Leishmania spp. are protozoans that survive and replicate intracellularly in mammalian macrophages. Antileishmanial immunity requires gamma interferon (IFN-␥)-mediated macrophage activation and generation of microbicidal effector molecules. The presence of intracellular Leishmania sp. impairs macrophage responses to IFN-␥, which has led to the description of macrophages as deactivated. It has recently become apparent that in addition to classical activation, macrophages can be activated by distinct triggers to express noninflammatory or anti-inflammatory genes. These nonclassical activation programs have been called alternative or type II pathways. We hypothesized that during initial contact with a phagocyte, leishmaniae activate one of these nonclassical pathways, resulting in expression of genes whose products suppress microbicidal responses. Using DNA microarrays, we studied gene expression in RNAs from BALB/c bone marrow macrophages with and without Leishmania chagasi infection. Some changes were verified by an RNase protection assay, reverse transcription-PCR, immunoblotting, or a bioassay. The pattern of genes activated by leishmania phagocytosis differed from the pattern of genes activated by bacteria or lipopolysaccharide and IFN-␥. Genes encoding some proinflammatory cytokines, receptors, and Th1-type immune response genes were downmodulated, and some genes associated with anti-inflammatory or Th2-like immune responses were up-regulated. Nonetheless, some markers of alternative (arginase) or type II activation (interleukin-10, tumor necrosis factor alpha) were unchanged. These data suggest that macrophages infected with L. chagasi exhibit a hybrid activation profile that is more characteristic of alternative or type II activation than of classical activation but does not strictly fall into either of these categories. We speculate that the pattern of genes upregulated by leishmania phagocytosis optimizes the chance of parasite survival in this hostile environment.Leishmania spp. are parasitic protozoans with a two life stages that correspond to the transit of the organisms between two hosts. The extracellular promastigote is found in the gut of the sand fly vector, and the obligate intracellular amastigote resides in the mammalian host macrophage. During a blood meal the sand fly inoculates the promastigote into a pool of blood in the skin, where it is phagocytosed by a resident mononuclear phagocyte. Inside the macrophage the parasite changes to an amastigote, which is the obligate intracellular form and the only form of the parasite found in a mammalian host. Intracellular parasites spread to other mononuclear

A periurban outbreak of visceral leishmaniasis (VL) caused by the protozoan Leishmania chagasi is ongoing outside Natal, northeast Brazil. Manifestations range from asymptomatic infection to disseminated visceral disease. Literature reports suggest that both genetic and environmental factors influence the outcome of infection. Due to the association of the tumor necrosis factor (TNF) locus with other infectious diseases, we examined whether polymorphic alleles at this locus are associated with the outcome of L. chagasi infection. Neighborhoods with ongoing transmission were identified through patients admitted to local hospitals. Altogether, 1,024 individuals from 183 families were classified with the following disease phenotypes: (i) symptomatic VL, (ii) asymptomatic infection (positive delayed-type hypersensitivity [DTH؉]), or (iii) no evidence of infection (DTH؊). Genotypes were determined at a microsatellite marker (MSM) upstream of the TNFB gene encoding TNF-␤ and at a restriction fragment length polymorphism (RFLP) at position ؊307 in the promoter of the TNFA gene encoding TNF-␣. Analyses showed that the distribution of TNFA RFLP alleles (TNF1 and TNF2) and the TNF MSM alleles (TNFa1 to TNFa15) differed between individuals with VL and those with DTH؉ phenotypes. TNF1 was transmitted more frequently than expected from heterozygous parents to DTH؉ offspring (P ‫؍‬ 0.0006), and haplotypes containing TNF2 were associated with symptomatic VL (P ‫؍‬ 0.0265, transmission disequilibrium test). Resting serum TNF-␣ levels were higher in TNF1/2 heterozygotes than in TNF1/1 homozygotes (P < 0.05). These data led us to hypothesize that an individual's genotype at the TNF locus may be associated with whether he or she develops asymptomatic or symptomatic disease after L. chagasi infection. The results preliminarily suggest that this may be the case, and follow-up with larger populations is needed for verification.The Leishmania spp. are obligate intracellular protozoa that cause a spectrum of diseases, including cutaneous, mucocutaneous, and visceral leishmaniasis (VL), in tropical or subtropical countries. In Latin America, the form of leishmaniasis leading to most fatalities, VL, is caused by Leishmania chagasi (44). The outcome of L. chagasi infection ranges in severity from asymptomatic infection to a severe progressive wasting disease called VL, which has 10% mortality even with medical treatment (5,33,34,44). The factors that determine whether an individual will develop asymptomatic or symptomatic disease after L. chagasi infection are not defined.Malnutrition, young age, and gender in those over age 10 predispose individuals to develop VL (21, 31). In addition, studies of inbred mice have led investigators to hypothesize that genetic factors may also contribute to the outcome of human disease (10,12). Supporting this hypothesis, several groups of researchers have independently documented familial aggregation of either symptomatic VL or positive delayed-type hypersensitivity (DTH) to leishmania antigen, a measure ...

Leishmania spp. are intracellular protozoa residing in mononuclear phagocytes. Leishmania organisms are susceptible to microbicidal responses generated in response to phagocytosis. Assuming that both phagocyte and parasite populations are heterogeneous, it is advantageous to examine the response of individual cells phagocytosing living parasites. Because Leishmania spp. lose virulence during the raising of transfectants, we developed a method to label live Leishmania chagasi short-term with fluorescent dyes. Up to six parasite divisions were detected by flow cytometry after labeling with carboxyfluorescein diacetate succinimidyl ester (CFSE), dioctadecyl-tetramethylindo carbocyanine perchlorate, or chloromethyl tetramethylrhodamine. Labeled parasites entered mononuclear phagocytes as determined by confocal and time-lapse microscopy. Dihydroethidium (DHE) was used to detect macrophage-derived oxidants generated during phagocytosis. Presumably Leishmania organisms are opsonized with host serum/tissue components such as complement prior to phagocytosis. Therefore, we investigated the effects of opsonization and found that this increased the efficiency of CFSE-labeled parasite entry into monocytes (84.6% ؎ 8.8% versus 20.2% ؎ 3.8% monocytes infected; P < 0.001). Opsonization also increased the percentage of phagocytes undergoing a respiratory burst (66.0% ؎ 6.3% versus 41.0% ؎ 8.3% of monocytes containing CFSE-labeled parasites; P < 0.001) and the magnitude of oxidant generation by each infected monocyte. Inhibitor data indicated that DHE was oxidized by products of the NADPH oxidase. These data suggest that opsonized serum components such as complement lead to more efficient entry of Leishmania into their target cells but at the same time activate the phagocyte oxidase to generate microbicidal products in infected cells. The parasite must balance these positive and negative survival effects in order to initiate a viable infection.Leishmania protozoa are intracellular parasites residing in macrophages or other mononuclear cells of their mammalian hosts (13). The parasites are inoculated into mammalian cutaneous tissue in the form of a promastigote, which must locate an appropriate phagocyte and stimulate its own internalization in order to survive. However, promastigotes are susceptible to killing by the reactive oxygen species generated by the macrophage during phagocytosis (19,20,27). Once intracellular, parasites convert to the more resistant obligate intracellular amastigote form. The Leishmania spp. have developed a number of mechanisms to protect themselves from oxidant-mediated killing while in their susceptible promastigote form. These include a surface glycolipid coat called lipophosphoglycan, antioxidant defense molecules such as iron superoxide dismutase, peroxidoxins, and a unique thiol-reducing factor, trypanothione (16,33,34). Furthermore, Leishmania spp. have the ability to suppress gamma interferon (IFN-␥)-mediated activation of macrophages through enhancing macrophage phosphatase activity (9, 30).Th...

The effects of short-term diet change from high fat (F) to high carbohydrate (C) (or vice versa) on the storage and utilization of glycogen and triacylglycerol (TG) in muscle and liver were studied in untrained rats. Rats were fed on an F or C diet for 28 days. For an additional 3 days, half of the rats in both F and C groups were fed the same diets as before (F-F and C-C) and the other half of the rats were switched to the counterpart diets (F-C and C-F). On the final day of the experiment, half of the rats in each diet group were exercised by swimming for 1.5 h and the other half were rested. Short-term diet change from F to C diets increased, but the change from C to F diets decreased, glycogen stores of soleus and plantaris muscles and liver, resulting in no difference in glycogen stores between F-C and C-C, and between F-F and C-F. The dietary change also had an affect on TG stores of red gastrocnemius muscle and liver-however, muscle TG stores were still higher in F-C than in C-C and C-F, and there were no differences in liver TG stores between F-C and C-F. Exercise decreased muscle glycogen contents markedly in F-C and C-C, whereas, it decreased muscle TG concentrations in F-F and C-F. Liver glycogen depletion was lower in F-C than in other groups. Lipolytic activities of epididymal adipose tissue at rest and postexercise were no differences between F-F and F-C, and were higher in F-C than in C-C and C-F. beta-adrenergic receptor binding was determined with [125I] iodocyanopindolol, and maximal numbers of beta-adrenergic receptor of plasma membrane from perirenal adipose tissue were approximately 170%-200% higher in F-C than in other groups at rest and postexercise. These results suggested that short-term C diet fed rats adapted to F diet enhanced not only glycogen stores of muscle and liver but also did not decrease lipolytic activity of adipose tissue with increased beta-adrenergic receptor density, resulting in the preservation of energy reserves (glycogen and TG) of muscle at rest, and liver glycogen sparing during exercise.

The general pharmacological properties of YJA20379-8 (3-butyryl-4-[(R)-1-methylbenzylamino]-8-ethoxy-1,7-naphthyridine, CAS 187654-40-6), a new H+/K(+)-ATPase inhibitor with anti-ulcer activities, were investigated in mice, rats and guinea pigs. YJA20379-8 at oral doses of 25, 50 and 100 mg/kg did not affect the locomotor activity, hexobarbital hypnosis and motor coordination in mice. The drug did not have analgesic action and anticonvulsant action at the doses of 100 mg/kg p.o. The respiration and blood pressure were not affected at 10 mg/kg i.v. in rats. YJA20379-8 at 2 x 10(-4) g/ml did neither produce any contraction nor relaxation of isolated organs, such as guinea pig ileum, rat fundus, rat uterus and guinea pig vas deferens, and the drug antagonized the contractile responses to several spasmogens, such as acetylcholine, histamine, serotonin, L-phenylephrine, oxytocin and BaCl2. The drug up to 100 mg/kg p.o. did not affect pupil size and the intestinal propulsion of mice. And it did not show an anticarrageenan action at 100 mg/kg. In this general pharmacology study, hypothermic effect in mice, retardation in gastric emptying in rats, decreases in urine excretion in rats at oral doses of 50 and 100 mg/kg of YJA20379-8 and the spasmolytic activity could be found. However, no other effects were exhibited at a high oral dose of 100 mg/kg in animals in this study.