Matrix metalloproteinase-2 and matrix metalloproteinase-9 in mice with ocular toxocariasis.

In ocular toxocariasis, Toxocara canis-induced inflammatory reaction can lead to eye destruction and granuloma, which is formed by immune cell infiltration and concurrent extensive remodeling tissue. Herein, the histomorphology of granuloma and proteinase production in the eye of T. canis-infected BALB/c mice were investigated. Pathological effects substantially increased after the infection culminated in a severe leukocyte infiltration and granuloma formation from days 4 to 56 post-inoculation. The matrix metalloproteinase (MMP)-2 and MMP-9 activities remarkably increased, compared with those of uninfected control, by gelatin zymography and Western blot analysis in ocular toxocariasis. Granuloma formation had a remarkably positive correlation with MMP-2 and MMP-9 levels. We suggested that T. canis larvae and leukocytes infiltrated from blood vessel both migrated into corpus adiposum orbitae. Activated leukocytes secreted MMP-2 and MMP-9, leading to fibronectin degradation. The imbalance of MMP-2/TIMP-2 and MMP-9/TIMP-1 may play a role in inflammatory cell infiltration and extracellular matrix degradation, forming granuloma, in ophthalmological pathogenesis of T. canis infection.

Changes to plasminogen activators and matrix metalloproteinase-9 in dogs with toxocarosis.

The relationship between proteolytic enzymes and hematological response to infection was studied in five 1-month-old dogs inoculated experimentally with 2000 eggs of Toxocara canis. Moderate leukocytosis and marked eosinophilia occurred 14 days post-infection with T. canis. Plasminogen activators (PAs) and matrix metalloproteinase-9 (MMP-9) activity in serum was significantly different in dogs infected with T. canis, compared with controls. Urokinase-type PA (uPA) activity was positively correlated with eosinophilia, and tissue-type PA (tPA) and MMP-9 activity was negatively correlated with eosinophilia. However, there was no correlation between inflammation and MMP-2. The use of uPA, tPA or MMP-9 proteolytic enzymes as laboratory reference markers for toxocarosis requires further study.

Induction of matrix metalloproteinase-9 in mice during Toxocara canis larvae migration.

The relationships between inflammation in organs with Toxocara canis larval migration and matrix metalloproteinase-9 (MMP-9) were investigated following the infection of mice with 1,000 infective eggs. Gelatinase activity was defined by gelatin zymography, optimum pH, inhibitor specificity and Western blot analysis. MMP-9 activity was present in the lungs, liver, muscles, and brain during T. canis larval migration. This enzyme had a molecular weight of about 94 kDa and showed maximum activity in the pH range of 6-8. The increased MMP-9 proteinases coincided with larval recovery and the degree of inflammation among the four organs. These results suggest that MMP-9 may be associated with the inflammatory reaction to larval toxocariasis during early migration, and may therefore be a useful marker during T. canis larvae migration.

Diagnosis of parasitic zoonoses by immunoenzymatic assays--analysis of cross-reactivity among the excretory/secretory antigens of Fasciola hepatica, Toxocara canis, and Ascaris suum.

Several parasitic infections such fasciolosis, toxocariosis or ascariosis are important zoonoses. During the infection with Fasciola hepatica, Toxocara canis and Ascaris suum, an important intraorganic phase in their hosts takes place, releasing antigens responsible for a humoral immune response, which enables the diagnosis of that parasitosis. A study to identify the existence of cross-reactivity among the excretory/ secretory antigens of F. hepatica, T. canis and A. suum was developed. One group of Sprague-Dawley rats was infected with 20 metacercariae of F. hepatica and another group remained uninfected as control. By means of an Indirect-ELISA, the rat humoral immune response (IgG and IgM) against the excretory/secretory antigens of F. hepatica was analysed and measured for cross reactivity with T. canis and A. suum. IgM cross-reaction was mainly observed in the first 10 weeks post-infection. IgG cross-reaction was observed throughout the study, and was maximal at the 2-3 weeks and 3-6 weeks post-infection, which corresponds to the intraorganic migratory phase of these parasites. The western-blot showed that the rat IgG recognised three proteins of 190, 160 and 33 kDa in the antigens from F. hepatica, T. canis and A. suum. The existence of cross-reactivity among these antigens seems to demonstrate also the presence of structural similarities, such as tegumental proteins. These results should be consider when immunoassay probes are used in the diagnosis of parasitic infections.

Eosinophil peroxidase levels in hearts and lungs of mice infected with Toxocara canis.

Toxocara canis infection of abnormal hosts results in a condition in which infective larvae migrate through the soft tissues of the body, exclusive of the skin. This condition is known as visceral larva migrans (VLM) and causes a syndrome characterized by hepatosplenomegaly, hyperglobulinemia, hypereosinophilia, and transient pulmonary infiltrates. Because of the known association between hypereosinophilia and eosinophilic heart disease, we have been studying the hearts of mice infected with T. canis for evidence of myocardial damage and have previously described a severe eosinophilic myocarditis that leads to a marked myocardial fibrosis. We have measured eosinophil peroxidase (EPO) levels (a marker enzyme for specific granules of eosinophils) in homogenized lungs, homogenized hearts, and eosinophils recovered from the lungs of mice infected with T. canis over a 6-wk period. A marked accumulation of EPO was observed in the lungs of infected mice from day 14 postinfection (PI) to at least 6 wk of infection. Most of the EPO was associated with eosinophils that comprise the bulk of the pulmonary infiltrates associated with the VLM syndrome. However, following bronchoalveolar lavage, cytochemical localization of EPO activity in lungs from infected mice suggested that eosinophil degranulation had resulted in this marker enzyme being deposited within the pulmonary parenchyma. Peak levels of EPO were found in the myocardium by day 14 PI and declined over the 6-wk period. These levels equaled about 1/3 of the levels seen in the lungs of the same mice. These studies suggest that in mice infected with T. canis, the presence of increased numbers of eosinophils may lead to marked peroxidatic cardiopulmonary damage.

A relation between antibody response and acetylcholinesterase activity in dogs and rabbits infected with Toxocara canis.

A relation between IgG antibodies and acetylcholinesterase (ACHE) activities was observed in the sera of dogs and rabbits infected with Toxocara canis (T. canis). No antibodies were found in the sera of dogs' fetuses and newborns, more over their ACHE levels were lower than the ones registered in the sera of adult dogs. The ACHE levels of both, the mother dog and the newborn puppies, reached the same level 3 months after delivery. Similar results were obtained in the sera of puppies and adult dogs. In the sera of two rabbits experimentally infected with T. canis, the IgG antibodies were sharply raised 2 weeks after infection and the ACHE levels gradually decreased during the course of this observation. The serum glutamic pyruvic transaminase further tested in these sera gradually increased. From the results obtained, the liver disfunction could be recognized in rabbits but not in dogs, and no relation between antibody and ACHE activities was found neither in the sera of dogs nor rabbits.