No abstract

Cutaneous leishmaniasis affects millions of people around the world. Several species ofLeishmaniainfect mouse strains, and murine models closely reproduce the cutaneous lesions caused by the parasite in humans. Mouse models have enabled studies on the pathogenesis and effector mechanisms of host resistance to infection. Here, we review the role of nitric oxide (NO), reactive oxygen species (ROS), and peroxynitrite (ONOO−) in the control of parasites by macrophages, which are both the host cells and the effector cells. We also discuss the role of neutrophil-derived oxygen and nitrogen reactive species during infection withLeishmania. We emphasize the role of these cells in the outcome of leishmaniasis early after infection, before the adaptive Th-cell immune response.

The gene encoding the cytolytic protein perforin is selectively expressed by activated killer lymphocytes. To understand the mechanisms underlying the cell-type-specific expression of this gene, we have characterized the regulatory functions and the DNA-protein interactions of the 5'-flanking region of the mouse perforin gene (Pfp). A region extending from residues +62 through -141, which possesses the essential promoter activity, and regions further upstream, which are able to either enhance or suppress gene expression, were identified. The region between residues -411 and -566 was chosen for further characterization, since it contains an enhancer-like activity. We have identified a 32-mer sequence (residues -491 to -522) which appeared to be capable of enhancing gene expression in a killer cell-specific manner. Within this segment, a 9-mer motif (5'-ACAGGAAGT-3', residues -505 to -497; designated NF-P motif), which is highly homologous to the Ets proto-oncoprotein-binding site, was found to interact with two proteins, NF-P1 and NF-P2. NF-P2 appears to be induced by reagents known to up-regulate the perforin message level and is present exclusively in killer cells. Electrophoretic mobility shift assay and UV cross-linking experiments revealed that NF-P1 and NF-P2 may possess common DNA-binding subunits. However, the larger native molecular mass of NF-P1 suggests that NF-P1 contains an additional non-DNA-binding subunit(s). In view of the homology between the NF-P motif and other Ets proto-oncoprotein-binding sites, it is postulated that NF-P1 and NF-P2 belong to the Ets protein family. Results obtained from the binding competition assay, nevertheless, suggest that NF-P1 and NF-P2 are related to but distinct from Ets proteins, e.g., Ets-1, Ets-2, and NF-AT/Elf-1, known to be expressed in T cells.Perforin, the pore-forming protein (PFP), is a cytolytic protein specifically expressed in killer lymphocytes, including cytotoxic T lymphocytes (CTL), natural killer cells, lymphokine-activated killer (LAK) cells, and y/8 T cells. Together with other effector molecules, PFP is thought to play a crucial role in lymphocyte-mediated cytotoxicity by forming transmembrane pores that lead to target cell lysis (18,32,42,48 and their cognate trans-acting factors are involved in regulating the transcription of P4,, virtually no information is available concerning the regulatory functions and the DNAprotein interactions of the P4, 5'-flanking region.The tissue-and cell-specific manner of gene expression has been shown to rely primarily on the combined actions of various transcription factors. These transcription factors may interact with their cognate regulatory sequences (promoters, enhancers, or silencers) of the gene to be transcribed (27,29). The presence of multiple regulatory elements allows specific transcription factors to initiate efficient transcription in a coordinated manner within a cell at different times during its growth, differentiation, or activation. Mechanisms that control the expression of lymphocyte-specific...

Intracellular parasites of the genus Leishmania are the causative agents of leishmaniasis. The disease is transmitted by the bite of a sand fly vector, which inoculates the parasite into the skin of mammalian hosts, including humans. During chronic infection the parasite lives and replicates inside phagocytic cells, notably the macrophages. An interesting, but overlooked finding, is that other cell types and even non-phagocytic cells have been found to be infected by Leishmania spp. Nevertheless, the mechanisms by which Leishmania invades such cells had not been previously studied. Here, we show that L. amazonensis can induce their own entry into fibroblasts independently of actin cytoskeleton activity, and, thus, through a mechanism that is distinct from phagocytosis. Invasion involves subversion of host cell functions, such as Ca 2+ signaling and recruitment and exocytosis of host cell lysosomes involved in plasma membrane repair.This article has an associated First Person interview with the first author of the paper.

C57BL/6 and BALB/c mice are prototype hosts for the study of resistance and susceptibility to several infectious diseases. In many cases, resistance of C57BL/6 is due to the microbicidal effect of nitric oxide (NO) produced by macrophages in response to interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha), mainly secreted by Th1 cells and macrophages, respectively. BALB/c, usually unable to give rise to Th1 lymphocytes, does not control certain infections. However, we and others have previously observed that regardless of the adaptive immune response, C57BL/6 (M-1) macrophages are far more sensitive to the stimulus of IFN-gamma-plus lipopolysaccharide (LPS) for the production of NO than are BALB/c (M-2) cells, a feature that might also account for resistance. Here, we report that the differential production of NO by M-1 and M-2 macrophages correlates with the accumulation of inducible nitric oxide synthase (iNOS) mRNA and protein, which shows that expression of iNOS is differentially regulated in M-1 and M-2 cells. The higher accumulation of iNOS mRNA in M-1 cells is independent of its stability, and, thus, it is possible that transcription of the iNOS gene in these cells may be more efficient than in M-2 cells. A remarkable finding is that the level of iNOS protein is much higher in M-1 macrophages than in M-2 cells, as compared with the mRNA levels, which makes us speculate that differential translational or posttranslational controls of iNOS gene are operative.