Evaluation of the immunoreactivity of nerve growth factor and tropomyosin receptor kinase A in the esophagus of noninfected and infected individuals with Trypanosoma cruzi.

Megaesophagus is one of the major manifestations of the chronic phase of Chagas disease. Its primary symptom is generally dysphagia due to disturbance in the lower esophageal sphincter. Microscopically, the affected organ presents denervation, which has been considered as consequence of an inflammatory process that begins at the acute phase and persists in the chronic phase. Inflammatory infiltrates are composed of lymphocytes, macrophages, natural killer cells, mast cells, and eosinophils. In this study, we evaluated the immunoreactivity of nerve growth factor (NGF), and of its receptor tropomyosin receptor kinase A (TrkA), molecules that are well known for having a relevant role in neuroimmune communication in the gastrointestinal tract. Esophageal samples obtained via autopsy or surgery procedures from six noninfected individuals, six infected individuals without megaesophagus, and six infected individuals with megaesophagus were analyzed. Infected individuals without megaesophagus presented increased numbers of NGF immunoreactive (IR) mast cells and increased areas of TrkA-IR epithelial cells and inner muscle cells. Infected individuals with megaesophagus showed increased numbers of NGF-IR eosinophils and mast cells, TrkA-IR eosinophils and mast cells, increased area of NGF-IR epithelial cells, and increased areas of TrkA-IR epithelials cells and inner muscle cells. The data presented here point to the participation of NGF and its TrkA receptor in the pathology of chagasic megaesophagus.

Mast cell-nerve interaction in the colon of Trypanosoma cruzi-infected individuals with chagasic megacolon.

Chagas disease is an infection caused by the parasite Trypanosoma cruzi that affects millions of people worldwide and is endemic in Latin America. Megacolon is the most frequent complication of the digestive chronic form and happens due to lesions of the enteric nervous system. The neuronal lesions seem to initiate in the acute phase and persist during the chronic phase, albeit the mechanisms involved in this process are still debated. Among the cells of the immune system possibly involved in this pathological process is the mast cell (MC) due to its well-known role in the bi-directional communication between the immune and nervous systems. Using ultrastructural analysis, we found an increased number of degranulated MCs in close proximity to nerve fibers in infected patients when compared with uninfected controls. We also immunostained MCs for the two pro-inflammatory molecules tryptase and chymase, the first being also important in neuronal death. The number of MCs immunostained for tryptase or chymase was increased in patients with megacolon, whereas increased tryptase staining was additionally observed in patients without megacolon. Moreover, we detected the expression of the tryptase receptor PAR2 in neurons of the enteric nervous system, which correlated to the tryptase staining results. Altogether, the data presented herein point to the participation of MCs on the denervation process that occurs in the development of T. cruzi-induced megacolon.

Mast cells in the colon of Trypanosoma cruzi-infected patients: are they involved in the recruitment, survival and/or activation of eosinophils?

Megacolon is frequently observed in patients who develop the digestive form of Chagas disease. It is characterized by dilation of the rectum-sigmoid portion and thickening of the colon wall. Microscopically, the affected organ presents denervation, which has been considered as consequence of an inflammatory process that begins at the acute phase and persists in the chronic phase of infection. Inflammatory infiltrates are composed of lymphocytes, macrophages, natural killer cells, mast cells, and eosinophils. In this study, we hypothesized that mast cells producing tryptase could influence the migration and the activation of eosinophils at the site, thereby contributing to the immunopathology of the chronic phase. We seek evidence of interactions between mast cells and eosinophils through (1) evaluation of eosinophils, regarding the expression of PAR2, a tryptase receptor; (2) correlation analysis between densities of mast cells and eosinophils; and (3) ultrastructural studies. The electron microscopy studies revealed signs of activation of mast cells and eosinophils, as well as physical interaction between these cells. Immunohistochemistry and correlation analyses point to the participation of tryptase immunoreactive mast cells in the migration and/or survival of eosinophils at the affected organ.

Neuroimmunopathology of Trypanosoma cruzi-induced megaoesophagus: Is there a role for mast cell proteases?

Tryptase and chymase are mast cell (MC)-specific proteases, which influence in the activation of inflammatory cells. In this study, we quantified tryptase- or chymase-expressing MCs in the oesophaguses of Chagas patients, and searched for a correlation between those data with area of nerve fibres that expressed either PGP9.5 (pan-marker) or vasoactive intestinal polypeptide (VIP), which is a neuromediator that has anti-inflammatory activity. Samples from the oesophaguses of 14 individuals Trypanosoma cruzi-infected and from six uninfected individuals were analysed by immunohistochemistry. It was demonstrated that the number of tryptase-IR MCs in infected individuals increased when compared with controls, regardless of whether the individuals had megaoesophagus, whereas the number of chymase-IR MCs increased only in infected individuals without megaoesophagus. Negative correlations were observed between tryptase-IR MCs and the density of nerve fibres that expressed VIP or PGP 9.5-IR. The participation of chymase and tryptase in this type of immunopathology is discussed.

An imbalance between substance P and vasoactive intestinal polypeptide might contribute to the immunopathology of megaesophagus after Trypanosoma cruzi infection.

Megaesophagus is one of the major causes of morbidity in chronic Chagas disease, and extensive denervation, associated with an inflammatory process, is recognized as the key factor for alterations in motility and disease development. Here, we analyzed esophagus samples from necropsied, infected individuals--6 cases with megaesophagus and 6 cases without megaesophagus--for the relative areas of expression of 2 neuromediators, substance P and vasoactive intestinal peptide, which are known to activate or inhibit, respectively, local immune cells. Samples from 6 noninfected individuals were used as controls. Esophageal sections were immunohistochemically stained for protein gene product 9.5, vasoactive intestinal peptide, and substance P, and the relative areas of expression of the latter 2 were calculated. Morphometric analyses revealed increased substance P and decreased vasoactive intestinal peptide relative areas in esophageal sections from patients with megaesophagus. Furthermore, in the group of patients without megaesophagus, the loss of vasoactive intestinal peptide positively correlated with the denervation process. We suggest that an imbalance between vasoactive intestinal peptide and substance P production results in the reestablishment and maintenance of the inflammatory process, leading to denervation and, consequently, promoting the development of megaesophagus.

Characterization of enteroglial cells and denervation process in chagasic patients with and without megaesophagus.

Chagas disease is caused by infestation with the parasite Trypanosoma cruzi, and some patients who are serologically positive develop chronic megaesophagus, whereas others are symptom-free. Gastrointestinal form of Chagas disease involves an inflammatory invasion of the enteric plexuses and degeneration of enteric neurons and previous works related that enteroglial cells would be involved in enteric inflammatory responses. Because of this, the aims of this study were to determine the relation of enteroglial cells with the denervation process in chagasic patients with and without megaesophagus and seronegative individuals. Our results indicated that the innervation of the esophageal muscle was substantially reduced in patients with megaesophagus, but asymptomatic seropositive subjects were not different to seronegative controls. Besides, patients with megaesophagus had significant decreased of enteroglial cells labeled with S-100 and glial fibrillary acidic protein, whereas patients without megaesophagus presented an increased of both labels. We believe that enteroglial cells would operate a mechanism of defense in the enteric nervous system against the Trypanosoma cruzi infection, which could prevent the organ denervation and preserve the esophagus function.