Tumor necrosis factor alpha (TNF-alpha) receptor-II is required for TNF-alpha-induced leukocyte-endothelial interaction in vivo

Tumor necrosis factor-alpha (TNF-alpha) binds to 2 distinct cell-surface receptors: TNF-alpha receptor-I (TNFR-I: p55) and TNF-alpha receptor-II (TNFR-II: p75). TNF-alpha induces leukocyte adhesion molecules on endothelial cells (ECs), which mediate 3 defined steps of the inflammatory response; namely, leukocyte rolling, firm adhesion, and transmigration. In this study, we have investigated the role of p75 in TNF-alpha-induced leukocyte adhesion molecules using cultured ECs derived from wild-type (WT), p75-null (p75-/-), or p55-null (p55-/-) mice. We observed that p75 was essential for TNF-alpha-induced E-selectin, vascular cell adhesion molecule 1 (VCAM-1), and intercellular adhesion molecule 1 (ICAM-1) expression. We also investigated the putative role of p75 in inflammation in vivo using an intravital microscopic approach with a mouse cremaster muscle model. TNF-alpha-stimulated leukocyte rolling, firm adhesion to ECs, and transmigration were dramatically reduced in p75-/- mice. Transplanted WT cremaster in p75-/- mice showed a robust leukocyte rolling and firm adhesion upon TNF-alpha activation, suggesting that the impairment in EC-leukocyte interaction in p75-/- mice is due to EC dysfunction. These results demonstrate, for the first time, that endothelial p75 is essential for TNF-alpha-induced leukocyte-endothelial-cell interaction. Our findings may contribute to the identification of novel p75-targeted therapeutic approaches for inflammatory diseases.     

Accumulation of eosinophils in intestine-draining mesenteric lymph nodes occurs after Trichuris muris infection

Eosinophils have recently been demonstrated capable of localizing to lymph nodes that drain mucosal surfaces, in particular during T helper 2 (Th2) responses. Resistance of mice to infection with the gastrointestinal nematode Trichuris muris depends critically on mounting of a Th2 response and represents a useful model system to investigate Th2 responses. Following infection of resistant BALB/c mice with T. muris, we observed accumulation of eosinophils in intestine-draining mesenteric lymph nodes (MLNs). The accumulation of MLN eosinophils was initiated during the second week of infection and peaked during worm expulsion. In contrast, we detected a comparably late and modest increase in eosinophil numbers in the MLNs of infected susceptible AKR mice. MLN eosinophils localized preferentially to the medullary region of the lymph node, displayed an activated phenotype and contributed to the interleukin-4 (IL-4) response in the MLN. Despite this, mice genetically deficient in eosinophils efficiently generated IL-4-expressing CD4(+) T cells, produced Th2 cytokines and mediated worm expulsion during primary T. muris infection. Thus, IL-4-expressing eosinophils accumulate in MLNs of T. muris-infected BALB/c mice but are dispensable for worm expulsion and generation of Th2 responses, suggesting a distinct or subtle role of MLN eosinophils in the immune response to T. muris infection.     

NK1.1+ cell depletion in vivo fails to prevent protection against infection with the murine nematode parasite Trichuris muris

Protection against the murine nematode parasite Trichuris muris has been shown to involve interleukin 4 (IL-4). NK1.1+ T cell receptor alphabeta+ cells, designated Natural Killer T (NKT) cells, produce a large amount of IL-4 in response to anti-CD3 stimulation and numerous pieces of evidence suggest that NKT cells provide the initial source of IL-4 for T helper 2 (Th2) priming. These observations allow the hypothesis that NKT cells produce a large amount of IL-4 in response to T. muris infection and augment Th2 responses and IL-4 production, thus achieving protection against T. muris. To investigate the involvement of NKT cells in protection against T. muris infection, NK1.1+ cell-depleted B10.BR mice were prepared by anti-NK1.1 monoclonal antibody injection. Efficient expulsion of T. muris worms occurred in NK1.1+ cell-depleted infected mice, and the expulsion kinetics of T. muris worms, the levels of IL-4 production by mesenteric lymph node cells, and the kinetics of the specific IgG1 and IgG2a responses to T. muris were similar to those in mouse IgG-treated or non-treated control B10.BR mice. These observations suggest that NK1.1+ cells and NKT cells are not involved in the induction of Th2 responses and protective immunity to T. muris infection.     

The role of IL-4, IL-13 and IL-4Ralpha in the development of protective and pathological responses to Trichinella spiralis

T helper type 2 (Th2) responses have been shown to be important in protective responses to gastrointestinal (GI) helminth infections and in the development of the intestinal pathology accompanying expulsion of the parasite. Different inbred mouse strains have been shown to develop different cytokine profiles following infection with GI helminths with increased resistance observed in those strains where Th2 cytokines predominate. The aim of this study was to determine the role of IL-4, IL-13 and IL-4Ralpha and the impact of host background on the development of the protective and pathological responses induced by infection with the gastrointestinal helminth Trichinella spiralis. IL-4, IL-13 and IL-4Ralpha were required for the generation of Th2 responses to T. spiralis; however, the role these responses play in the development of protection and enteropathy was less clear. IL-4Ralpha-deficiency mice resulted in substantially reduced parasite expulsion, intestinal pathology and Th2 responses. Similarly, lack of IL-13 resulted in inhibited expulsion and the development of enteropathy. Although Th2 responses were reduced in BALB/c IL-4-/- mice, neither expulsion nor enteropathy were different from wild-type mice. In contrast, C57BL/6 IL-4-/- exhibited delayed expulsion and reduced pathology, suggesting that host genetics are important in the function of individual cytokines. Thus, differences in background genotype may be an important component in the development host protection and the development of intestinal pathology accompanying the loss of GI helminths.     

Interleukin-4 causes susceptibility to invasive pulmonary aspergillosis through suppression of protective type I responses

Aspergillus fumigatus, an opportunistic fungal pathogen, causes multiple allergic and nonallergic airway diseases. Invasive pulmonary aspergillosis (IPA) is a nonallergic, life-threatening disease of immunocompromised patients. In a murine model of IPA, interleukin (IL)-4-deficient (IL-4-/-) BALB/c mice were used to examine the role of IL-4 in lung pathology and immune responses. IL-4-/- mice were more resistant than wild-type mice to infection caused by multiple intranasal injections of viable A. fumigatus conidia. Resistance was associated with decreased lung inflammatory pathology, impaired T helper (Th)-2 responses (including lung eosinophilia), and an IL-12-dependent Th1 response. In contrast, development of host-detrimental antifungal Th2 cells occurred in IL-12-/- and interferon-gamma-/- mice and in IL-4-/- mice when subjected to IL-12 neutralization. These results demonstrate that IL-4 renders mice susceptible to infection with A. fumigatus by inhibition of protective Th1 responses. IL-4 appears to have a distinct role in the pathogenesis of allergic and nonallergic lung diseases caused by the fungus.     

Modulation of cytokine production and response phenotypes in murine trichuriasis

BALB/K mice are usually resistant to infection with the intestinal nematode parasite Trichuris muris and exhibit a Th2 dominated (IL-5, IL-9) response. Conversely in B10.BR mice, which are unable to expel T. muris, Th1 type (IFN-gamma producing) cells predominate. We have manipulated the course of infection in these two strains of mice such that the period of host-parasite contact is extended in the former and curtailed in the latter. Extension of host-parasite contact in BALB/K mice beyond normal (day 21) resulted in the modulation of cytokines produced by in vitro concanavalin A (Con-A) stimulated MLNC away from IL-5 and IL-9 (Th2-type cytokines) in favour of the Th1-type cytokine IFN-gamma. Curtailment of host parasite contact in B10.BR mice to less than 21 days resulted in elevated production of IL-5 and IL-9 by MLNC in the absence of elevated IFN-gamma levels. Thus modulation of expulsion phenotype also modulates cytokine production by T-cells in the MLN draining the site of infection, with a Th2 response being associated with resistance and a Th1 type response with the inability to expel the parasite. Mechanisms by which the modulated cytokine profiles arise are discussed.     

Mucosal cytokine and antigen-specific responses to Cryptosporidium parvum in IL-12p40 KO mice

Studies of cellular immune responses to Cryptosporidium parvum have been limited in part by lack of suitable animal models. IL-12p40(-/-)mice are susceptible to initial infection with C. parvum but recover within 2 weeks, rendering the animals resistant to reinfection. Because the host responses that determine duration and severity of primary infection are not yet understood, we studied the cellular immune response to primary infection with C. parvum in IL-12p40(-/-)mice and also explored possible mechanisms for this response. Female IL-12p40(-/-)mice were inoculated with 10,000 oocysts. Uninfected age-matched mice served as controls. At different time intervals following exposure to oocysts, mice were sacrificed and their intestine, spleen, and mesenteric lymph node tissues were harvested. Cellular immune responses to C. parvum were characterized. Infection of IL-12p40(-/-)mice induced changes in the gene expression of the cytokines IFN-gamma, IL-4, IL-15, IL-18, TNF-alpha and TGF-beta during primary infection. There was also a significant increase in total numbers of lymphocytes and CD19/CD62L-expressing cells in mesenteric lymph nodes. These MLN cells exhibited increased antigen-specific proliferation and cytokine production (IL-6 and IFN-gamma) levels when stimulated in vitro. These observations delineate the cellular immune responses during acute C. parvum infection of the IL-12p40(-/-)mouse model.     

Mechanisms of TNF-alpha-induced insulin resistance.

There is now substantial evidence linking TNF-alpha to the presentation of insulin resistance in humans, animals and in vitro systems. We explored the relationship between TNF-alpha and insulin resistance using knockout mice deficient for either TNF-alpha or one or both of its receptors, p55 and p75. In studies of TNF-alpha-deficient knockout mice with diet-induced obesity, obese TNF-alpha knockouts responded to an exogenous dose of insulin or glucose much more efficiently than TNF-alpha wild-type animals. This finding suggests that deletion of TNF-alpha leads to increased insulin sensitivity, ie decreased insulin resistance. In studies using genetically obese ob/ob mice, TNF-alpha receptor wild-type and p75 receptor knockout animals developed a pronounced hyperinsulinemia and transient hyperglycaemia, whereas p55 receptor and double-knockout animals did not. Moreover, in glucose and insulin tolerance tests, we found that p75 knockout animals exhibited profiles identical to those of the wild-type animals, but that p55 knockout animals and double mutants showed a mild improvement in insulin sensitivity, relative to the wild type. Since the improvement in sensitivity was slightly greater with double mutants, p55 alone cannot be responsible for TNF-alpha's promotion of insulin resistance in obese mice, despite the likelihood that it is more important than p75. How TNF-alpha-related insulin resistance is mediated is not fully clear, although phosphorylation of serine residues on IRS-1 has previously been shown to be important. When we monitored Glut 4 expression in obese TNF-alpha wild-type and knockout mice, we found no convincing evidence that TNF-alpha mediation of the down-regulation of Glut 4 mRNA expression is responsible for insulin resistance. However, we found an approximately 2-fold increase in insulin-stimulated tyrosine phosphorylation of the insulin receptor in the muscle and adipose tissue of TNF-alpha knockout mice, suggesting that insulin receptor signalling is an important target for TNF-alpha. Other possible mediators of TNF-alpha-induced insulin resistance include circulating free fatty acids (FFAs) and leptin.     

Mast cells, eosinophils and antibody-mediated cellular cytotoxicity are not critical in resistance to Trichuris muris

The murine intestinal nematode Trichuris muris provides an invaluable model of human infection with T. trichiura. Hence, analysis of the immunological responses in the mouse may elucidate the mechanisms of immunity to trichuriasis in man. The work described here investigates the roles of eosinophils, mast cells and antibody-dependent cell-mediated cytotoxicity (ADCC) in the elimination of T. muris from the host gut. Following ablation of IL-5, and hence eosinophilia, mice usually resistant to T. muris infection remained so. Further, blocking the stem cell factor receptor, c-kit, to facilitate complete ablation of mast cells over the period of parasite expulsion in resistant mice had no effect on the development of protective immunity. Therefore it can be deduced that eosinophils and mast cells are not critical in resistance. In addition to these studies, the role of antibody-mediated cellular cytotoxic mechanisms was investigated via the analysis of an infection time course in Fc gamma R-/- mice. These animals, on a resistant background, were fully immune and expelled the parasites before development of the adult stage. Thus this model provides evidence against a major role for ADCC in resistance to infection with T. muris. The studies described here have eliminated some of the major effector mechanisms traditionally associated with helminth infection, and work continues to elucidate the critical immune responses associated with resistance.     

Interleukin-4 influences the production of microfilariae in a mouse model of Brugia infection

Sub-cutaneous infection of interleukin (IL)-4-/- mice on the BALB/c background with third stage larva (L3) of Brugia pahangi revealed an altered cytokine profile consistent with the absence of the Th2 promoting cytokine IL-4. Splenocytes from IL-4-/- mice secreted significantly more antigen (Ag)-specific IL-2 and interferon-gamma and significantly less Ag-specific IL-5, compared to those from L3-infected wild-type mice. However, levels of Ag-specific IL-13 were similar between groups. Despite the alteration in immune responses, there was no significant difference in recovery of developing worms from the peritoneal cavity of the two strains of mice at any time postinfection. However, at later time points of infection, the IL-4-/- mice contained large numbers of microfilariae (Mf) in the peritoneal cavity while the wild-type mice contained comparatively few Mf. The differences in Mf levels appear to relate to differences in worm fecundity in the two strains of mice, with adult female worms from the wild-type mice containing few developing Mf. Moreover, implantation of sexually mature adult female worms into the peritoneal cavity of both strains of mice resulted in equal levels of Mf, confirming that the primary role of IL-4 is to limit fecundity during the maturation phase of infection.     

IL-33: a Janus cytokine

Interleukin (IL) 33, a member of the IL-1 family, is the ligand of ST2 that is expressed mainly on activated Th2 cells and mast cells. IL-33 can skew a predominantly Th1 cell population to a mainly Th2 cells phenotype in vivo. IL-33 messenger RNA is expressed early during infection of the intestinal-dwelling nematode Trichuris muris in mice. IL-33 treatment enhances resistance to Trichuris infection. IL-33 also effectively attenuates sepsis by mobilising the innate cells, neutrophils, to the site of infection, helping to clear the pathogens. Thus, IL-33 may be evolutionally preserved for the host defence against infections. IL-33 can reduce an ongoing atherosclerosis in ApoE(-/-) mice and attenuate adipocytes mainly by inducing the production of type II cytokines. In contrast, IL-33 can also exacerbate allergy and the inflammation in collagen-induced or serum-induced arthritis. Hence, IL-33 is a double-edged sword, and targeting IL-33 should be approached with caution.     

RELMbeta/FIZZ2 is a goblet cell-specific immune-effector molecule in the gastrointestinal tract

Gastrointestinal (GI) nematode infections are an important public health and economic concern. Experimental studies have shown that resistance to infection requires CD4(+) T helper type 2 (Th2) cytokine responses characterized by the production of IL-4 and IL-13. However, despite >30 years of research, it is unclear how the immune system mediates the expulsion of worms from the GI tract. Here, we demonstrate that a recently described intestinal goblet cell-specific protein, RELMbeta/FIZZ2, is induced after exposure to three phylogenetically distinct GI nematode pathogens. Maximal expression of RELMbeta was coincident with the production of Th2 cytokines and host protective immunity, whereas production of the Th1 cytokine, IFN-gamma, inhibited RELMbeta expression and led to chronic infection. Furthermore, whereas induction of RELMbeta was equivalent in nematode-infected wild-type and IL-4-deficient mice, IL-4 receptor-deficient mice showed minimal RELMbeta induction and developed persistent infections, demonstrating a direct role for IL-13 in optimal expression of RELMbeta. Finally, we show that RELMbeta binds to components of the nematode chemosensory apparatus and inhibits chemotaxic function of a parasitic nematode in vitro. Together, these results suggest that intestinal goblet cell-derived RELMbeta may be a novel Th2 cytokine-induced immune-effector molecule in resistance to GI nematode infection.     

Distinct roles for IL-13 and IL-4 via IL-13 receptor alpha1 and the type II IL-4 receptor in asthma pathogenesis

IL-13 and IL-4 are central T helper 2 (Th2) cytokines in the immune system and potent activators of inflammatory responses and fibrosis during Th2 inflammation. Recent studies using Il13ra1(-/-) mice have demonstrated a critical role for IL-13 receptor (IL-13R) alpha1 in allergen-induced airway responses. However, these observations require further attention especially because IL-4 can induce similar lung pathology to IL-13, independent of IL-13, and is still present in the allergic lung. Thus, we hypothesized that IL-13Ralpha1 regulates IL-4-induced responses in the lung. To dissect the role of IL-13Ralpha1 and the type I and II IL-4Rs in experimental asthma, we examined lung pathology induced by allergen, IL-4, and IL-13 challenge in Il13ra1(-/-) mice. We report that IL-13Ralpha1 is essential for baseline IgE production, but Th2 and IgE responses to T cell-dependent antigens are IL-13Ralpha1-independent. Furthermore, we demonstrate that increased airway resistance, mucus, TGF-beta, and eotaxin(s) production, but not cellular infiltration, are critically dependent on IL-13Ralpha1. Surprisingly, our results identify a CCR3- and IL-13Ralpha1-independent pathway for lung eosinophilia. Global expression profiling of lungs from mice stimulated with allergen or IL-4 demonstrated that marker genes of alternatively activated macrophages are differentially regulated by the type I and type II IL-4R. Taken together, our data provide a comprehensive mechanistic analysis of the critical role by which IL-13Ralpha1 mediates allergic lung pathology and highlight unforeseen roles for the type II IL-4R.     

小鼠感染泡球蚴后细胞因子水平的变化

目的　观察人工接种泡球蚴(EM)昆明小鼠体内6种细胞因子水平的动态变化,研究泡球蚴病的免疫学机制。　方法　实验组和对照组小鼠分别腹腔接种泡球蚴及生理盐水,持续观察260d,收集各组血清用ELISA法检测血清中白细胞介素2(IL2)、γ干扰素 (IFNγ)、肿瘤坏死因子α(TNFα)、IL4、IL5、IL10水平。　结果　实验组6种细胞因子水平始终高于正常对照组,其中辅助性T细胞1(Th1)类细胞因子IL2水平在感染后 80d达到峰值,感染140d后迅速降低;TNFα水平在感染后40d较对照组明显上升,感染100d左右达到峰值,140d后迅速下降;IFNγ在感染80d后达到峰值,140d后缓慢下降;而在80d以前,辅助性T细胞2(Th2)类细胞因子IL4、IL5、IL10维持在较低水平;100d后,这3类细胞因子明显上升,其中IL4、IL10水平在100d达到峰值,IL5水平在140d达到峰值,以后维持在较高水平。　结论　Th2细胞介导的体液免疫与Th1细胞介导的细胞免疫共同参与了宿主抗棘球蚴免疫。感染早期以Th1细胞介导的细胞免疫应答为主,感染中晚期转化为以Th2细胞介导的体液免疫为主。     

Contrasting effects of Trichinella spiralis and Trichuris muris antigens on the infection by Leishmania infantum in BALB/c mice

The interaction of Trichinella spiralis and Trichuris muris derived antigens with the infection by Leishmania infantum was investigated in BALB/c mice. Infection with 10(6) promastigotes of L. infantum did not induce relevant serum antibody (IgG subclasses), nor cytokine (IFN-gamma, IL-4) responses despite that mice could partially control the infection. Immunization with T. spiralis activated a moderate IgG1 and secondarily an IgG2a anti-leishmanial response whereas immunization with T. muris elicited only a weak and late activation of IgG1 anti-leishmanial response. Immunization with T. muris caused an elevation of serum IFN-gamma levels which was drastically reinforced by the L. infantum infection, and that was accompanied by almost complete parasitological cure of infected mice. Immunization with T. spiralis induced an elevation of serum IL-4 levels but this response was greatly (about 60%) neutralized by the infection with L. infantum, and this was associated to exacerbation of the infection.     

Serum levels of tumor necrosis factor-alpha (TNF-alpha), interleukin 6 (IL-6), and their soluble receptors in coal workers' pneumoconiosis

The aim of this study was to investigate whether systemic tumor necrosis factor alpha (TNF-alpha), soluble TNF-alpha receptors (p55, p75), interleukin 6 (IL-6), and soluble IL-6 receptor could be markers of biological activities of coal workers' pneumoconiosis (CWP). The study population was composed of 182 Chinese retired coal miners who had similar dust exposure histories. Among them, 71 were cases with CWP and 111 were controls. Chest radiographs were classified according to International Labour Organization Criteria (ILO, 1980). Individual dust exposure variables were estimated from work histories, and smoking information was obtained from interviews. Serum concentrations of TNF-alpha, TNF-alpha receptors (p55, p75), IL-6, and IL-6 receptor were measured by ELISA techniques. Mean serum levels of p55, p75 and IL-6 were significantly higher in cases than in controls (P < or = 0.01 for each comparison by crude analyses). Results from logistic regression models, adjusted for age, dust exposure variables, and smoking habits, found similar associations between soluble p55 and p75 levels and the presence of CWP. Linear regression analysis revealed that CWP radiographic stage (by ILO criteria) was significantly correlated with the individual serum concentrations of p55, p75 and IL-6. Serum concentrations of all measured cytokines were notcorrelated to age, dust exposure, or smoking, but there were correlations between soluble p75 and p55 levels, and between p75 and IL-6 levels. The results of this study suggest that serum levels of TNF receptors and IL-6 are associated with the fibrotic process of CWP and serum cytokine levels may be correlated with the severity of CWP.     

Altered Serum Cytokine Profiles in Relapse Phase of Relapsing-Remitting Multiple Sclerosis

Background: Multiple sclerosis (MS) is a demyelinating disease of the central nervous system and cytokines may play a role in the development of MS lesions. Objective: To determine levels of different cytokines in patients with relapsing-remitting MS (RR-MS) compared to healthy controls. Methods: Profiles of pro-inflammatory, Th1-, Th2-, and Th17-related cytokines were compared by quantitative multiplexed ELISA-based chemiluminescent assay in 44 RR-MS and 44 healthy age- and sex-matched individuals from the same ethnicity. Results: Among pro-inflammatory cytokines, the levels of IL-6 (p=0.003), IL-8 (p=0.05) and TNF-α (p=0.002) were higher in patients than controls, though IL-4 and IL-10 as well as ΣTh2 cytokines were lower in patients (p=0.05, p=0.02 and p=0.05, respectively). After gender classification, the higher levels of IL-4 in male patients remained significant and IL-13 also showed significantly higher levels in male patients compared to male controls (p=0.003 and p=0.05, respectively). A significant negative correlation was detected between EDSS and IL-10 or ΣTh2 levels (p=0.005). In addition, IL-1α (r=0.4, p=0.05) and IFN-γ (r=0.35, p=0.05) were also directly correlated with EDSS in female patients. Conclusions: Patients with RR‑MS who are in the relapse clinical phase exhibit higher levels of pro-inflammatory cytokines and reduction in protective Th2-related cytokines.     

The lack of suppressor of cytokine signalling-1 (SOCS1) protects mice from the development of cerebral malaria caused by Plasmodium berghei ANKA

Cerebral malaria is a severe complication of infection with Plasmodium berghei ANKA involving the Th1 cytokines TNF-alpha and IFN-gamma. Suppressor of cytokine signalling-1 (SOCS1) is an important component in the regulatory cascade controlling inflammatory responses and signalling through IFN-gamma. Contrary to the expectation that SOCS1-deficient mice, in which IFN-gamma responses are uncontrolled and which are more sensitive to IFN-gamma, may show heightened susceptibility, mice lacking SOCS1 were protected from cerebral malaria. Unlike the controls and despite similar parasitaemia, infected SOCS1 null mice showed no inflammation or haemorrhaging in the brains. Mice lacking SOCS1 exhibited decreased splenic cellularity and a reduced ratio of CD4 : CD8 lymphocytes, which were maintained during infection. However, the ratio of IFN-gamma to IL-4 mRNA expression during infection was similar in SOCS1 -/- and control mice suggesting that a dramatic shift in the ratio of Th1 : Th2 responses does not account for the resistance to disease. Resistance conferred by the lack of SOCS1 is specific since the related SOCS2, also implicated in Th1-mediated responses, did not seem to be involved in the development of disease. Understanding the mechanism by which SOCS1 deficiency protects mice from cerebral malaria may allow the manipulation of its activity and alleviate pathology.