Ifeoma Okwor

Infection with arginase deficient Leishmania major reveals a parasite number-dependent and cytokine-independent regulation of host cellular arginase activity and disease pathogenesis

The balance between the products of l-arginine metabolism in macrophages regulates the outcome of Leishmania major infection. l-arginine can be oxidized by host inducible NO synthase to produce NO, which contributes to parasite killing. In contrast, l-arginine hydrolysis by host arginase blocks NO generation and provides polyamines, which can support parasite proliferation. Additionally, Leishmania encode their own arginase which has considerable potential to modulate infectivity and disease pathogenesis. In this study, we compared the infectivity and impact on host cellular immune response in vitro and in vivo of wild-type (WT) L. major with that of a parasite arginase null mutant (arg−) L. major. We found that arg− L. major are impaired in their macrophage infectivity in vitro independent of host inducible NO synthase activities. As with in vitro results, the proliferation of arg− L. major in animal infections was also significantly impaired in vivo, resulting in delayed onset of lesion development, attenuated pathology, and low parasite burden. Despite this attenuated pathology, the production of cytokines by cells from the draining lymph node of mice infected with WT and arg− L. major was similar at all times tested. Interestingly, in vitro and in vivo arginase levels were significantly lower in arg− than in WT-infected cases and were directly correlated with parasite numbers inside infected cells. These results suggest that Leishmania-encoded arginase enhances disease pathogenesis by augmenting host cellular arginase activities and that contrary to previous in vitro studies, the host cytokine response does not influence host arginase activity.

Vaccines and vaccination strategies against human cutaneous leishmaniasis

One might think that the development of a vaccine against cutaneous leishmaniasis would be relatively straightforward because the type of immune response required for protection is known and natural immunity occurs following recovery from primary infection. However, there is as yet no effective vaccine against the disease in humans. Although vaccination in murine studies has yielded promising results, these vaccines have failed miserably when tested in primates or humans. The reasons behind these failures are unknown and remain a major hurdle for vaccine design and development against cutaneous leishmaniasis. In contrast, recovery from natural, deliberate or experimental infections results in development of long-lasting immunity to re-infection. This so called infection-induced resistance is the strongest anti-Leishmania immunity known. Here, we briefly review the different approaches to vaccination against cutaneous leishmaniasis and argue that vaccines composed of genetically modified (attenuated) parasites, which induce immunity akin to infection-induced resistance, may provide best protection against cutaneous leishmaniasis in humans.

LIGHT Is Critical for IL-12 Production by Dendritic Cells, Optimal CD4+ Th1 Cell Response, and Resistance to Leishmania major

Although studies indicate LIGHT (lymphotoxin (LT)-like, exhibits inducible expression and competes with HSV glycoprotein D for herpes virus entry mediator (HVEM), a receptor expressed by T lymphocytes) enhances inflammation and T cell-mediated immunity, the mechanisms involved in this process remain obscure. In this study, we assessed the role of LIGHT in IL-12 production and development of CD4+ Th cells type one (Th1) in vivo. Bone marrow-derived dendritic cells from LIGHT−/− mice were severely impaired in IL-12p40 production following IFN-γ and LPS stimulation in vitro. Furthermore, blockade of LIGHT in vitro and in vivo with HVEM-Ig and LT β receptor (LTβR)-Ig leads to impaired IL-12 production and defective polyclonal and Ag-specific IFN-γ production in vivo. In an infection model, injection of HVEM-Ig or LTβR-Ig into the usually resistant C57BL/6 mice results in defective IL-12 and IFN-γ production and severe susceptibility to Leishmania major that was reversed by rIL-12 treatment. This striking susceptibility to L. major in mice injected with HVEM-Ig or LTβR-Ig was also reproduced in LIGHT−/− → RAG1−/− chimeric mice. In contrast, L. major-infected LTβ−/− mice do not develop acute disease, suggesting that the effect of LTβR-Ig is not due to blockade of membrane LT (LTα1β2) signaling. Collectively, our data show that LIGHT plays a critical role for optimal IL-12 production by DC and the development of IFN-γ-producing CD4+ Th1 cells and its blockade results in severe susceptibility to Leishmania major.

Inoculation of killed Leishmania major into immune mice rapidly disrupts immunity to a secondary challenge via IL-10-mediated process

Recovery from natural or experimental Leishmania major infection, the causative agent of cutaneous leishmaniasis, results in development of durable immunity in mice and humans that is manifested as rapid control of parasite replication and resolution of cutaneous lesion after secondary challenge. This form of “infection-induced” immunity is thought to occur naturally in endemic areas and is generally considered the gold standard for any effective vaccine against cutaneous leishmaniasis. To determine factors that might heighten or abrogate infection-induced immunity, we investigated the impact of inoculating dead antigen in the form of killed Leishmania parasites to healed mice. We show that inoculation of killed parasites into mice that resolved their primary virulent L. major infection results in rapid and relatively sustained loss of infection-induced immunity. This loss of immunity was not due to the inability of killed parasites to induce inflammatory responses (such as delayed type hypersensitivity), but it was related to their failure to induce robust IFN-γ response. Furthermore, inoculation of killed Leishmania parasites into healed mice led to rapid expansion of IL-10-producing CD4+CD25+Foxp3+ T cells in lymph nodes draining the primary infection site. Treatment with anti-CD25 or anti-IL-10R mAb abolished killed parasite-induced loss of immunity. Our study suggests that vaccination with killed parasites could predispose naturally immune individuals to become susceptible to new infections and/or disease reactivation. This may account for the lack of efficacy of such vaccines in field trials in endemic regions. These findings have important implications for vaccine design and vaccination strategies against human cutaneous leishmaniasis.

The immunology of Leishmania/HIV co-infection

Leishmaniases are emerging as an important disease in human immunodeficiency virus (HIV)–infected persons living in several sub-tropical and tropical regions around the world, including the Mediterranean. The HIV/AIDS pandemic is spreading at an alarming rate in Africa and the Indian subcontinent, areas with very high prevalence of leishmaniases. The spread of HIV into rural areas and the concomitant spread of leishmaniases to suburban/urban areas have helped maintain the occurrence of Leishmania/HIV co-infection in many parts of the world. The number of cases of Leishmania/HIV co-infection is expected to rise owing to the overlapping geographical distribution of the two infections. In Southwestern Europe, there is also an increasing incidence of Leishmania/HIV co-infection (particularly visceral leishmaniasis) in such countries as France, Italy, Spain and Portugal. Studies suggest that in humans, very complex mechanisms involving dysregulation of host immune responses contribute to Leishmania-mediated immune activation and pathogenesis of HIV. In addition, both HIV-1 and Leishmania infect and multiply within cells of myeloid or lymphoid origin, thereby presenting a perfect recipe for reciprocal modulation of Leishmania and HIV-1-related disease pathogenesis. Importantly, because recovery from leishmaniases is associated with long-term persistence of parasites at the primary infection sites and their draining lymph nodes, there is very real possibility that HIV-mediated immunosuppression (due to CD4+ T cell depletion) could lead to reactivation of latent infections (reactivation leishmaniasis) in immunocompromised patients. Here, we present an overview of the immunopathogenesis of Leishmania/HIV co-infection and the implications of this interaction on Leishmania and HIV disease outcome.

Protective Immunity and Vaccination Against Cutaneous Leishmaniasis

Although a great deal of knowledge has been gained from studies on the immunobiology of leishmaniasis, there is still no universally acceptable, safe, and effective vaccine against the disease. This strongly suggests that we still do not completely understand the factors that control and/or regulate the development and sustenance of anti-Leishmania immunity, particularly those associated with secondary (memory) immunity. Such an understanding is critically important for designing safe, effective, and universally acceptable vaccine against the disease. Here we review the literature on the correlate of protective anti-Leishmania immunity and vaccination strategies against leishmaniasis with a bias emphasis on experimental cutaneous leishmaniasis.

Social and Economic Burden of Human Leishmaniasis

Leishmaniasis continues to pose a major public health problem worldwide. With new epidemics occurring in endemic areas and the spread of the disease to previously free areas because of migration, tourism, and military activities, there is a great need for the development of an effective vaccine. Leishmaniasis is a disease of the poor, occurring mostly in remote rural villages with poor housing and little or no access to modern health-care facilities. In endemic areas, diagnosis of any form of leishmaniasis puts a huge financial strain on an already meagre financial resource at both the individual and community levels. Most often families need to sell their assets (land and livestock) or take loans from informal financial outfits with heavy interest rates to pay for the diagnosis and treatment of leishmaniasis. Here, we discuss the disease with special emphasis on its socioeconomic impact on the affected individual and community. In addition, we highlight the reasons why continued research aimed at developing an effective Leishmania vaccine is necessary.

Parasite-Derived Arginase Influences Secondary Anti-Leishmania Immunity by Regulating Programmed Cell Death-1–Mediated CD4+ T Cell Exhaustion

The breakdown of L-arginine to ornithine and urea by host arginase supports Leishmania proliferation in macrophages. Studies using arginase-null mutants show that Leishmania-derived arginase plays an important role in disease pathogenesis. We investigated the role of parasite-derived arginase in secondary (memory) anti-Leishmania immunity in the resistant C57BL/6 mice. We found that C57BL/6 mice infected with arginase-deficient (arg−) L. major failed to completely resolve their lesion and maintained chronic pathology after 16 wk, a time when the lesion induced by wild-type L. major is completely resolved. This chronic disease was associated with impaired Ag-specific proliferation and IFN-γ production, a concomitant increase in programmed cell death-1 (PD-1) expression on CD4+ T cells, and failure to induce protection against secondary L. major challenge. Treatment with anti–PD-1 mAb restored T cell proliferation and IFN-γ production in vitro and led to complete resolution of chronic lesion in arg− L. major–infected mice. These results show that infection with arg− L. major results in chronic disease due in part to PD-1–mediated clonal exhaustion of T cells, suggesting that parasite-derived arginase contributes to the overall quality of the host immune response and subsequent disease outcome in L. major–infected mice. They also indicate that persistent parasites alone do not regulate the quality of secondary anti-Leishmania immunity in mice and that the quality of the primary immune response may be playing a hitherto unrecognized dominant role in this process.

CD4+CD25+ regulatory T cells attenuate lipopolysaccharide-induced systemic inflammatory responses and promotes survival in murine Escherichia coli infection.

ABSTRACT It is well established that CD4+CD25+ regulatory T cells (Tregs) downregulate inflammatory immune responses and help to maintain immune homeostasis. Recent reports have shown that ligation of germline encoded pattern recognition receptors such as Toll-like receptors can stimulate Tregs and therefore implicate Tregs in the pathophysiology of sepsis and other inflammatory diseases. In this report, we show that injection of lipopolysaccharide (LPS) leads to expansion of CD4+CD25+FoxP3+ Tregs, suggesting that these cells may play an important role in immune regulation in LPS-induced acute inflammation. Indeed, genetic or immunological inhibition of Treg function using mice lacking functional Tregs (CD25 KO mice) or anti-CD25 monoclonal antibody (anti-CD25 mAb), respectively, led to acute death in an otherwise nonlethal LPS challenge. This was accompanied by exaggerated production of proinflammatory cytokines. Strikingly, adoptive transfer of CD4+CD25+ Tregs to CD25 KO mice before LPS challenge rescues mice from death. Unlike LPS, depletion of Tregs followed by concanavalin A (Con A) challenge does not result in mortality, suggesting that Treg depletion does not globally influence all models of acute inflammation. We authenticate our findings by showing that depletion of Tregs leads to mortality in a nonlethal Escherichia coli challenge accompanied by elevated serum levels of proinflammatory cytokines. Collectively, our results indicate that in addition to regulation of LPS-induced acute inflammation, Tregs help to improve bacterial clearance and promote survival in an acute model of bacterial infection.

Immunotherapy as a strategy for treatment of leishmaniasis: a review of the literature

Leishmaniasis occurs as a spectrum of clinical syndromes divided into cutaneous, mucocutaneous and visceral forms. The epidemiology and clinical features are highly variable owing to the interplay of many factors ranging from parasite species and strains, vectors, host genetics and environment. Currently, there is no effective licensed vaccine for use in humans against leishmaniasis. Most traditional and low-cost treatment options, particularly in poor and endemic areas, are toxic with many adverse reactions and they require a long course of administration. The use of more effective, less toxic drugs is limited because total treatment cost is very high (expensive) and there are fears of development of drug resistance. Recent studies indicate that certain strategies aimed at modulating the host immune response (collectively called immunotherapy) could result in prophylactic and/or therapeutic cure of leishmaniasis under both laboratory and field conditions. In this review, we focus on treatment of leishmaniasis with a particular emphasis on immunotherapy/immunochemotherapy as an alternative to conventional drug treatment.