Differences in affinity of binding of lymphocytic choriomeningitis virus strains to the cellular receptor alpha-dystroglycan correlate with viral tropism and disease kinetics.

alpha-Dystroglycan (alpha-DG) was recently identified as a receptor for lymphocytic choriomeningitis virus (LCMV) and several other arenaviruses, including Lassa fever virus (W. Cao, M. D. Henry, P. Borrow, H. Yamada, J. H. Elder, E. V. Ravkov, S. T. Nichol, R. W. Compans, K. P. Campbell, and M. B. A. Oldstone, Science 282:2079-2081, 1998). Data presented in this paper indicate that the affinity of binding of LCMV to alpha-DG determines viral tropism and the outcome of infection in mice. To characterize this relationship, we evaluated the interaction between alpha-DG and several LCMV strains, variants, and reassortants. These viruses could be divided into two groups with respect to affinity of binding to alpha-DG, dependence on this protein for cell entry, viral tropism, and disease course. Viruses that exhibited high-affinity binding to alpha-DG displayed a marked dependence on alpha-DG for cell entry and were blocked from infecting mouse 3T6 fibroblasts by 1 to 4 nM soluble alpha-DG. In addition, high-affinity binding to alpha-DG correlated with an ability to infiltrate the white pulp (T-dependent) area of the spleen, cause ablation of the cytotoxic T-lymphocyte (CTL) response by day 7 postinfection, and establish a persistent infection. In contrast, viruses with a lower affinity of binding to alpha-DG were only partially inhibited from infecting alpha-DG(-/-) embryonic stem cells and required a concentration of soluble alpha-DG higher than 100 nM to prevent infection of mouse 3T6 fibroblasts. These viruses that bound at low affinity were mainly restricted to the splenic red pulp, and the host generated an effective CTL response that rapidly cleared the infection. Reassortants of viruses that bound to alpha-DG at high and low affinities were used to map genes responsible for the differences described to the S RNA, containing the virus attachment protein glycoprotein 1.

B cell-deficient mice are highly resistant to Leishmania donovani infection, but develop neutrophil-mediated tissue pathology.

Resolution of Leishmania infection is T cell-dependent, and B lymphocytes have been considered to play a minimal role in host defense. In this study, the contribution of B lymphocytes to the response against Leishmania donovani was investigated using genetically modified IgM transmembrane domain (muMT) mutant mice, which lack mature B lymphocytes. When compared with wild-type mice, muMT mice cleared parasites more rapidly from the liver, and infection failed to establish in the spleen. The rapid clearance of parasites in muMT mice was associated with accelerated and more extensive hepatic granuloma formation compared with wild-type mice. However, the liver of infected muMT mice also showed signs of destructive pathology, associated with the presence of increased numbers of neutrophils. The role of neutrophils in controlling parasite growth in the viscera was determined by depletion with the mAb RB6-8C5. This treatment led to a dramatic enhancement of parasite growth in both the liver and spleen of muMT and wild-type mice. As assessed by transfer of both normal and chronic-infection serum, Ig protects microMT mice from destructive hepatic pathology, but minimally alters their resistance compared with wild-type mice. However, adoptive transfer of CD4+ and CD8+ T cells into recombinase activating gene 1 (RAG1-/-) recipients, suggested that T cell function was not altered by maturation in a B cell-deficient environment. Taken together, these data suggest an inhibitory role for B lymphocytes in resistance to L. donovani unrelated to the presence or absence of Ig. However, Ig protects muMT mice from the exaggerated pathology that occurs during infection.

Neutralization of IL-12 demonstrates the existence of discrete organ-specific phases in the control of Leishmania donovani.

IL-12 plays a key role in stimulating both innate and antigen-specific immune responses against a number of intracellular pathogens. A neutralizing anti-IL-12 monoclonal antibody (mAb) was used to define and compare the role of endogenous IL-12 in the liver and spleen of mice infected with Leishmania donovani. IL-12 neutralization both early and late in infection caused delayed resolution of parasite load, a transient decrease in IFN-gamma, IL-4, TNF-alpha and inducible nitric oxide synthase (NOS-2) production, and suppressed tissue granuloma formation in the liver of genetically susceptible BALB/c mice. In contrast to the liver of BALB/c mice, neutralization of IL-12 had no effect on parasite burden in the spleen over the first 28 days of infection. However, IL-12 appeared to be critical for the development of mechanisms which subsequently contain the growth of persistent parasites in this organ in that neutralization of IL-12 dramatically enhanced parasite growth after day 28 of infection. Following IL-12 neutralization, the later unchecked growth of parasites in the spleen was coincident with an extensive breakdown of the tissue microarchitecture. Immunohistochemical studies revealed that IL-12 was largely produced by uninfected cells in L. donovani-infected BALB/c mice. In contrast, the course of infection in the liver and spleen of genetically resistant CBA/n mice was unaffected by the administration of anti-IL-12 mAb. These results suggest that the liver and spleen in susceptible BALB/c mice have different temporal requirements for IL-12 in controlling L. donovani infection, whereas IL-12 plays little role in either organ in resistant CBA/n mice. In addition, IL-12 appears to be involved in the generation of both Th1 and Th2 responses during L. donovani infection in BALB/c mice.

Destruction of follicular dendritic cells during chronic visceral leishmaniasis.

Follicular dendritic cells (FDCs) play a pivotal role in the germinal center (GC) response and in the development and regulation of B lymphocytes. Pathologic changes in GCs and a loss of FDCs have previously been noted in various viral infections, notably HIV-1. However, such changes have not been formally described in a chronic parasitic infection. In BALB/c mice infected with Leishmania donovani, parasites persist in the spleen for long periods, with associated splenomegaly. To examine the fate of FDC during the course of this chronic infection, we used 1) immunohistology, with FDC-specific mAbs; and 2) passive immunization with immune complexes, followed by light and electron microscopy. This study demonstrates that destruction of FDCs and a concomitant loss of GCs are associated with chronic visceral leishmaniasis. These pathologic effects are notable from 4 wk postinfection. At 8 wk postinfection and beyond, FDC are almost undetectable by both immunohistology and functional immune complex trapping. The loss of FDCs is associated with the infiltration of heavily parasitized macrophages into the GC, and reduction in parasite burden by chemotherapy is able to retard the process of FDC destruction. These data directly demonstrate for the first time the loss of FDCs during a chronic parasite infection and suggest a mechanism underlying the aberrant regulation of B cell function in murine visceral leishmaniasis.

An in vivo analysis of cytokine production during Leishmania donovani infection in scid mice.

Leishmania donovani, the causative agent of visceral leishmaniasis, fails to induce NK cell activation in scid mice. In order to further our understanding of the host response to L. donovani, we analysed cytokine mRNA accumulation and TNF alpha protein synthesis in the liver of scid and BALB/c mice infected with this parasite. scid mice infected with L. donovani exhibited very little proinflammatory response, as measured by cytokine mRNA accumulation and TNF alpha protein expression, supporting the notion of a relatively "quiet" interaction between L. donovani and macrophages in these animals. In contrast, immunocompetent BALB/c mice were found to generate an early IFN gamma response, coincident with a rise in IL-2 mRNA levels and elaboration of a tissue response involving TNF alpha production by infected Kupffer cells. These results extend our understanding of the response of BALB/c and scid mice to L. donovani infection and highlight the value of cytokine analysis at both the tissue and cellular levels.