Induction of gelatinase in eosinophilic meningitis of BALB/c-strain mice was caused by Angiostrongylus cantonensis. Time-course studies showed that the molecular weight of 94-kDa gelatinase was detected at day 10 post-inoculation (PI), and reached a high intensity from days 15 to 25 PI. The 94-kDa gelatinase activity was clearly inhibited by EDTA and 1,10-phenanthroline, but not by leupeptin and phenylmethanesulphonyl fluoride. When immunoblots were performed using specific antiserums against the 94-kDa gelatinase B (matrix metalloproteinase-9; MMP-9) with cerebrospinal fluid (CSF), the 94-kDa immunopositive band was MMP-9. Immunohistochemistry studies demonstrated MMP-9 localisation within eosinophils and macrophages. The increased MMP-9 activity was closely associated with the rapid rise of CSF eosinophils, and the inflammatory reaction of the subarachnoid space. In contrast to changes in MMP-9, MMP-2 activity was constitutive and unaffected in this parasitic meningitis. These results show that MMP-9 was associated with eosinophilic meningitis, and that the enzyme may be a useful marker for angiostrongyliasis meningitis.

Matrix metalloproteinase 9 (MMP-9) is involved in the pathogenesis of parasitic meningitis caused by the nematode Angiostrongylus cantonensis. The present study evaluated the efficacy of albendazole therapy in BALB/c mice infected with the third stage larvae of this nematode. Albendazole showed a pronounced larvicidal activity. Eosinophil numbers significantly increased in infected mice but decreased upon administration of albendazole. Densitometric scanning indicated that albendazole reduced gelatinolytic activity detected by gelatin-substrate zymography. In the cerebrospinal fluid, albendazole reduced the lytic area intensity of the 94 kDa MMP-9 band by 46.5% within 7 days, and by 51.5% by day 14. Examination of brain tissue revealed a similar pattern of decrease (48.6% by day 7, and 53.9% by day 14). Albendazole may thus be an effective compound for the treatment of angiostrongyliasis through its larvicidal activity and facilitation of an improved inflammatory response via the reduction of MMP-9 activity.

Ingestion of the larval nematode Angiostrongylus cantonensis can cause the human eosinophilic meningitis known as angiostrongyliasis. Analysis of the extracts and excretory-secretory (ES) products of A. cantonensis larvae and adult stages on gelatin substrate zymography demonstrated the presence of distinct gelatinolytic enzymes. In worm extracts, inhibitor studies showed that the metalloproteinases revealed in L(1) (23 kDa), L(3) (66, 42 and 30 kDa), young adult worm (72 and 94 kDa) and adult worm (72 and 94 kDa). In ES products, the L(1) revealed one low (42 kDa) and two high (105 and 94 kDa) molecular weight proteolytic bands that degraded gelatin in substrate gels. The L(3) revealed three low (66, 50, and 30 kDa) and one high (105 kDa) molecular weight proteolytic bands. Inhibitor studies confirmed that the 105 and 94 proteolytic bands of the L(1), and the 50 and 30 kDa proteolytic bands of the L(3) classification were metalloproteinases. These metalloproteinases secreted in the infective larvae may be associated with the parasite dissemination or pathogenesis.

The histomorphology of granuloma formation and gelatinase production were investigated in the brains, hearts, lungs and livers of Sprague-Dawley rats infected with Angiostrongylus cantonensis. The relationships between two gelatinases and granulomatous fibrosis were explored, following infection of each rat with 60 infective larvae of the nematode. Worm recovery from the brain was maximal on day 15 post-inoculation whereas peak recovery from the lungs was maximal 75 days later, on day 90. The granulomatous reactions and fibrosis were marked in the lungs but only mild, if present at all, in the brain, heart and liver. Gelatin zymography revealed that matrix metalloproteinase2 (MMP-2) was present, at all time-points, in the heart and lungs, although only in the lungs was there partial conversion of the 72-kDa pro-enzyme to the 64-kDa active form during granulomatous fibrosis. The activity of the MMP-9 pro-enzyme was significantly higher at the time-points when granuloma formation was observed than at other times. Immuno-histochemistry revealed MMP-2 and MMP-9 within the lung granulomas, around infiltrating leucocytes and the epithelial cells of the alveoli. As the granulomatous fibrosis appeared to be strongly associated with MMP-2 and MMP-9, these enzymes may be useful markers in the lungs of rats infected with A. cantonensis.

Angiostrongylus cantonensis, the rat lungworm, is the principal cause of human eosinophilic meningitis or meningoencephalitis world-wide. In the present study, the efficacies of early-stage treatment with the Chinese herbal medicine long-dan-xie-gan-tan (LDXGT) and albendazole, used alone or in combination, were evaluated in BALB/c mice with A. cantonensis-induced dysfunction of the blood-central-nervous-system barrier and eosinophilic meningo-encephalitis. Indicators of the therapeutic effect included worm recovery, histopathological scores for the meningitis, assays of tissue-type plasminogen activator (PA), urokinase-type PA and matrix metalloproteinase-9 (MMP-9) in the brain, the ratio between albumin concentrations in the cerebrospinal fluid (CSF) and serum, and counts of eosinophils in the CSF. Combined treatment with albendazole and LDXGT gave better results than monotherapy based on either drug, apparently inhibiting eosinophilic meningitis via antagonists of the PA/MMP-9 system. LDXGT may have a therapeutic role in reducing inflammatory reaction in the subarachnoid space. Monotherapy with such an anti-inflammatory drug may relieve the symptoms of mild infection and the host's immune responses to A. cantonensis larvae. In severe infection, however, co-therapy with an anthelmintic (to kill the larvae) and an anti-inflammatory agent (to provide symptomatic relief) is probably a better approach. The therapeutic strategy should be tailored to the severity of the illness and the numbers of eosinophils in the CSF.