Biomedical subjects

N E Reiner

Publications and source records attributed to N E Reiner.

At least 37 records · Page 2Linked to original sources

PubMed1994

Altered cell signaling and mononuclear phagocyte deactivation during intracellular infection.

Given the critical antimicrobial properties of mononuclear phagocytes, an important concern in cell biology and immunology has been to understand how intracellular microbes are able to establish states of chronic infection within these cells. Recent studies indicate that mononuclear phagocytes become functionally deactivated during intracellular infection. Here, Neil Reiner considers the experimental evidence to indicate that this is a frequent event that may be accounted for by induced defects in the signaling pathways required to bring cells to an activated state.

Animals↗

PubMed1994

Gamma interferon induces rapid and coordinate activation of mitogen-activated protein kinase (extracellular signal-regulated kinase) and calcium-independent protein kinase C in human monocytes.

Gamma interferon plays an important role in regulating the functional properties of mononuclear phagocytes. In the present study, the role of activated protein kinases in the mechanism of action of gamma interferon cell signaling in human peripheral blood monocytes was investigated. Analysis in vitro of 100,000 x g cytosolic fractions from untreated and interferon-treated cells showed that agonist treatment resulted in time- and concentration-dependent increases in phosphotransferase activity when myelin basic protein (MBP) was used as the substrate. Anion-exchange chromatography of high-speed supernatants prepared from detergent extracts of interferon-treated cells revealed two discrete peaks of MBP phosphotransferase activity. Immunoblotting of fractions from these peaks with antiphosphotyrosine antibodies and with antibodies that specifically recognize the family of mitogen-activated protein (MAP) kinases detected a MAP kinase with a subunit M(r) of 42,000 in the earliest-eluting peak (peak 1). Phosphorylation of the 42,000-M(r) protein on tyrosine was observed only after treatment of cells with interferon. The contribution of MAP kinase to the interferon-stimulated activity in peak 1 was confirmed by quantitative immunoprecipitation with anti-MAP kinase and antiphosphotyrosine antibodies. The conclusion that the interferon-activated MBP kinase in peak 1 could be accounted for by an activated MAP kinase was also supported by the finding that fractions from Mono Q peak 1 demonstrated activity towards a MAP kinase-specific substrate. The later-eluting peak of interferon-activated MBP phosphotransferase activity appeared to be accounted for by an activated protein kinase C (PKC). This conclusion is based upon analyses of immunoblotting and immunoprecipitation experiments with antibodies to PKC and was also supported by the observed inhibition of this kinase with a PKC pseudosubstrate peptide. The interferon-stimulated PKC present in Mono Q peak 2 was active in the absence of calcium ions, suggesting that it is a calcium-independent isoform of PKC.

Amino Acid Sequence↗

PubMed1993

Expression of 65- and 67-kilodalton heat-regulated proteins and a 70-kilodalton heat shock cognate protein of Leishmania donovani in macrophages.

Heat shock protein (HSP) expression was examined in murine bone marrow-derived macrophages infected with stationary-phase promastigotes of Leishmania donovani. Immunoblotting performed with a rabbit polyclonal antiserum raised against HSP60 from Heliothis virescens (moth) revealed the de novo appearance of 65- and 67-kDa proteins in leishmania-infected macrophages. A third protein of 60 kDa, which represented murine HSP60, was also detected, and its expression did not change in response to infection. In contrast, expression of the novel 65- and 67-kDa proteins in infected cells was coordinately regulated and, at 24 h of infection, reached maximal levels of 52 to 100% increases above initial levels determined at 3 h. Proteins which had identical electrophoretic mobilities and were similarly regulated in response to heat were also detected in promastigotes. The appearance of these proteins in macrophages was specific to leishmania infection in that neither protein was detected in noninfected cells either in the basal state or following several treatments, including (i) infection with Yersinia pseudotuberculosis, (ii) phagocytosis of Staphylococcus aureus, (iii) NaAsO2 treatment, and (iv) heat shock. Expression of the 65- and 67-kDa heat-regulated Leishmania proteins was also observed to be selective, in that as their concentration was increasing, the abundance of the Leishmania surface protease gp63 in infected cells was noted to decrease. Murine HSP60 but not the Leishmania heat-regulated proteins was also recognized by a distinct rabbit antiserum raised against human HSP60, suggesting the presence of specific determinants within these Leishmania proteins. A monoclonal antibody that recognizes both mammalian HSP70 and HSP70 from plasmodia detected single isoforms of both Leishmania and murine HSP70 in infected cells, and the level of neither protein changed during infection. Moreover, although a murine HSP of 73 kDa was induced in response to both heat shock and NaAsO2 treatment, it was not induced to detectable levels by infection. The rapid and relatively high level of expression of inducible HSP60-related proteins of L. donovani and Leishmania HSP70 in infected macrophages suggests that these proteins are involved in pathogenesis and may be important targets of the immune response.

Animals↗

PubMed1992

Defective stimulus-response coupling in human monocytes infected with Leishmania donovani is associated with altered activation and translocation of protein kinase C.

Stimulus-response coupling through protein kinase C (PKC) was shown to be defective in mononuclear phagocytes (M phi) infected with Leishmania donovani. Phorbol 12-myristate 13-acetate (PMA)-induced oxidative burst activity and protein phosphorylation were markedly attenuated in infected M phi. These results were not explained either by quantitative alterations in amounts of PKC or by altered phorbol ester binding but were related to defects in kinase activation. Analysis in vitro of the kinetic properties of PKC from infected M phi revealed an approximately 2-fold increase in the concentration of 1,2-dioleoyl-rac-glycerol required to achieve half-maximal kinase activation. Evidence for abnormal PKC activation in vivo was reflected by attenuation of PMA-induced translocation of enzyme to the particulate fraction of infected cells. These results provide direct evidence that infection with Leishmania inhibits activation of, and therefore intracellular signaling dependent on, PKC. Inhibition of stimulus-response coupling through PKC provides a basis for understanding impairment of cellular activation by Leishmania and may contribute to chronic infection.

Animals↗

PubMed1992

Stimulus-response coupling in monocytes infected with Leishmania. Attenuation of calcium transients is related to defective agonist-induced accumulation of inositol phosphates.

Mononuclear phagocytes infected with Leishmania have been shown to have defective responses to extracellular stimuli. To investigate the potential relationship of these findings to alterations in calcium-dependent signaling pathways, the regulation of [Ca2+]i concentrations was examined in human peripheral blood monocytes infected with amastigotes of Leishmania donovani. Measurements of [Ca2+]i in fura-2-loaded monocytes were made at the single cell level by microfluorimetry. In normal monocytes, resting [Ca2+]i was 56 +/- 2 nM (mean +/- SEM). In contrast, in monocytes infected with Leishmania there was an approximately twofold increase in basal [Ca2+]i (122 +/- 5 nM, p less than 0.01 vs control). Treatment of cells with pertussis toxin before infection did not abrogate infection-induced increases in basal [Ca2+]i, suggesting that this effect was not mediated via the activation of a G protein coupled to phospholipase C. However, elevated resting [Ca2+]i did correlate with increased rates of 45Ca2+ uptake by infected monocytes. As expected, in response to treatment with 10(-7) M FMLP, control monocytes showed rapid net increases in [Ca2+]i of 303 +/- 19 nM. In contrast, net transients of [Ca2+]i in infected monocytes in response to FMLP were attenuated to only 137 +/- 9 nM (p less than 0.01 vs control). This result was not related to excess buffering of [Ca2+]i in infected cells as both control and infected monocytes showed equivalent transients of [Ca2+]i in response to the calcium ionophore A23187. Rather, inhibition of agonist-induced calcium release in infected cells appeared related to defective generation of second messenger because compared to control cells labeled with myo-[2-3H]inositol, little accumulation of inositol 1,4,5-trisphosphate was detected in infected monocytes. Attenuation of inositol phosphate accumulation and calcium release in response to chemotactic peptide correlated with decreased FMLP-induced superoxide and hydrogen peroxide production by infected monocytes. These results provide direct evidence for defective regulation of [Ca2+]i and calcium-dependent signaling in Leishmania-infected monocytes and provide a basis for understanding abnormalities in activation-related responses that involve signaling through Ca(2+)-regulated pathways.

Animals↗

PubMed1992

Inhibition of expression of major histocompatibility complex class II molecules in macrophages infected with Leishmania donovani occurs at the level of gene transcription via a cyclic AMP-independent mechanism.

Among the important pleiotropic responses to gamma interferon (IFN-gamma) during the activation of macrophages (M phi) is the increased expression of major histocompatibility complex class II genes. In the present study, infection with Leishmania donovani was shown to inhibit in parallel the induction by IFN-gamma of H-2 A beta gene transcription, class II mRNA accumulation, and H-2 Ad protein expression in cells of the murine macrophage cell line P388D1. Treatment of P388D1 cells with either the adenylate cyclase activator cholera toxin or the protein kinase A activator N6-2'-O-dibutyryl cyclic AMP (dibutyryl cAMP) similarly inhibited the induction by IFN-gamma of class II protein expression, and in parallel with Leishmania infection, cholera toxin inhibited the induction of mRNA for the H-2 A alpha and H-2 A beta proteins. Concentrations of intracellular cAMP were significantly increased in cholera toxin-treated cells but not in leishmania-infected cells. These findings indicate that at least one mechanism by which Leishmania infection attenuates the activation of M phi by IFN-gamma involves selective, transcriptional inhibition of major histocompatibility complex class II genes via a cAMP-independent mechanism.

Animals↗

PubMed1991

Modification of GP63 genes from diverse species of Leishmania for expression of recombinant protein at high levels in Escherichia coli.

Toward the future development of a defined subunit vaccine against leishmaniasis is, high levels of recombinant GP63 for diverse species of Leishmania were produced in Escherichia coli. Several features of Leishmania GP63 genes were simultaneously modified with the polymerase chain reaction (PCR) using either cloned genes or total genomic DNA from Leishmania as template DNA for the PCR amplification reactions. The PCR products included only the coding region for the predicted mature form of GP63 that occurs on the surface of Leishmania, flanked by the appropriate translation signals and cloning sites for the production of recombinant GP63 as nonfusion protein in E. coli. When the codon usage in the GP63 gene was modified to reduce the guanine and cytosine content for the codons adjacent to the ATG initiation codon, rGP63 represented about 50% of total protein in E. coli. Mouse monoclonal antibodies raised against purified Leishmania major rGP63 had equivalent immunoblotting characteristics for native GP63 and recombinant GP63 with respect to linear determinants on GP63 expressed in diverse species of Leishmania. Human T cell lines and clones were derived from a patient infected with Leishmania braziliensis panamensis using rGP63 purified from an L. major GP63 expression clone as antigen.

Animals↗

PubMed1990

Modulation of in vitro monocyte cytokine responses to Leishmania donovani. Interferon-gamma prevents parasite-induced inhibition of interleukin 1 production and primes monocytes to respond to Leishmania by producing both tumor necrosis factor-alpha and interleukin 1.

Cytokines produced by mononuclear cells are important regulatory and effector molecules and evidence has been presented to support a role at least for tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) in host defense against Leishmania. In the present study, we examined the production of TNF-alpha and interleukin 1 (IL-1) by resting and IFN-gamma-primed peripheral blood monocytes infected in vitro with Leishmania donovani. Monocytes produced neither IL-1 nor TNF-alpha during challenge with Leishmania. Cells preinfected with Leishmania synthesized normal amounts of TNF-alpha, but had diminished production of IL-1 in response to stimulation with either S. aureus or lipopolysaccharide (LPS). The induction by S. aureus or LPS of IL-1 beta mRNA accumulation in infected cells was normal despite diminished intracellular or supernatant IL-1 protein and bioactivity. Thus, inhibition of IL-1 production by Leishmania most probably reflected diminished translation of IL-1 beta mRNA. Pretreatment of cells with IFN-gamma abrogated infection-induced inhibition of IL-1 production and primed cells for the production of both IL-1 and TNF-alpha upon subsequent exposure to Leishmania. These results indicate that L. donovani has evolved the capacity to infect mononuclear phagocytes, without stimulating the production of two potentially host-protective monokines. The ability of IFN-gamma to prime monocytes to produce TNF-alpha and IL-1 in response to infection with Leishmania and to prevent inhibition of IL-1 production may have implications for immunotherapy with this lymphokine.

Animals↗

PubMed1989

Genetic heterogeneity in Peruvian Leishmania isolates.

Leishmania were isolated from Peruvian patients with uta or espundia; genomic DNA was examined for restriction fragment length differences by Southern blot analysis using DNA probes for beta-tubulin and for the major surface antigen gp63. Using 5 different restriction endonucleases, Peruvian isolates show homogeneity when examined at the beta-tubulin locus. In contrast, the organisms demonstrated heterogeneity both within and between disease groups when examined for restriction site differences within the gp63 locus. The differences observed did not correlate with the 2 disease groups. Comparison of these Peruvian isolates to New World reference strains of the Leishmania braziliensis complex reveals no consistent pattern of identity with either L. b. guyanensis, L. b. panamensis, or L. b. braziliensis.

Animals↗

PubMed1988

Kinetics of gamma interferon binding and induction of major histocompatibility complex class II mRNA in Leishmania-infected macrophages.

Cells of the monocyte-macrophage series must carry out discrete accessory-cell functions during the process of antigen-specific T-cell activation. One of these functions is the cell-surface expression of major histocompatibility complex (MHC) class II gene products, which are involved in the presentation of foreign antigen to T cells. Previously, we reported that murine peritoneal macrophages infected with the obligate intracellular protozoan Leishmania donovani had suppressed responses to gamma interferon (IFN-gamma) for the induction of MHC class II antigen expression. To determine the molecular basis for this suppression, we examined in the present series of experiments the interaction of this organism with cells of the murine macrophage tumor cell line P388D1. When infected with Leishmania, these cells were also markedly unresponsive to IFN-gamma for the induction of MHC class II antigen expression. This finding was not the result of a defect at the level of the IFN-gamma receptor. Thus, when 125I-labeled IFN-gamma was used, infected macrophages were found to express normal numbers of high-affinity IFN-gamma receptors, and ligand-receptor binding resulted in rapid internalization of labeled IFN-gamma. Despite normal ligand-receptor interactions, the induction in infected cells of mRNA encoding MHC (H-2) class II I-A alpha and beta chains in response to IFN-gamma was markedly suppressed. However, infected cells had normal levels of mRNA encoding the cytoskeletal protein actin. These findings indicate that Leishmania interferes with IFN-gamma induction of macrophage MHC class II antigen expression by down-regulating lymphokine induction of MHC class II mRNA. Suppression of class II expression by this intracellular parasite may prevent subsequent T-cell recognition of infected macrophages and thus favor parasite survival.

Animals↗

PubMed1988

Eicosanoid metabolism by Leishmania donovani-infected macrophages: mouse strain responses in prostanoid synthesis.

Arachidonic acid (20:4) conversion to prostanoids was examined in murine peritoneal macrophages infected in vitro with Leishmania donovani. Four strains of mice differing in resistance to in vivo L. donovani infection were studied. Normal macrophages from all strains converted 20:4 to prostanoids and this was augmented by L. donovani infection. Although cells from each strain synthesized elevated levels of prostaglandin-E2 (PGE2), there were differences with respect to relative increases of this product, with infected macrophages from the C3H/HeJ strain showing the smallest increase above basal levels. These results indicate that macrophages from mouse strains with distinct levels of in vivo resistance to L. donovani all respond to infection with augmented prostanoid synthesis. Although some heterogeneity in strain-specific PGE2 responses to in vitro infection were observed (with respect to increases in PGE2 synthesis above basal levels), it is unlikely that differential resistance of these strains to in vivo infection is strictly related to these relative differences. It seems likely that other genetically controlled factors may have a major impact on disease expression.

Alprostadil↗

PubMed1987

Parasite-accessory cell interactions in murine leishmaniasis. II. Leishmania donovani suppresses macrophage expression of class I and class II major histocompatibility complex gene products.

In the present study, we examined the modulation of MHC class II and class I gene products on BALB/c macrophages infected with the obligate intracellular protozoan Leishmania donovani. Our findings indicated that this organism suppressed macrophage expression of both classes of MHC antigens. These effects varied somewhat, depending on whether cells were in the basal state or were stimulated with interferon-gamma. Thus, class II density on interferon-gamma-treated infected macrophages was suppressed by as much as 90%, relative to lymphokine-stimulated control cells. Induction of H-2K and H-2D by lymphokine treatment of infected macrophages was also markedly reduced. In the basal (non-lymphokine-treated) state, infected cells also showed reduced expression of H-2K and H-2D, but not I-A or I-E. The latter result was related to minimal levels of class II molecules on normal, in vitro cultured macrophages. Suppression of MHC gene products correlated with both the duration and intensity of leishmania infection and could not be overcome by increasing doses of interferon-gamma. Culture of cells under conditions of cyclooxygenase inhibition completely abolished elevated synthesis of prostaglandin E2 by infected macrophages and augmented their responsiveness to lymphokine induction of class II antigens by 60 to 80%. These results indicate that L. donovani is capable of subverting a critical macrophage accessory function required for the induction of T lymphocyte immunity. This mechanism could account, at least in part, for defective parasite-specific cell-mediated immunity seen during infections with this protozoan.

Animals↗

PubMed1987

Parasite accessory cell interactions in murine leishmaniasis. I. Evasion and stimulus-dependent suppression of the macrophage interleukin 1 response by Leishmania donovani.

Interleukin 1 (IL 1) is a principal mediator of the host immune response to microbial challenge. Accessory cells of the monocyte-macrophage series are a major source of this cytokine and are also chronically parasitized by protozoa of the genus Leishmania. This suggests that characterization of the macrophage IL 1 response to Leishmania would increase our understanding of the regulation of host immunity to these organisms. In the present study, the macrophage IL 1 response to Leishmania donovani was examined because infections with this organism have findings consistent with parasite-specific T cell unresponsiveness. Cytokine activity was measured either by direct stimulation or by co-stimulation of thymocytes. Conditioned media from BALB/c resident peritoneal macrophages infected with amastigotes of L. donovani contained no more IL 1 than did supernatant fluids of control cells. In contrast, supernatants from cells stimulated with lipopolysaccharide or heat-killed Listeria monocytogenes had significantly increased cytokine content. Resident cells infected with L. donovani for 4 hr before being stimulated with Listeria demonstrated a suppressed IL 1 response (approximately 40% of Listeria alone) to this secondary particulate stimulus. In contrast, the secondary response of leishmania-preinfected cells to lipopolysaccharide was not affected. To examine whether accessory cell nonresponsiveness to L. donovani (with respect to IL 1) was related to the state of macrophage activation, elicited peritoneal macrophages obtained by injection of proteose peptone were also studied. These cells responded to stimulation with lipopolysaccharide and fixed Staphylococcus aureus with increases in intracellular, membrane, and secreted cytokine activities. In contrast, L. donovani failed to induce any of these activities. This was found to be the case irrespective of whether amastigotes were alive or killed or opsonized with specific antibodies. Elicited cells preinfected with Leishmania responded normally to secondary stimulation with lipopolysaccharide, but not S. aureus (64% of Staphylococcus alone). In addition, attachment and penetration of L. donovani promastigotes and their subsequent conversion to amastigotes within macrophages failed to induce IL 1 synthesis. The findings of this study indicate that L. donovani has the ability to both evade and suppress the macrophage IL 1 response. Because this monokine provides an obligatory signal during macrophage-dependent T cell activation, evasion of signal transduction for IL 1 synthesis may be related to defects in cell-mediated immunity which occur during infections with this organism.

Animals↗

PubMed1985

Arachidonic acid metabolism by murine peritoneal macrophages infected with Leishmania donovani: in vitro evidence for parasite-induced alterations in cyclooxygenase and lipoxygenase pathways.

Leishmania donovani is an obligate intracellular protozoan that resides within mononuclear phagocytes of infected mammals. Affected human and rodent hosts commonly show abnormalities of T cell function, which may be related to altered macrophage physiology resulting from intracellular parasitism. To examine this possibility, we studied the metabolism of endogenous arachidonyl-phospholipids and [3H]-arachidonyl-phospholipids by murine peritoneal exudate macrophages infected with amastigotes of L. donovani. Our results indicated that infected cells synthesized increased amounts of both cyclooxygenase and lipoxygenase metabolites of arachidonic acid. Increased synthesis of immunoreactive prostaglandin (PG)E2 was evident as early as 1 to 4 hr after infection, was correlated with the fraction of cells infected, and was inhibited by sodium meclofenamate (0.2 and 20 microM) but not nordihydroguaiaretic acid (3 microM). As determined by thin-layer chromatography, infected cells also produced markedly increased amounts of prostaglandin F2 alpha (also inhibited by sodium meclofenamate) with insignificant increases in thromboxane B2 and the stable metabolite of prostacyclin, 6-oxo-PGF1 alpha. In contrast, stimulation of cells with opsonized zymosan resulted in significantly increased synthesis of all four eicosanoids. L. donovani infection was also found to induce marked increases in synthesis of lipoxygenase metabolites of arachidonic acid by infected cells. This was evidenced by increased amounts of [3H]-labeled material in cell extracts that co-migrated with authentic standards of 5 and 12/15-hydroxy-eicosate-traenoic acids in thin-layer chromatograms. Increased synthesis of these products was largely inhibited by both NDGA (3 microM) and sodium meclofenamate (20 and 0.2 microM). Additional evidence for augmentation of 5-lipoxygenase by Leishmania was provided by the demonstration of increased leukotriene-C4 in conditioned medium from infected cells. These results indicate that macrophages infected with L. donovani produce increased amounts of arachidonic acid metabolites with the potential for influencing cellular immune function and the inflammatory response to infection.

Animals↗

PubMed1984

Arachidonic acid metabolism in murine leishmaniasis (Donovani): ex-vivo evidence for increased cyclooxygenase and 5-lipoxygenase activity in spleen cells.

This study examined the metabolism of arachidonic acid (20:4) by splenic mononuclear cells from BALB/c mice infected with Leishmania donovani. Spleen cells removed from mice after either 4 or 8 weeks of infection and cultured in the presence of phytohemagglutinin (PHA) incorporated 60-70% less [3H]thymidine and synthesized 2- to 5-fold more prostaglandin E2 than did spleen cells from normal mice. Inhibition of cyclooxygenation by sodium meclofenamate was associated with restoration of PHA-induced spleen cell blastogenesis. Thin-layer chromatography of spleen cell extracts showed that PHA-stimulated spleen cells from infected animals synthesized 54 and 27% more 5-hydroxyeicosatetraenoic acid (5-HETE) and 12/15-HETE respectively, when compared to spleen cells from noninfected mice. Culture medium of spleen cells from infected mice also contained 25% more immunoreactive leukotriene C4. Nordihydroguaiaretic acid (3 microM) reduced spleen cell synthesis of 12/15-HETE while minimally affecting 5-HETE production. These data indicated that increased cyclooxygenase and lipoxygenase activities in spleen cells from mice infected with L. donovani resulted in the generation of 20:4 metabolites with the capability of altering T-lymphocyte function.

Animals↗

PubMed1984

Pathologic changes in murine leishmaniasis (Leishmania donovani) with special reference to the dynamics of granuloma formation in the liver.

The histopathologic changes in liver, spleen, and bone marrow of BALB/c mice infected for 6 months with Leishmania donovani are described. Granulomas were the most important histologic lesions found; and the dynamics of their formation, collagen deposition, and resolution in the liver were studied. The number of hepatic granulomas increased until the eighth week and then decreased steadily. In contrast to the liver granulomas, those of spleen and bone marrow do not mature or show collagen deposition. Actual granuloma counts in the liver support the idea that mature granulomas revert to poorly formed ones and finally resolve without scarring. BALB/c mice are suitable for the study of the dynamics of granuloma formation and resolution and the survival of L donovani in an in vivo system. The relevance of these changes to the pathology of L donovani infection in man are discussed.

Animals↗

PubMed1983

Interleukin 2 deficiency in murine Leishmaniasis donovani and its relationship to depressed spleen cell responses to phytohemagglutinin.

This study examined the kinetics and mechanisms of depressed spleen cell responses to phytohemagglutinin (PHA) that occur during Leishmania donovani infection of BALB/c mice. In co-culture experiments, neither spleen cells from infected animals nor parasite-infected macrophages suppressed PHA responses of normal spleen cells. In addition, parasite-mediated suppression of PHA-stimulated spleen cell proliferation could not be demonstrated. Mice with 2 wk of infection did manifest an impairment in spleen cell production of interleukin 2 (IL 2) and by 8 wk IL 2 activity in supernatants from these cells was reduced by approximately 95%. This finding was not explained by an alteration in the kinetics of IL 2 production. Furthermore, diminished IL 2 activity in supernatants of PHA-activated spleen cells from infected animals was not caused by suppressive factors in these fluids as shown by their inability to suppress IL 2 stimulation of IL 2-dependent T cells. When spleen cells from mice with 8 wk of infection were cultured with PHA and supplemented with exogenous IL 2, there was an approximately 48% increase in mitogenesis. These data indicate that abnormal PHA-induced spleen cell activation in BALB/c mice with L. donovani infection is associated with impaired production of IL 2. In addition, the observation that supplementation of spleen cells from infected mice with IL 2 resulted in partial reconstitution of the PHA response is consistent with a defect in IL 2 responsiveness.

Animals↗

PubMed1982

Oxidant-mediated damage of Leishmania donovani promastigotes.

Dissemination of Leishmania within the host is related to parasites undergoing unchecked proliferation. We therefore studied the effects of oxidant generating systems on promastigote multiplication by (i) direct determinations of organism proliferation and (ii) the incorporation of [3H]uracil into promastigote nucleoprotein. These two parameters correlated closely as measures of organism replication as demonstrated by parallel suppression of them by the protein synthesis inhibitors puromycin and cycloheximide and the nucleic acid synthesis inhibitors actinomycin D and mitomycin C. Promastigotes showed dose-related susceptibility to reagent and generated hydrogen peroxide (H2O2) as reflected in quantitatively similar decreases in multiplication and [3H]uracil incorporation. These effects were specific for H2O2 as catalase abrogated the dimunition in multiplication. The generation of superoxide anion by acetaldehyde-xanthine oxidase (10 mU/ml) did not alter promastigote replication or nucleoprotein synthesis. These results indicate that Leishmania donovani promastigotes are damaged by H2O2 and that the incorporation of [3H]uracil into promastigote nucleoprotein may be useful for studying the interaction of this parasite with host effector cells.

Acetaldehyde↗