2) in combination with saponin as the adjuvant. Therefore, tests for immunogenicity, polyfunctional T-cells, and the ability to induce central and effector memory in T-lymphocytes capacity in reducing the parasite load on the spleen for Cockt-1 and Cockt-2 were performed. Among the vaccines under study, Cockt-1 showed the best results, eliciting CD4+ and CD8+ polyfunctional T-cells, with a reduction in spleen parasitism that correlates to the generation of T CD4+ central memory and T CD8+ effector memory cells. In this way, our findings corroborate the use of immunoinformatics as a tool for the development of future vaccines against VL
A vaccine therapy for canine visceral leishmaniasis promoted significant improvement of clinical and immune status with reduction in parasite burden.
Herein, we evaluated the treatment strategy employing a therapeutic heterologous
vaccine composed of antigens of Leishmania braziliensis associated with MPL adjuvant
(LBMPL vaccine) for visceral leishmaniasis (VL) in symptomatic dogs naturally infected
by Leishmania infantum. Sixteen dogs received immunotherapy with MPL adjuvant
(n = 6) or with a vaccine composed of antigens of L. braziliensis associated with MPL
(LBMPL vaccine therapy, n = 10). Dogs were submitted to an immunotherapeutic
scheme consisting of 3 series composed of 10 subcutaneous doses with 10-day
interval between each series. The animals were evaluated before (T0) and 90 days after
treatment (T90) for their biochemical/hematological, immunological, clinical, and parasitological
variables. Our major results showed that the vaccine therapy with LBMPL
was able to restore and normalize main biochemical (urea, AST, ALP, and bilirubin)
and hematological (erythrocytes, hemoglobin, hematocrit, and platelets) parameters.
In addition, in an ex vivo analysis using flow cytometry, dogs treated with LBMPL
vaccine showed increased CD3+ T lymphocytes and their subpopulations (TCD4+ and
TCD8+), reduction of CD21+ B lymphocytes, increased NK cells (CD5?CD16+) and
CD14+ monocytes. Under in vitro conditions, the animals developed a strong antigen-
specific lymphoproliferation mainly by TCD4+ and TCD8+ cells; increasing in both
TCD4+IFN-?+ and TCD8+IFN-?+ as well as reduction of TCD4+IL-4+ and TCD8+IL-4+
lymphocytes with an increased production of TNF-? and reduced levels of IL-10.
Concerning the clinical signs of canine visceral leishmaniasis, the animals showed an important reduction in the number and intensity of the disease signs; increase body
weight as well as reduction of splenomegaly. In addition, the LBMPL immunotherapy
also promoted a reduction in parasite burden assessed by real-time PCR. In the bone
marrow, we observed seven times less parasites in LBMPL animals compared with
MPL group. The skin tissue showed a reduction in parasite burden in LBMPL dogs
127.5 times higher than MPL. As expected, with skin parasite reduction promoted by
immunotherapy, we observed a blocking transmission to sand flies in LBMPL dogs
with only three positive dogs after xenodiagnosis. The results obtained in this study
highlighted the strong potential for the use of this heterologous vaccine therapy as an
important strategy for VL treatment
A vaccine therapy for canine visceral leishmaniasis promoted significant improvement of clinical and immune status with reduction in parasite burden.
Herein, we evaluated the treatment strategy employing a therapeutic heterologous
vaccine composed of antigens of Leishmania braziliensis associated with MPL adjuvant
(LBMPL vaccine) for visceral leishmaniasis (VL) in symptomatic dogs naturally infected
by Leishmania infantum. Sixteen dogs received immunotherapy with MPL adjuvant
(n = 6) or with a vaccine composed of antigens of L. braziliensis associated with MPL
(LBMPL vaccine therapy, n = 10). Dogs were submitted to an immunotherapeutic
scheme consisting of 3 series composed of 10 subcutaneous doses with 10-day
interval between each series. The animals were evaluated before (T0) and 90 days after
treatment (T90) for their biochemical/hematological, immunological, clinical, and parasitological
variables. Our major results showed that the vaccine therapy with LBMPL
was able to restore and normalize main biochemical (urea, AST, ALP, and bilirubin)
and hematological (erythrocytes, hemoglobin, hematocrit, and platelets) parameters.
In addition, in an ex vivo analysis using flow cytometry, dogs treated with LBMPL
vaccine showed increased CD3+ T lymphocytes and their subpopulations (TCD4+ and
TCD8+), reduction of CD21+ B lymphocytes, increased NK cells (CD5?CD16+) and
CD14+ monocytes. Under in vitro conditions, the animals developed a strong antigen-
specific lymphoproliferation mainly by TCD4+ and TCD8+ cells; increasing in both
TCD4+IFN-?+ and TCD8+IFN-?+ as well as reduction of TCD4+IL-4+ and TCD8+IL-4+
lymphocytes with an increased production of TNF-? and reduced levels of IL-10.
Concerning the clinical signs of canine visceral leishmaniasis, the animals showed an important reduction in the number and intensity of the disease signs; increase body
weight as well as reduction of splenomegaly. In addition, the LBMPL immunotherapy
also promoted a reduction in parasite burden assessed by real-time PCR. In the bone
marrow, we observed seven times less parasites in LBMPL animals compared with
MPL group. The skin tissue showed a reduction in parasite burden in LBMPL dogs
127.5 times higher than MPL. As expected, with skin parasite reduction promoted by
immunotherapy, we observed a blocking transmission to sand flies in LBMPL dogs
with only three positive dogs after xenodiagnosis. The results obtained in this study
highlighted the strong potential for the use of this heterologous vaccine therapy as an
important strategy for VL treatment
In vivo antileishmanial efficacy of a naphthoquinone derivate incorporated into a Pluronic? F127-based polymeric micelle system against Leishmania amazonensis infection.
New therapeutic strategies against leishmaniasis are desirable, since the treatment against disease presents
problems, such as the toxicity, high cost and/or parasite resistance. As consequence, new antileishmanial
compounds are necessary to be identified, as presenting high activity against Leishmania parasites, but low
toxicity in mammalian hosts. Flau-A is a naphthoquinone derivative recently showed to presents an in vitro
effective action against Leishmania amazonensis and L. infantum species. In the present work, the in vivo efficacy
of Flau-A, which was incorporated into a Poloxamer 407-based micelle system, was evaluated in a murine model
against L. amazonensis infection. Amphotericin B (AmB) and Ambisome? were used as controls. The animals were
infected and later treated with the compounds. Thirty days after the treatment, parasitological and immunological parameters were evaluated. Results showed that AmB, Ambisome?
, Flau-A or Flau-A/M-treated
animals presented significantly lower average lesion diameter and parasite burden in tissue and organs evaluated, when compared to the control (saline and micelle) groups. Flau-A or Flau-A/M-treated mice were those
presenting the most significant reductions in the parasite burden, when compared to the others. These animals
developed also a more polarized antileishmanial Th1 immune response, which was based on significantly higher
levels of IFN-?, IL-12, TNF-?, GM-CSF, and parasite-specific IgG2a isotype; associated with low levels of IL-4, IL10, and IgG1 antibody. The absence of toxicity was found in these animals, although mice receiving AmB have
showed high levels of renal and hepatic damage markers. In conclusion, results suggested that the Flau-A/M
compound may be considered as a possible therapeutic target to be evaluated against human leishmaniasis
Vaccination with a CD4+ and CD8+ T-cell epitopes-based recombinant chimeric protein derived from Leishmania infantum proteins confers protective immunity against visceral leishmaniasis.
Vaccination seems to be the best approach to control visceral leishmaniasis (VL). Resistance against infection is based on the development of a Th1 immune response characterized by the production of interferons-? (IFN-?), interleukin-12 (IL-12), granulocyte-macrophage-colony-stimulating factor (GM-CSF), and tumor necrosis factor-? (TNF-?), among others. A number of antigens have been tested as potential targets against the disease; few of them are able to stimulate human immune cells. In the present study, 1 prediction of MHC class I and II molecules-specific epitopes in the amino acid sequences of 3 Leishmania proteins: 1 hypothetical, prohibitin, and small glutamine-rich tetratricopeptide repeat-containing proteins, was performed using bioinformatics tools, and a T-cell epitopes-based recombinant chimeric protein was constructed, synthetized and purified to be evaluated in invitro and in vivo experiments. The purified protein was tested regarding its immunogenicity in peripheral blood mononuclear cells (PBMCs) from healthy subjects and VL patients, as well as to its immunogenicity and protective efficacy in a murine model against Leishmania infantum infection. Results showed a Th1 response based on high IFN-? and low IL-10 levels derived from in chimera-stimulated PBMCs in both healthy subjects and VL patients. In addition, chimera and/or saponin-immunized mice presented significantly lower parasite burden in distinct evaluated organs, when compared to the controls, besides higher levels of IFN-?, IL-2, IL-12, and GM-CSF, and an IgG2a isotype-based humoral response. In addition, the CD4+ and CD8+ T-cell subtypes contributed to IFN-? production in the protected animals. The results showed the immunogenicity in human cells and the protective efficacy against L. infantum in a murine model, and well indicate that this recombinant chimera can be considered as a promising strategy to be used against human disease
Prevalence and Factors Associated with Leishmania infantum Infection of Dogs from an Urban Area of Brazil as Identified by Molecular Methods
Visceral leishmaniasis (VL) is a disease caused by the parasite Leishmania infantum, and dogs are the most important domestic reservoirs of the agent. During recent decades, VL has expanded to large Brazilian urban centers. In the present work, we have demonstrated by using molecular techniques that the rate of canine infection as detected by serology has been considerably underestimated. Two groups of seronegative dogs (infected and non-infected according to molecular methods) were further evaluated from data obtained through interviews with owners of the animals. The factors associated with Leishmania infection in dogs were a family income of less than two minimum salaries, the knowledge of the owner regarding the vector, the dog spending most of its time in the backyard and the dog never having had a previous serological examination. Awareness regarding the factors associated with canine infection will improve health services and the understanding of the disease's expansion in urban areas
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