Nerve growth factor (NGF) could be involved in the development of hyperalgesia as well as in nervous remodeling consequence of inflammation. Both dysmotility and increase of visceral sensitivity have been described in functional gastrointestinal disorders such as irritable bowel syndrome. Trichinella spiralis-infected rats show an exacerbated spontaneous motility and a significant increase of the excitatory response to cholecystokinin (CCK), both associated with a reversible inflammatory process and the hypertrophy of the muscle layers. In this study we determined the intestinal expression of NGF mRNA by polymerase chain reaction and NGF by enzyme-linked immunosorbent assay. We implanted serosal strain gauge transducers on duodenum, jejunum, and ileum of anesthetized Sprague-Dawley rats to record circular muscle contractions. The experimental protocol included the evaluation of intestinal spontaneous motor activity (SMA), the response to CCK-8, and the ascending contraction induced by electrical mucosal stimulation. This protocol was performed in healthy and infected nontreated rats, in healthy rats with an NGF antibody treatment (1.6 mg/rat i.p.), and in infected rats with the same treatment applied at 0 or 3 days postinfection. NGF and NGF mRNA levels in the bowel were increased during inflammation. Although anti-NGF treatments did not prevent or reverse inflammatory response, the treatment was effective in preventing the motor alterations induced by the T. spiralis infection, i.e., inhibited increased SMA, reversed altered response to CCK, and reversed in part exacerbated response to electrical stimulation. (+info)
Effect of cyclosporin A on immunological response in lungs of guinea pigs infected with Trichinella spiralis.
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The effects of cyclosporin A (CsA), a potent immunosuppressive drug with antiparasitic activity, on the innate immunological response in guinea pig lungs during an early period (6th and 14th days) after T. spiralis infection were studied. CsA treatment of T. spiralis-infected guinea pigs caused a significant attenuation of immunological response in lungs by decreasing lymphocyte infiltration into pulmonary alveolar space, inhibiting alveolar macrophage superoxide anion production and lowering both the production of NO metabolites measured in bronchoalveolar lavage fluid and expression of the iNOS protein in lung homogenates, allowing us to speculate that the T. spiralis-dependent immunological response is dependent on lymphocyte T function. Interestingly, CsA itself had a pro-inflammatory effect, promoting leucocyte accumulation and macrophage superoxide production in guinea pig lungs. This observation may have a relevance to the situation in patients undergoing CsA therapy. Macrophage expression of the iNOS protein, evaluated by immunoblotting was not influenced by treatment of animals with CsA or anti-TGF-antibody, indicating different regulation of the guinea pig and murine enzymes. (+info)
Nucleotidase cascades are catalyzed by secreted proteins of the parasitic nematode Trichinella spiralis.
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Extracellular nucleotides are signaling molecules whose receptor-mediated effects are involved in a variety of physiological responses in mammalian tissues. An overwhelming body of data indicate that inflammatory and other immune responses can be modulated by the availability and local concentrations of nucleotides via nucleotide receptor signaling, but this is only just beginning to be investigated in the context of infectious disease. Evidence is provided here that the parasitic nematode Trichinella spiralis can catalyze the conversion and thus modulate both the availability and concentration of extracellular nucleotides by means of the following secreted exoenzymes: apyrase, 5'-nucleotidase, and adenosine deaminase. These enzymes were characterized in terms of substrate specificity, kinetic behavior, pH, divalent cation preferences, and response to a series of compounds. The secreted 5'-nucleotidase was identified as a protein with an apparent molecular mass of 67 kDa after N-terminal amino acid sequencing of the purified protein. The presence of adenosine deaminase was confirmed in the secreted products by Western blotting with an antibody against a mammalian enzyme, as a protein with an apparent molecular mass of 38 kDa. These secreted proteins constitute an enzymatic cascade which catalyzes the degradation of extracellular nucleotides, with a potential physiological role in the regulation of purinergic signaling. (+info)
IL-18 regulates intestinal mastocytosis and Th2 cytokine production independently of IFN-gamma during Trichinella spiralis infection.
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Expulsion of the gastrointestinal nematode Trichinella spiralis is associated with pronounced mastocytosis mediated by a Th2-type response involving IL-4, IL-10, and IL-13. Here we demonstrate that IL-18 is a key negative regulator of protective immune responses against T. spiralis in vivo. IL-18 knockout mice are highly resistant to T. spiralis infection, expel the worms rapidly and subsequently develop low levels of encysted muscle larvae. The increased speed of expulsion is correlated with high numbers of mucosal mast cells and an increase in IL-13 and IL-10 secretion. When normal mice were treated with rIL-18 in vivo, worm expulsion was notably delayed, and the development of mastocytosis and Th2 cytokine production was significantly reduced. The treatment had no effect on intestinal eosinophilia or goblet cell hyperplasia but specifically inhibited the development of mastocytosis. Addition of rIL-18 to in vitro cultures of bone marrow-derived mast cells resulted in a significant reduction in cell yields as well as in the number of IL-4-secreting mast cells. In vivo treatment of T. spiralis-infected IFN-gamma knockout mice with rIL-18 demonstrated that the inhibitory effect of IL-18 on mastocytosis and Th2 cytokine secretion is independent of IFN-gamma. Hence, IL-18 plays a significant biological role as a negative regulator of intestinal mast cell responses and may promote the survival of intestinal parasites in vivo. (+info)
Intestinal nematode infection ameliorates experimental colitis in mice.
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Epidemiological studies suggest that inflammatory bowel disease (IBD) is common in developed countries and rare in countries where intestinal nematode infections are common. T cells are critical in many immune responses, including those associated with IBD and nematode infection. Among the distinct T helper (Th) cell subsets, Th1-type immune response is predominantly associated with Crohn's disease, while many nematode infections generate a strong Th2 response. The reciprocal cross regulation between Th1 and Th2 cells suggests that generation of a Th2 response by nematodes could prevent or reduce the effects of Th1-mediated diseases. In the present study, we investigated the effect of polarizing the immune response toward the Th2 type, using intestinal nematode infection, on subsequent experimental colitis. Mice were infected with the intestinal nematode Trichinella spiralis and allowed to recover before colitis was induced with dinitrobenzene sulfonic acid. The mice were sacrificed postcolitis to assess colonic damage macroscopically, histologically, and by myeloperoxidase (MPO) activity and Th cytokines. Prior nematode infection reduced the severity of colitis both macroscopically and histologically together with a decreased mortality and was correlated with a down-regulation of MPO activity, Th1-type cytokine expression in colonic tissue, and emergence of a Th2-type immune response. These results indicate a protective role of nematode infection in Th1 cell-driven inflammation and prompt consideration of a novel therapeutic strategy in IBD based on immunological distraction. (+info)
Nasal immunization with homogenate and peptide antigens induces protective immunity against Trichinella spiralis.
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Mice were successfully immunized against the intestinal nematode Trichinella spiralis by intranasal administration of a 30-mer peptide antigen with cholera toxin B. Immunized mice developed antigen-specific serum immunoglobulin G1, intestinal immunoglobulin A, and a type 2-biased cytokine response. Intranasal immunization therefore generates the Th2-mediated responses required for immunity against intestinal parasites. (+info)
Trichinella spiralis-specific monoclonal antibodies and affinity-purified antigen-based diagnosis.
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Hybridomas secreting monoclonal antibodies (MAbs) to Trichinella spiralis were produced. Myeloma cells were fused with splenocytes of a mouse immunized with excretory-secretory (E-S) antigen of infective larvae. A large percentage of growing hybrids secreted antibodies cross-reactive to many of 23 heterologous parasites tested. Only 6 monoclones (designated 3F2, 5D1, 10F6, 11E4, 13D6 and 14D11) secreted MAbs specific to the E-S antigen and/or a crude extract (CE) of T. spiralis infective larvae. The 6 monoclones secreted IgM, IgG3, IgM, IgG3, IgG3 and IgG3, respectively. Clone 5D1 was selected to mass produce MAbs which were then coupled to CNBr-activated Sepharose CL-4B to prepare an affinity-purified antigen. Dot-blot ELISA with either purified antigen or CE was evaluated. There were 17 patients with acute trichinellosis and 76 individuals convalescing from T. spiralis infection (group 1). Controls were 170 patients with parasitic infections other than trichinellosis (group 2) and 35 healthy parasite-free controls (group 3). CE-ELISA was positive in all group 1 patients. However, sera from many group 2 patients also were reactive (opisthorchiasis-44.2%, schistosomiasis-44%, gnathostomiasis-30%, paragonimiasis-28.6%, taeniasis-27.3%, strongyloidiasis-23.1% and hookworm infections-20%). Affinity-purified antigen was 100% specific, all sera from group 2 and group 3 individuals tested negative. Although 74 of 76 patients (97.4%) with convalescing trichinellosis tested positive, sera from only 3 of 17 patients (17.6%) with acute T. spiralis were reactive. Thus, CE antigen is appropriate when sensitivity is needed, while purified antigen should be used when specificity is required. Dot-blot ELISA is easier to perform, more rapid and less expensive than indirect ELISA. Many samples can be assayed simultaneously, special equipment is not required, and results can be preserved for retrospective analysis. Dot-blot ELISA is therefore the method of choice for the rapid diagnosis of trichinellosis, particularly when more complex laboratory tests are unavailable. (+info)
Modulation of intestinal muscle contraction by interleukin-9 (IL-9) or IL-9 neutralization: correlation with worm expulsion in murine nematode infections.
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Immune responses associated with intestinal nematode infections are characterized by the activation of T-helper 2 (Th2) cells. Previous studies demonstrated that during Trichinella spiralis infection, Th2 cells contribute to the development of intestinal muscle hypercontractility and to worm eviction from the gut, in part through signal transducer and activator of transcription factor 6 (Stat6). Interleukin-9 (IL-9), a Th2-cell-derived cytokine, has pleiotropic activities on various cells that are not mediated through Stat6. In this study, we investigated the role of IL-9 in the generation of enteric muscle hypercontractility in mice infected with the intestinal parasite T. spiralis and the cecal parasite Trichuris muris. Treatment of mice with IL-9 enhanced infection-induced jejunal muscle hypercontractility and accelerated worm expulsion in T. spiralis infection. These effects were associated with an up-regulation of IL-4 and IL-13 production from in vitro-stimulated spleen cells. In addition, increases in the level of intestinal goblet cells and in the level of mouse mucosal mast cell protease 1 (MMCP-1) in serum were observed in infected mice following IL-9 administration. However, the neutralization of IL-9 by anti-IL-9 vaccination or by anti-IL-9 antibody had no significant effect on worm expulsion or muscle contraction in T. spiralis-infected mice. In contrast, the neutralization of IL-9 significantly attenuated T. muris infection-induced colonic muscle hypercontractility and inhibited worm expulsion. The attenuated expulsion of the parasite by IL-9 neutralization was not accompanied by changes in goblet cell hyperplasia or the MMCP-1 level. These findings suggest that IL-9 contributes to intestinal muscle function and to host protective immunity and that its importance and contribution may differ depending on the type of nematode infection. (+info)