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Leishmania major Adenylate kinase immunization offers partial protection to a susceptible host
Abstract
Leishmania major causes mild to severe cutaneous lesions resulting in significant disfigurations, if untreated. The drugs are toxic and drug-resistance parasites are emerging. Therefore, a prophylactic vaccination is an urgent need. As no vaccine is available, we compared the genes expressed by virulent and avirulent parasites. We identify L. major adenylate kinase (AdeK) as a probable vaccine candidate after a series of experimentations. We cloned the gene in mammalian pcDNA6/HisA and pet28a+ vector for in vivo expression following immunization and in vitro protein expression for booster, respectively. We observed that immunization of susceptible BALB/c mice with AdeK resulted in significant protection against L. major challenge infection. The protection was accompanied increased IFN-γ producing lymphoctyes and reduced IL-4, IL-17 and IL-10 secreting central and effector Th2, Th17 and Treg memory cells respectively. These observations indicate L. major AdeK as a potential vaccine candidate.
... The role of ADKs has been studied in Schistosoma japonicum (Gao et al. 2017(Gao et al. , 2020, Schistosoma mansoni (de Almeida Marques et al. 2012), Clonorchis sinensis (Kwon et al. 2018;Liang et al. 2013), Toxoplasma gondii (Recacha et al. 2000), Plasmodium falciparum (Ma et al. 2012;Taku et al. 2021), Leishmania spp. (Kulkarni et al. 2019;Villa et al. 2003;Zutshi et al. 2020), and Trypanosoma brucei (Bouvier et al. 2006). Some studies have shown that ADKs play a vital role in maintaining the energy metabolism of the parasites and promoting their growth and development. ...
... S. japonicum ADK1 can provide partial protective immunity against S. japonicum infection (Gao et al. 2017), and Leishmania ADK1 has been proposed as a potential vaccine candidate (Zutshi et al. 2020). According to the present findings, EgADK1 and EgADK8 not only have multiple phosphorylation sites, but also are localized in various developmental stages of E. granulosus s.s. ...
Adenylate kinases (ADKs) are one of the important enzymes regulating adenosine triphosphate (ATP) metabolism in Echinococcus granulosus sensu lato. The objective of the present study was to explore the molecular characteristics and immunological properties of E. granulosus sensu stricto (G1) adenylate kinase 1 (EgADK1) and adenylate kinase 8 (EgADK8). EgADK1 and EgADK8 were cloned and expressed, and the molecular characteristics of EgADK1 and EgADK8 were analyzed through different bioinformatics tools. Western blotting was used to examine the reactogenicity of recombinant adenylate kinase 1 (rEgADK1) and recombinant adenylate kinase 8 (rEgADK8) and to evaluate their diagnostic value. The expression profiles of EgADK1 and EgADK8 in 18-day-old strobilated worms and protoscoleces were analyzed by quantitative real-time PCR, and their distribution in 18-day-old strobilated worms, the germinal layer, and protoscoleces was determined by immunofluorescence localization. EgADK1 and EgADK8 were successfully cloned and expressed. Bioinformatics analysis predicted that EgADK1 and EgADK8 have multiple phosphorylation sites and B-cell epitopes. Compared with EgADK8, EgADK1 and other parasite ADKs have higher sequence similarity. In addition, both cystic echinococcosis (CE)–positive sheep sera and Cysticercus tenuicollis–infected goat sera could recognize rEgADK1 and rEgADK8. EgADK1 and EgADK8 were localized in protoscoleces, the germinal layer, and 18-day-old strobilated worms. EgADK1 and EgADK8 showed no significant difference in their transcription level in 18-day-old strobilated worms and protoscoleces, suggesting that EgADK1 and EgADK8 may play an important role in the growth and development of E. granulosus sensu lato. Since EgADK1 and EgADK8 can be recognized by other parasite-positive sera, they are not suitable as candidate antigens for the diagnosis of CE.
... The target protein is tagged with an array of histidine residues and bound protein is then purified by passing imidazole, which replaces the protein-Ni 2+ interaction owing to its high affinity with metal ions. In vaccination studies, this technique is used for purification of the recombinant form of antigenic proteins [44]. ...
Search for an efficacious antileishmanial vaccine has led to clinical trials of numerous vaccine candidates in the past few decades. As no promising candidate has emerged from these studies, novel vaccine modalities and vaccine assessment techniques are still emerging for antileishmanial vaccine development. Briefly, this chapter discusses: (a) history and timeline of antileishmanial vaccine development; (b) techniques utilized for developing whole-parasite and subunit-based antileishmanial vaccine formulations, and (c) immunogenicity and post-challenge protective efficacy assessment of vaccine candidates.
Leishmaniasis is an enigmatic disease that has very restricted options for chemotherapy and none for prophylaxis. As a result, deriving therapeutic principles for curing the disease has been a major objective in Leishmania research for a long time. Leishmania is a protozoan parasite that lives within macrophages by subverting or switching cell signaling to the pathways that ensure its intracellular survival. Therefore, three groups of molecules aimed at blocking or eliminating the parasite, at least, in principle, include blockers of macrophage receptor- Leishmania ligand interaction, macrophage-activating small molecules, peptides and cytokines, and signaling inhibitors or activators. Macrophages also act as an antigen-presenting cell, presenting antigen to the antigen-specific T cells to induce activation and differentiation of the effector T cell subsets that either execute or suppress anti-leishmanial functions. Three groups of therapeutic principles targeting this sphere of Leishmania-macrophage interaction include antibodies that block pro-leishmanial response of T cells, ligands that activate anti-leishmanial T cells and the antigens for therapeutic vaccines. Besides these, prophylactic vaccines have been in clinical trials but none has succeeded so far. Herein, we have attempted to encompass all these principles and compose a comprehensive review to analyze the feasibility and adoptability of different therapeutics for leishmaniasis.
Leishmaniasis is endemic in at least 98 countries. Due to the high toxicity and resistance associated with the drugs, we chose lignans as an alternative, due to their favorable properties of absorption, distribution, metabolism, excretion, and toxicity (ADMET). To investigate their leishmanicidal potential, the biological activities of a set of 160 lignans were predicted using predictive models that were built using data for Leishmania major and L. (Viannia) braziliensis. A combined analysis, based on ligand and structure, and several other computational approaches were used. The results showed that the combined analysis was able to select 11 lignans with potential activity against L. major and 21 lignans against L. braziliensis, with multitargeting effects and low or no toxicity. Of these compounds, four were isolated from the species Justicia aequilabris (Nees) Lindau. All of the identified compounds were able to inhibit the growth of L. braziliensis promastigotes, with the most active compound, (159) epipinoresinol-4-O-β-d-glucopyranoside, presenting an IC50 value of 5.39 µM and IC50 value of 36.51 µM for L. major. Our findings indicated the potential of computer-aided drug design and development and demonstrated that lignans represent promising prototype compounds for the development of multitarget drugs against leishmaniasis.
Masoud Ghorbani, Ramin Farhoudi Department of Viral Vaccine Production, Pasteur Institute of Iran, Research and Production Complex, Karaj, Iran Abstract: Leishmania is an obligate intracellular pathogen that invades phagocytic host cells. Approximately 30 different species of Phlebotomine sand flies can transmit this parasite either anthroponotically or zoonotically through their bites. Leishmaniasis affects poor people living around the Mediterranean Basin, East Africa, the Americas, and Southeast Asia. Affected regions are often remote and unstable, with limited resources for treating this disease. Leishmaniasis has been reported as one of the most dangerous neglected tropical diseases, second only to malaria in parasitic causes of death. People can carry some species of Leishmania for long periods without becoming ill, and symptoms depend on the form of the disease. There are many drugs and candidate vaccines available to treat leishmaniasis. For instance, antiparasitic drugs, such as amphotericin B (AmBisome), are a treatment of choice for leishmaniasis depending on the type of the disease. Despite the availability of different treatment approaches to treat leishmaniasis, therapeutic tools are not adequate to eradicate this infection. In the meantime, drug therapy has been limited because of adverse side effects and unsuccessful vaccine preparation. However, it can immediately make infections inactive. According to other studies, vaccination cannot eradicate leishmaniasis. There is no perfect vaccine or suitable drug to eradicate leishmaniasis completely. So far, no vaccine or drug has been provided to induce long-term protection and ensure effective immunity against leishmaniasis. Therefore, it is necessary that intensive research should be performed in drug and vaccine fields to achieve certain results. Keywords: leishmania, leishmania treatment, vaccine, recombinant antigens
Leishmaniasis is caused by an intracellular parasite transmitted to humans by the bite of a sand fly. It is endemic in Asia, Africa, the Americas, and the Mediterranean region. Worldwide, 1.5 to 2 million new cases occur each year, 350 million are at risk of acquiring the disease, and leishmaniasis causes 70,000 deaths per year. Clinical features depend on the species of Leishmania involved and the immune response of the host. Manifestations range from the localized cutaneous to the visceral form with potentially fatal outcomes. Many drugs are used in its treatment, but the only effective treatment is achieved with current pentavalent antimonials.
Leishmaniasis is an important disease mediated by the protozoan parasite Leishmania via the bite of the female sandfly insect vector. Leishmaniasis is endemic in the tropical and subtropical regions. The most common form of the disease is cutaneous leishmaniasis, which affects more than 10 million people worldwide and includes at least 1.5 million new cases every year. So far, treatment of the disease relies on unsatisfactory chemotherapy that can be complicated by the rising appearance of drug-resistant parasites. Furthermore, it is challenging to achieve solid control of the insect vector and animal reservoir. Therefore, the development of a safe and effective vaccine is urgently needed for the treatment and prevention of leishmaniasis. This review focuses on the recent advances in the development of a safe vaccine that could be used for prevention and treatment of cutaneous leishmaniasis. A short outlook for future research efforts is also presented.
No licensed human vaccines are currently available against any parasitic disease including leishmaniasis. Several anti-leishmanial vaccine formulations have been tested in various animal models including genetically modified live attenuated parasite vaccines. Experimental infection studies have shown that Leishmania parasites utilize a broad range of strategies to undermine effector properties of host phagocytic cells i.e., dendritic cells (DC) and macrophages (M). Further, Leishmania parasites have evolved strategies to actively inhibit TH1 polarizing functions of DCs, and to condition the infected M towards anti-inflammatory/alternative/M2 phenotype. The altered phenotype of phagocytic cells is characterized by decreased production of anti-microbial reactive oxygen, nitrogen molecules and pro-inflammatory cytokines such as IFN-, IL-12 and TNF-α. These early events limit the activation of TH1 effector cells and set the stage for pathogenesis. Further, this early control of innate immunity by the virulent parasites results in substantial alteration in the adaptive immunity characterized by reduced proliferation of CD4+ and CD8+ T cells, and TH2 biased immunity that results in production of anti-inflammatory cytokines such as TGF-, and IL-10. More recent studies have also documented the induction of co-inhibitory ligands such as CTLA-4, PD-L1, CD200 and Tim-3 that induce exhaustion and/or non-proliferation in antigen experienced T cells. Most of these studies focus on viral infections in chronic phase thus limiting the direct application of these results from these studies to parasitic infections and much less to parasitic vaccines. However, these studies suggest that vaccine induced protective immunity can be modulated using strategies that enhance the co-stimulation that might reduce the threshold necessary for T cell activation and conversely by strategies that reduce or block inhibitory molecules such as PD-L1 and CD200. In this review we will focus on the polarization of antigen presenting cells and subsequent role of co-stimulatory and co-inhibitory molecules in mediating vaccine induced immunity using live attenuated Leishmania parasites as specific examples.
Leishmania (L.) tropica is the causative agent of different forms of human leishmaniasis. There is little information about the role of Leishmania-specific antibodies in the immune response against L. tropica infection. The aim of this study is to evaluate the role of Leishmania-specific antibodies and their immunoglobulin G (IgG) isotypes in L. tropica infection.
L. tropica at two different doses (high dose, 10(6) parasites/mouse and low dose, 10(3) parasites/mouse) were used for infection of BALB/c mice. BALB/c mice infected with Leishmania major were used for comparison. Anti-Leishmania antibodies of the IgG1 and IgG2a isotypes were assayed by enzyme-linked immunosorbent assay.
Our data showed that (1) a higher parasite dose results in higher levels of antibody. (2) L. tropica infection results in a lower IgG1 antibody response, compared with L. major infection. (3) The IgG2a/IgG1 antibody response in L. tropica infection is higher than that in L. major infection.
A higher IgG2a/IgG1 ratio is associated with protective immune response in L. tropica infection. These data can help to approach the complex profile of immunity against L. tropica infection.
Copyright © 2015. Published by Elsevier B.V.
HLA class I molecules reflect the health state of cells to cytotoxic T cells by presenting a repertoire of endogenously derived peptides. However, the extent to which the proteome shapes the peptidome is still largely unknown. Here we present a high-throughput mass spectrometry based workflow that allows stringent and accurate identification of thousands of such peptides and direct determination of binding motifs. Applying the workflow to seven cancer cell lines and primary cells, yielded more than 22,000 unique HLA peptides across different allelic binding specificities. By computing a score representing the HLA class I sampling density, we show a strong link between protein abundance and HLA-presentation (P<0.0001). When analyzing over-presented proteins, those with at least five-fold higher density score than expected for their abundance, we noticed that they are degraded almost 3 hours faster than similar but non-presented proteins (top 20% abundance class; median half-life 20.8h vs. 23.6h, p<0.0001). This validates protein degradation as an important factor for HLA presentation. Ribosomal, mitochondrial respiratory chain and nucleosomal proteins are particularly well presented. Taking a set of proteins associated with cancer, we compared the predicted immunogenicity of previously validated T cell epitopes with other peptides from these proteins in our dataset. The validated epitopes indeed tend to have higher immunogenic scores than the other detected HLA peptides, suggesting the usefulness of combining MS-analysis with immunogenesis prediction for ranking and selecting peptides for therapeutic use.
Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
TLRs recognize pathogen-expressed Ags and elicit host-protective immune response. Although TLR2 forms heterodimers with TLR1 or TLR6, recognizing different ligands, differences in the functions of these heterodimers remain unknown. In this study, we report that in Leishmania major-infected macrophages, the expression of TLR1 and TLR2, but not TLR6, increased; TLR2-TLR2 association increased, but TLR2-TLR6 association diminished. Lentivirus-expressed TLR1-short hairpin RNA (shRNA) or TLR2-shRNA administration reduced, but TLR6-shRNA increased L. major infection in BALB/c mice. Corroboratively, Pam3CSK4 (TLR1-TLR2 ligand) and peptidoglycan (TLR2 ligand) increased L. major infection but reduced TLR9 expression, whereas pegylated bisacycloxypropylcysteine (BPPcysMPEG; TLR2-TLR6 ligand) reduced L. major number in L. major-infected macrophages, accompanied by increased TLR9 expression, higher IL-12 production, and inducible NO synthase expression. Whereas MyD88, Toll/IL-1R adaptor protein, and TNFR-α-associated factor 6 recruitments to TLR2 were not different in Pam3CSK4-, peptidoglycan-, or BPPcysMPEG-treated macrophages, only BPPcysMPEG enhanced p38MAPK and activating transcription factor 2 activation. BPPcysMPEG conferred antileishmanial functions to L. major-infected BALB/c-derived T cells in a macrophage-T cell coculture and in BALB/c mice; the protection was TLR6 dependent and IL-12 dependent, and it was accompanied by reduced regulatory T cell number. BPPcysMPEG administration during the priming with fixed L. major protected BALB/c mice against challenge L. major infection; the protection was accompanied by low IL-4 and IL-10, but high IFN-γ productions and reduced regulatory T cells. Thus, BPPcysMPEG, a novel diacylated lipopeptide ligand for TLR2-TLR6 heterodimer, induces IL-12-dependent, inducible NO synthase-dependent, T-reg-sensitive antileishmanial protection. The data reveal a novel dimerization partner-dependent duality in TLR2 function.
Leishmaniasis is a neglected disease resulting in a global morbidity of 2,090 thousand Disability-Adjusted Life Years and a mortality rate of approximately 60,000 per year. Among the three clinical forms of leishmaniasis (cutaneous, mucosal, and visceral), visceral leishmaniasis (VL) accounts for the majority of mortality, as if left untreated VL is almost always fatal. Caused by infection with Leishmania donovani or L. infantum, VL represents a serious public health problem in endemic regions and is rapidly emerging as an opportunistic infection in HIV patients. To date, no vaccine exists for VL or any other form of leishmaniasis. In endemic areas, the majority of those infected do not develop clinical symptoms and past infection leads to robust immunity against reinfection. Thus the development of vaccine for Leishmania is a realistic public health goal, and this paper summarizes advances in vaccination strategies against VL.
An effective vaccine usually requires more than one time immunization in the form of prime-boost. Traditionally the same vaccines are given multiple times as homologous boosts. New findings suggested that prime-boost can be done with different types of vaccines containing the same antigens. In many cases such heterologous prime-boost can be more immunogenic than homologous prime-boost. Heterologous prime-boost represents a new way of immunization and will stimulate better understanding on the immunological basis of vaccines.
Resistance to leishmaniasis in C57BL/6 mice depends on Th1/Tc1 cells. BALB/c mice preferentially develop Th2 immunity and succumb to infection. We now assessed the role of IL-17 in cutaneous leishmaniasis. During the course of Leishmania major infection, BALB/c CD4 cells and neutrophils produced increased amounts of IL-17 as compared with cells from C57BL/6 mice. This increase was associated with significantly increased IL-23 release from L. major-infected BALB/c dendritic cells (DC), whereas IL-6 and TGF-beta1 production by BALB/c and C57BL/6 DC were comparable. Interestingly, lesion sizes in infected IL-17-deficient BALB/c mice were dramatically smaller and failed to progress as compared with those in control mice. Similar amounts of IL-4, IL-10, and IFN-gamma were produced by T cells from IL-17-deficient mice and control mice consistent with development of Th2-predominant immunity in all animals. Improved disease outcome was associated with decreased CXCL2-accumulation in lesion sites and decreased neutrophil immigration into lesions of infected IL-17-deficient mice confirming prior observations that enhanced neutrophil recruitment contributes to disease susceptibility in BALB/c mice. This study excludes an important facilitating role for IL-17 in Th1/Th2 development in L. major-infected BALB/c mice, and suggests that IL-23 production by L. major-infected DC maintains IL-17(+) cells that influence disease progression via regulation of neutrophil recruitment.
Determination of antigen-specific cytokine responses of T lymphocytes after vaccination is made difficult by the low frequency
of responder cells. In order to detect these responses, the profile of intracellular cytokines was analyzed using flow cytometry
after antigenic expansion. Peripheral blood mononuclear cells were stimulated with antigens for 5 days, further expanded with
interleukin (IL)-2, and then restimulated on day 10. Cytokine production was detected by intracellular staining with monoclonal
antibodies after saponin-based permeabilization. Influenza expansion resulted in specific interferon-γ (IFN-γ) production of 6%–20%, with less IL-4 production (0%–2%). Tetanus toxoid resulted in even greater production. IL-4 and IFN-γ were produced mainly by memory cells of the CD45RO+ phenotype. IFN-γ production was contributed by both CD4 and CD8 populations. These methods were then applied to a clinical trial of a candidate
human immunodeficiency virus type 1 vaccine. Antigen-specific increases in IFN-γ were measured, which corresponded to antibody production, lymphoproliferation, and skin testing.
The acquisition of immunity following subclinical or resolved infection with the intracellular parasite Leishmania donovani suggests that vaccination could prevent visceral leishmaniasis (VL). The LACK (Leishmania homolog of receptors for activated C kinase) antigen is of interest as a vaccine candidate for the leishmaniases because
of its immunopathogenic role in murine L. major infection. Immunization of mice with a truncated (24-kDa) version of the 36-kDa LACK antigen, delivered in either protein
or DNA form, was found previously to protect against cutaneous L. major infection by redirecting the early T-cell response away from a pathogenic interleukin-4 (IL-4) response and toward a protective
Th1 response. The amino acid sequence of theLeishmania p36(LACK) antigen is highly conserved, but the efficacy of this vaccine antigen in preventing disease caused by strains other
than L. major has not been determined. We investigated the efficacy of a p36(LACK) DNA vaccine against VL because of the serious nature
of this form of leishmaniasis and because it was unclear whether the LACK vaccine would be effective in a model where there
was not a dominant pathogenic IL-4 response. We demonstrate here that although the LACK DNA vaccine induced a robust parasite-specific
Th1 immune response (IFN-γ but not IL-4 production) and primed for an in vivo T-cell response to inoculated parasites, it
did not induce protection against cutaneous or systemic L. donovanichallenge. Coadministration of IL-12 DNA with the vaccine did not enhance the strong vaccine-induced Th1 response or augment
a protective effect.
Macrophages play host to Leishmania major, a parasite that causes leishmaniasis in 500,000 people annually. Macrophage-expressed CD40, a costimulatory molecule, induces interleukin-12 (IL-12)-dependent and interferon-gamma (IFN-gamma)-dependent host-protective immune responses to Leishmania and other intracellular pathogens. Paradoxically, IL-10, another CD40-induced cytokine in macrophages, promotes Leishmania infection. How CD40 signaling regulates the secretion of these two counteractive cytokines remains unknown. Here we show that weak CD40 signals induce extracellular stress-related kinase-1/2 (ERK-1/2)-dependent IL-10 expression, whereas stronger signals induce p38 mitogen-activated protein kinase (p38MAPK)-dependent IL-12 production. p38MAPK and ERK-1/2 therefore have counter-regulatory actions. Leishmania skews CD40 signaling toward ERK-1/2, inducing IL-10, which inhibits activation of CD40-induced p38MAPK and expression of inducible nitric oxide synthase-2 (iNOS-2) and IL-12. ERK-1/2 inhibition or IL-10 neutralization restores CD40-induced p38MAPK activation and parasite killing in macrophages and the BALB/c mouse, a susceptible host. These data uncover a new immune evasion strategy, whereby Leishmania differentially modulates CD40-engaged, reciprocally functioning signaling modules, and provide a new conceptual framework for immune homeostasis.
Long-term immunity to Leishmania may require the continued presence of parasites, but previous attempts to create attenuated parasites that persist without causing disease have had limited success. Since Leishmania major mutants that lack lipophosphoglycan and other secreted phosphoglycans, termed lpg2-, persist indefinitely in infected mice without inducing any disease, we tested their ability to provide protection to virulent L. major challenge. In response to leishmanial Ag stimulation, cells from lpg2--infected mice produced minimal levels of IL-4 and IL-10, as well as very low levels of IFN-gamma. Nevertheless, when BALB/c mice infected with lpg2- parasites were challenged with virulent L. major they were protected from disease. Thus, these findings report on attenuated parasites that may be used to induce long-term protection against leishmaniasis and indicate that the immunity induced can be maintained in the absence of a strong Th1 response.
This study reports the efficacy of a heterologous prime-boost vaccination using DNA and vaccinia viruses (Western Reserve
[WR] virus and modified [attenuated] vaccinia virus Ankara [MVA]) expressing the LACK antigen (Leishmania homologue of receptors for activated C kinase) and an intradermal murine infection model employing Leishmania infantum. At 1 month postinfection, vaccinated mice showed high levels of protection in the draining lymph node (240-fold reduction
in parasite burden) coupled with significant levels of gamma interferon (20 to 200 ng/ml) and tumor necrosis factor alpha/lymphotoxin
(8 to 134 pg/ml). Significant but lower levels of protection (6- to 30-fold) were observed in the spleen and liver. Comparable
levels of protection were found for mice boosted with either LACK-WR or LACK-MVA, supporting the use of an attenuated vaccinia
virus-based vaccine against human visceral leishmaniasis.
Leishmaniasis affects 12 million people, but there are no vaccines. Immunological correlates of vaccine efficacy are unclear. Polarized Th1 vs Th2 responses in Leishmania major-infected mice suggested that a shift in balance from IL-4 to IFN-gamma was the key to vaccine success. Recently, a role for IL-10 and regulatory T cells in parasite persistence was demonstrated, prompting re-evaluation of vaccine-induced immunity. We compared DNA/modified vaccinia virus Ankara heterologous prime-boost with Leishmania homolog of the receptor for activated C kinase (LACK) or tryparedoxin peroxidase (TRYP). Both induced low IL-4 and high IFN-gamma prechallenge. Strikingly, high prechallenge CD4 T cell-derived IL-10 predicted vaccine failure using LACK, whereas low IL-10 predicted protection with TRYP. The ratio of IFN-gamma:IL-10 was thus a clear prechallenge indicator of vaccine success. Challenge infection caused further polarization to high IL-10/low IFN-gamma with LACK and low IL-10/high IFN-gamma with TRYP. Ex vivo quantitative RT-PCR and in vitro depletion and suppression experiments demonstrated that Ag-driven CD4+ CD25+ T regulatory 1-like cells were the primary source of IL-10 in LACK-vaccinated mice. Anti-IL-10R treatment in vivo demonstrated that IL-10 was functional in determining vaccine failure, rendering LACK protective in the presence of high IFN-gamma/low IL-5 responses.
Cutaneous and visceral leishmaniasis are amongst the most devastating infectious diseases of our time, affecting millions of people worldwide. The treatment of these serious diseases rely on a few chemotherapeutic agents, most of which are of parenteral use and induce severe side-effects. Furthermore, rates of treatment failure are high and have been linked to drug resistance in some areas. Here, we reviewed data on current chemotherapy practice in leishmaniasis. Drug resistance and mechanisms of resistance are described as well as the prospects for applying drug combinations for leishmaniasis chemotherapy. It is clear that efforts for discovering new drugs applicable to leishmaniasis chemotherapy are essential. The main aspects on the various steps of drug discovery in the field are discussed.
Leishmania causes a spectrum of diseases that range from self-healing to fatal infections. Control of leishmania is dependent upon generating CD4+ Th1 cells that produce IFNγ, leading to macrophage activation and killing of the intracellular parasites. Following resolution of the disease, short-lived effector T cells, as well as long-lived central memory T cells and skin resident memory T cells, are retained and able to mediate immunity to a secondary infection. However, there is no vaccine for leishmaniasis, and the drugs used to treat the disease can be toxic and ineffective. While a live infection generates immunity, a successful vaccine will depend upon generating memory T cells that can be maintained without the continued presence of parasites. Since both central memory and skin resident memory T cells are long-lived, they may be the appropriate targets for a leishmaniasis vaccine.
The many factors impacting the efficacy of a vaccine can be broadly divided into three categories: features of the vaccine itself, including immunogen design, vaccine type, formulation, adjuvant and dosing; individual variations among vaccine recipients and vaccine administration-related parameters. While much literature exists related to vaccines, and recently systems biology has started to dissect the impact of individual subject variation on vaccine efficacy, few studies have focused on the role of vaccine administration-related parameters on vaccine efficacy. Parenteral and mucosal vaccinations are traditional approaches for licensed vaccines; novel vaccine delivery approaches, including needless injection and adjuvant formulations, are being developed to further improve vaccine safety and efficacy. This review provides a brief summary of vaccine administration-related factors, including vaccination approach, delivery route and method of administration, to gain a better understanding of their potential impact on the safety and immunogenicity of candidate vaccines.
BALB/c mice are susceptible to progressive infection with Leishmania major due to the preferential development of CD4(+) T cells that secrete Th2 cytokines. Although Th2 cell development and susceptibility are disrupted by blockade of CD86 function early in infection, CD28-deficient BALB/c mice remain susceptible to leishmaniasis. We therefore examined whether the alternative CD86 ligand, CTLA4, contributes to the expression of susceptibility. BALB/c mice treated for 2 weeks of infection with anti-CTLA4 monoclonal antibody developed more rapidly progressive disease than sham-treated mice, whereas normally resistant C57BL/6 mice were unaffected. The draining lymph node cells of anti-CTLA4-treated BALB/c mice produced up to sixfold more interleukin-4 (IL-4) and IL-13 than control mice in the first 2 weeks of infection, but IFN-gamma synthesis was reciprocally decreased. Anti-CTLA4 treatment of BALB/c mice pretreated with neutralizing anti-IL-4 antibody or genetically deficient in IL-4 also caused significant worsening of leishmaniasis. Exacerbation in IL-4 KO mice was associated with increased IL-13 and decreased gamma interferon (IFN-gamma) and inducible nitric oxide synthase (iNOS) mRNA expression in vivo. These data indicate that anti-CTLA4 antibody induced earlier and more-polarized Th2 responses in susceptible BALB/c mice infected with L. major. The mechanism of disease worsening was partially IL-4 independent, indicating that increased IL-13 and/or decreased IFN-gamma production may have disrupted nitric oxide-based microbicidal responses. We conclude that CTLA4 significantly modulates Th2 development in murine leishmaniasis and that the Th2-polarizing effects of anti-CTLA4 treatment result in IL-4-independent exacerbation of disease.
At the end of their growth in the sand fly, Leishmania parasites differentiate into the infective metacyclic promastigote stage, which is transmitted to the mammalian host. Thus, in experimental studies of parasite infectivity toward animals or macrophages, the use of purified metacyclics is generally preferred. While metacyclics of several Leishmania species can be efficiently purified with the aid of lectins or monoclonal antibodies, which differentially exploit stage-specific differences in the structure of the abundant surface glycolipid lipophosphoglycan (LPG), such reagents are unavailable for most species and they are unsuitable for studies involving LPG-deficient mutants. Here we describe a simple density gradient centrifugation method, which allows the rapid purification of infective metacyclic parasites from both wild-type and LPG-deficient Leishmania major. The purified metacyclic promastigotes are authentic, as judged by criteria such as their morphology, expression of the metacyclic-specific gene SHERP, and ability to invade and replicate within macrophages in vitro. Preliminary studies suggest that this method is applicable to other Leishmania species including L. donovani.