Genetic susceptibility to intracellular infections: Nramp1, macrophage function and divalent cations transport

Nramp1 is one of the few host resistance genes that have been characterized at the molecular level. Nramp1 is an integral membrane protein expressed in the lysosomal compartment of macrophages and is recruited to the membrane of bacterial phagosomes where it affects intracellular microbial replication. Nramp1 is part of a very large gene family conserved from bacteria and man that codes for transporters of divalent cations transporters. We propose that Nramp1 affects the intraphagosomal microbial replication by modulating divalent cations content in this organelle. Both mammalian and bacterial transporters may compete for the same substrate in the phagosomal space.     

Counter-regulatory anti-parasite cytokine responses during concurrent Plasmodium yoelii and intestinal helminth infections in mice

Malaria and helminth infections are two of the most prevalent parasitic diseases globally. While concomitant infection is common, mechanisms contributing to altered disease outcomes during co-infection remain poorly defined. We have previously reported exacerbation of normally non-lethal Plasmodium yoelii malaria in BALB/c mice chronically infected with the intestinal trematode Echinostoma caproni. The goal of the present studies was to determine the effect of helminth infection on IFN-γ and other key cytokines during malaria co-infection in the P. yoelii-E. caproni and P. yoelii-Heligmosomoides polygyrus model systems. Polyclonally stimulated spleen cells from both E. caproni- and H. polygyrus-infected mice produced significantly lower amounts of IFN-γ during P. yoelii co-infection than malaria-only infected mice. Furthermore, the magnitude of IFN-γ suppression was correlated with the relative amounts of IL-4 induced by these helminths (E. caproni = low; H. polygyrus = high), but not IL-10. Concurrent malaria infection also suppressed helminth-associated IL-4 responses, indicating that immunologic counter-regulation occurs during co-infection with malaria and intestinal helminths.     

Human deficiencies in type 1 cytokine receptors reveal the essential role of type 1 cytokines in immunity to intracellular bacteria

Studies on patients with idiopathic, severe infections due to poorly pathogenic mycobacteria and Salmonella have revealed that many of these patients are unable to produce or respond to interferon-gamma (IFN-gamma). This inability results from causative, deleterious genetic mutations in either one of four different genes in the type 1 cytokine cascade, encoding interleukin-12Rbeta1 (IL-12Rbeta1), IL-12p40, IFN-gammaR1 or IFN-gammaR2. The immunological phenotypes resulting from the seven groups of complete or partial deficiencies in type 1 cytokine (receptor) genes that have been distinguished thus far will be summarized and discussed, and placed in a broader context in relation to disease susceptibility.     

The facultative intracellular pathogen Candida glabrata subverts macrophage cytokine production and phagolysosome maturation

Although Candida glabrata is an important human pathogenic yeast, its pathogenicity mechanisms are largely unknown. Immune evasion strategies seem to play key roles during infection, since very little inflammation is observed in mouse models. Furthermore, C. glabrata multiplies intracellularly after engulfment by macrophages. In this study, we sought to identify the strategies that enable C. glabrata to survive phagosome biogenesis and antimicrobial activities within human monocyte-derived macrophages. We show that, despite significant intracellular proliferation, macrophage damage or apoptosis was not apparent, and production of reactive oxygen species was inhibited. Additionally, with the exception of GM-CSF, levels of pro- and anti-inflammatory cytokines were only marginally increased. We demonstrate that adhesion to and internalization by macrophages occur within minutes, and recruitment of endosomal early endosomal Ag 1 and lysosomal-associated membrane protein 1 indicates phagosome maturation. However, phagosomes containing viable C. glabrata, but not heat-killed yeasts, failed to recruit cathepsin D and were only weakly acidified. This inhibition of acidification did not require fungal viability, but it had a heat-sensitive surface attribute. Therefore, C. glabrata modifies the phagosome into a nonacidified environment and multiplies until the host cells finally lyse and release the fungi. Our results suggest persistence of C. glabrata within macrophages as a possible immune evasion strategy.     

Oral lactoferrin prevents body weight loss and increases cytokine responses during herpes simplex virus type 1 infection of mice

Lactoferrin (LF), a multifunctional milk protein, is known to inhibit in vitro infection by viruses such as herpes simplex virus type 1 (HSV-1). We evaluated the influence of LF feeding on the HSV-1 cutaneous infection of mice. Bovine LF was administered to mice and, after 10 d, the mice were infected with HSV-1. LF administration did not affect the viral clearance in the skin, but inhibited the appearance of skin lesions. LF prevented body weight loss and the decrease of splenocyte number associated with HSV-1 infection. LF increased the serum interleukin (IL)-18 level and splenocyte production of interferon-gamma and IL-12. These results suggest that LF feeding was not effective for eradication of the virus, but may contribute to the maintenance of homeostasis and the concomitant increases of cytokine responses during HSV-1 infection.     

In vivo intracellular cytokine production by leukocytes during haemodialysis

Several chronic inflammatory changes undergone during chronic haemodialysis are associated with increased pro-inflammatory cytokine production. Although generation of anaphylatoxins has been incriminated in the untoward effects of haemodialysis, it is still debated whether anaphylatoxins stimulate monocyte secretion of TNF-alpha and IL-1. We demonstrate that peripheral mononuclear cells isolated from healthy controls and cultured with complement-activated autologous serum or recombinant C5a induced high levels of IL-1, IL-1ra, IL-8 and MCP-1, low levels of TNFalpha and sTNFRII but no IL-10 and MIP-1alpha. Cytokine production by leukocytes was investigated by FACS analysis in six patients dialysed consecutively with three equivalent low permeability membranes known to activate the complement to different degrees: polysulfone (F6HPS), cellulose acetate (CA) and cuprophane (CP). Percentage of leukocytes expressing IL-1, IL-1ra, TNF-alpha and IL-8 is increased in patients dialysed with CP. Moreover, we show for the first time that haemodialysis is associated with the production of cytokines by circulating neutrophils. Predialysis plasma levels of MCP-1 and TNFRII did not increase during the dialysis session at the time when anaphylatoxin generation was highest. Dialysis with membranes that activate the complement to a high extent induce activation of leukocytes which may explain chronic complications associated with dialysing with CP.     

IFN-alpha/beta signaling is required for polarization of cytokine responses toward a protective type 1 pattern during experimental cryptococcosis

The antiviral activities of type I IFNs have long been established. However, comparatively little is known of their role in defenses against nonviral pathogens. We examined here the effects of type I IFNs on host resistance against the model pathogenic yeast Cryptococcus neoformans. After intratracheal or i.v. challenge with this fungus, most mice lacking either the IFN-alpha/beta receptor (IFN-alpha/betaR) or IFN-beta died from unrestrained pneumonia and encephalitis, while all wild-type controls survived. The pulmonary immune response of IFN-alpha/betaR-/- mice was characterized by increased expression of IL-4, IL-13, and IL-10, decreased expression of TNF-alpha, IFN-gamma, inducible NO synthetase, and CXCL10, and similar levels of IL-12 mRNA, compared with wild-type controls. Histopathological analysis showed eosinophilic infiltrates in the lungs of IFN-alpha/betaR-/- mice, although this change was less extensive than that observed in similarly infected IFN-gammaR-deficient animals. Type I IFN responses could not be detected in the lung after intratracheal challenge. However, small, but statistically significant, elevations in IFN-beta levels were measured in the supernatants of bone marrow-derived macrophages or dendritic cells infected with C. neoformans. Our data demonstrate that type I IFN signaling is required for polarization of cytokine responses toward a protective type I pattern during cryptococcal infection.     

白细胞介素1细胞信号转导机制研究现状

白细胞介素１是多种炎症增殖性疾病中炎症介导作用极强的细胞因子这一,对其细胞信号转导机制的研究令人瞩目。白细胞介素１与其特异受体结合后,通过某些调节蛋白作用,激活胞膜上或胞浆内的多种磷脂酶,产生多种信使类物质。此外,近年来还发现白细胞介素１可激活多种蛋白激酶和转录调节因子。不同传导途径的细胞信号与白细胞介素１导致的炎症效应密度切相关。     

Activation of rat and human alveolar macrophage intracellular microbicidal activity by a preformed LGL cytokine

An alveolar macrophage-activating factor was released from Percoll fractionated large granular lymphocytes (LGL) within minutes of contact with either the natural killer (NK)-sensitive K562 tumor or heat-killed Staphylococcus aureus. The factor enhanced the intracellular killing of S. aureus without altering the rate of phagocytosis. Factor release was blocked by treatment of LGL with monensin, a carboxylic ionophore that inhibits vesicular traffic, but was unaffected by actinomycin D and cycloheximide pretreatment, suggesting that the cytokine was performed. The cell producing the factor was found only in Percoll fractions containing high concentrations of lytic NK cells and LGL, and the phenotypes of the LGL were HNK-1+ and E rosette-. The macrophage activating factor was a small protein of 10,000 to 20,000 daltons, as determined by gel fractionation, and was sensitive to proteolytic enzymes and heat and pH labile. Active supernatants were devoid of antiviral (interferon; IFN) or interleukin 2 (IL 2) activity, and IFN-beta, IFN-gamma, IL 2, and interleukin 1 were unable to activate staphylococcidal activity, suggesting that the LGL macrophage activating factor was distinguishable from these cytokines.     

IL-18 promotes type 1 cytokine production from NK cells and T cells in human intracellular infection

We investigated the role of IL-18 in leprosy, a disease characterized by polar cytokine responses that correlate with clinical disease. In vivo, IL-18 mRNA expression was higher in lesions from resistant tuberculoid as compared with susceptible lepromatous patients, and, in vitro, monocytes produced IL-18 in response to Mycobacterium leprae. rIL-18 augmented M. leprae-induced IFN-gamma in tuberculoid patients, but not lepromatous patients, while IL-4 production was not induced by IL-18. Anti-IL-12 partially inhibited M. leprae-induced release of IFN-gamma in the presence of IL-18, suggesting a combined effect of IL-12 and IL-18 in promoting M. leprae-specific type 1 responses. IL-18 enhanced M. leprae-induced IFN-gamma production rapidly (24 h) by NK cells and in a more sustained manner (5 days) by T cells. Finally, IL-18 directly induced IFN-gamma production from mycobacteria-reactive T cell clones. These results suggest that IL-18 induces type 1 cytokine responses in the host defense against intracellular infection.     

IFN-gamma-independent autocrine cytokine regulatory mechanism in reprogramming of macrophage responses to bacterial lipopolysaccharide.

Macrophages are now well recognized to have a critical role in both innate and acquired immunity. The sentinel macrophage function is highly regulated and serves to allow for intrinsic plasticity of the innate immune responses to potential environmental signals. However, the mechanisms underlying the dynamic properties of the cellular arm of innate immunity are poorly understood. Therefore, we have conducted a series of in vitro studies to evaluate the contribution of immunoregulatory cytokines, such as IFN-gamma, IL-10, and IL-12, in modulation of macrophage responses. We found that macrophages from IFN-gamma knockout (IFN-gamma(-/-)) mice exhibit only marginal LPS-induced TNF-alpha, IL-12, and NO responses, all of which can be fully restored in the presence of rIFN-gamma. Pretreatment with substimulatory LPS concentrations led to reprogramming of IFN-gamma(-/-) macrophage responses in a dose-dependent manner that manifested by an increased TNF-alpha and IL-12, but not NO, production upon the subsequent LPS challenge. These reprogramming effects were substantially attenuated and profoundly enhanced in macrophages from IL-12(-/-) and IL-10(-/-) mice, respectively, as compared with those modulated in macrophages from the congenic wild-type mice. LPS-dependent reprogramming was also fully reproduced in macrophages isolated from SCID mice after immunodepletion of NK cells. Our data strongly imply that cytokine (TNF-alpha and IL-12), but not NO, responses in macrophages may, at least in part, be governed by an autocrine IFN-gamma-independent regulatory mechanism reciprocally controlled by IL-10 and IL-12. This mechanism may serve as an alternative/coherent pathway to the canonical IFN-gamma-dependent induction of antimicrobial and tumoricidal activity in macrophages.     

Current understanding of the interplays between host hormones and plant viral infections

Phytohormones mediate plant development and responses to stresses caused by biotic agents or abiotic factors. The functions of phytohormones in responses to viral infection have been intensively studied, and the emerging picture of complex mechanisms provides insights into the roles that phytohormones play in defense regulation as a whole. These hormone signaling pathways are not simple linear or isolated cascades, but exhibit crosstalk with each other. Here, we summarized the current understanding of recent advances for the classical defense hormones salicylic acid (SA), jasmonic acid (JA), and ethylene (ET) and also the roles of abscisic acid (ABA), auxin, gibberellic acid (GA), cytokinins (CKs), and brassinosteroids (BRs) in modulating plant–virus interactions.     

Quantifying macrophage defects in type 1 diabetes

Macrophages from animals prone to autoimmune (type 1) diabetes differ from those of diabetes-resistant animals in processing and clearing apoptotic cells. Using in vitro time-course assays of the number of engulfed apoptotic cells observed within macrophages, we quantified these differences in non-obese diabetic (NOD) versus Balb/c mice. Simple models lead to several elementary parameter estimation techniques. We used these to compute approximate rates of macrophage engulfment and digestion of apoptotic cells from basic features of the data (such as initial rise-times, phagocytic index and percent phagocytosis). Combining these estimates with full fitting of a sequence of model variants to the data, we find that macrophages from normal (Balb/c) mice engulf apoptotic cells up to four times faster than macrophages from the diabetes-prone (NOD) mice. Further, Balb/c macrophages appear to undergo an activation step before achieving their high engulfment rate. In NOD macrophages, we did not see evidence for this activation step. Rates of digestion of engulfed apoptotic cells by macrophages are similar in both types. Since macrophage clearance is an important mechanism of disposal of self-antigen, these macrophage defects could potentially be a factor in predisposition to type 1 diabetes.     

Intracellular iron availability modulates the requirement for Leishmania Iron Regulator 1 (LIR1) during macrophage infections

The Leishmania plasma membrane transporter Leishmania Iron Regulator 1 (LIR1) facilitates iron export and is required for parasite virulence. By modulating macrophage iron content, we investigated the host site where LIR1 regulates Leishmania amazonensis infectivity. In bone marrow-derived macrophages, LIR1 null mutants demonstrated a paradoxical increase in virulence during infections in heme-depleted media, while wild-type growth was inhibited under the same conditions. Loading the endocytic pathway of macrophages with cationized ferritin prior to infection reversed the effect of heme depletion on both strains. Thus, LIR1 contributes to Leishmania virulence by protecting the parasites from toxicity resulting from iron accumulation inside parasitophorous vacuoles.     

Effect of prolonged exercise and carbohydrate ingestion on type 1 and type 2 T lymphocyte distribution and intracellular cytokine production in humans.

The present study was undertaken to examine the role of the exercise-induced stress hormone response on the regulation of type 1 and type 2 T lymphocyte intracellular cytokine production. Subjects performed 2.5 h of cycling exercise at 65% maximal O2 uptake while ingesting a 6.4% carbohydrate (CHO) solution, 12.8% CHO solution, or a placebo. Peripheral whole blood samples were stimulated and stained for T lymphocyte surface antigens (CD4 and CD8). Cells were then permeabilized, stained for intracellular cytokines, and analyzed using flow cytometry. Exercise resulted in a decrease (P < 0.05) in the number and percentage of IFN-gamma positive CD4+ and CD8+ T lymphocytes. These stimulated cells produced less IFN-gamma immediately postexercise (P < 0.05) and 2-h postexercise (P < 0.05) compared with preexercise. However, CHO ingestion, which attenuated the exercise-induced stress hormone response compared with placebo (P < 0.05), prevented both the decrease in the number and percentage of IFN-gamma-positive CD4+ and CD8+ T lymphocytes and the suppression of IFN-gamma production from stimulated CD4+ and CD8+ T lymphocytes. There was no effect of exercise on the number of, or cytokine production from, IL-4-positive CD4+ or CD8+ T lymphocytes. These data provide support for the role of exercise-induced elevations in stress hormones in the regulation of type 1 T lymphocyte cytokine production and distribution.