Malnutrition Alters the Innate Immune Response and Increases Early Visceralization following Leishmania donovani Infection

Globally, protein-energy malnutrition is the most frequent cause of immunodeficiency (58). Epidemiologic and experimental studies have documented an increased risk for visceral leishmaniasis, caused by intracellular protozoan parasites of the Leishmania donovani complex, in the malnourished host (1, 2, 26). However, the immunologic basis for this association has not been established and standardized experimental models have not addressed this important issue. In this study, our goal was to investigate the mechanisms of the malnutrition-related susceptibility to visceral leishmaniasis. There were three components of the study. First, we needed to create a murine model of malnutrition that was relevant to human malnutrition in developing countries. Although the mouse has been extensively used in animal models of malnutrition, there is no standard murine model of protein-energy malnutrition (69). Human malnutrition is complex, typically involving deficiency of protein and energy with superimposed deficits of other nutrients. Zinc deficiency usually accompanies protein-energy malnutrition (19). Iron deficiency is highly prevalent in developing countries and may accompany zinc deficiency due to a common risk factor, cereal-based diets with little meat (61). Thus, in this model, in addition to protein and energy, zinc and iron were selected as deficient nutrients. Much of the vast body of data that has been collected on human malnutrition is based on anthropometric measures, i.e., weight-for-age (WA), height-for-age, and weight-for-height (13). However, in previous mouse models of malnutrition, there have been no efforts to relate morphometric measures of nutritional status to either human anthropometric scales or immunocompetence. Weight-for-age determination is advantageous because it can be measured unambiguously, provides a synthesis of linear growth and body proportion (13), and correlates with probability of death in children in developing countries (36). In this study, murine WA was correlated with specific measures of host defense and risk for visceral L. donovani infection. A second component of the study was to examine possible defects in the mediator network of the innate immune system produced by malnutrition, because it is the early events that are likely to determine whether the inoculated parasites are controlled locally or disseminate to visceral organs. The innate immune system provides a first line of defense against pathogens and instructs the differentiation of Th0 cells into Th1 and Th2 cells (18). It has been estimated that the innate immune system provides protection against 98% of the pathogens that are encountered (34). Malnutrition has been associated with an increased risk of many infections (3); however, there are no animal models that specifically examine the effect of malnutrition on the innate immune response to infection. The third part of the study was to investigate the mechanism of visceralization (the process whereby parasites disseminate from the site of cutaneous inoculation and the draining lymph nodes to the liver, spleen, and bone marrow). To reach this goal, we needed to develop a more natural animal model of visceral leishmaniasis, using the vector stage of the parasite (promastigote) and the intradermal route of infection. Although the immunopathogenesis of murine visceral leishmaniasis has been investigated, previous studies have used an unnatural (intravenous) route of infection and/or the mammalian host stage of the parasite (amastigote) as the infecting inoculum (35, 40, 43, 48, 65). The intravenous route of inoculation negates the immune processing that would occur in the skin and in the draining lymph node and therefore does not accurately reflect the immune response that would occur in a natural cutaneous infection. Furthermore, models of visceral leishmaniasis that utilize the intravenous route of inoculation do not facilitate an understanding of the mechanism of visceralization. Selection of either amastigote or promastigote as the infective inoculum may also have an important bearing on the course of infection, because these two stages of the parasite possess disparate virulence factors (9) and produce different immune responses (20). In this study, we establish a murine model of polynutrient deficiency that is similar to human malnutrition in developing regions of the world. We demonstrate that the malnourished mouse has an altered innate immune defense and is at increased risk of visceralization following cutaneous L. donovani infection.