IN-VITRO CHARACTERIZATION OF ACANTHAMOEBA-CASTELLANII CYTOPATHIC EFFECT

This study examined the mechanism of the cytopathic effect (CPE) of Ac anthamoeba castellanii on human target cells. Pathogenic Acanthamoeba trophozoites were incubated with human ocular melanoma (OCM1) cells fo r 30 min, 1 hr, and 3 hr. The amoebae were treated with a calcium iono phore (A23187), phorbol myristate ester (PMA), calcium channel blocker (Bepridil), cytochalasin D, and L-leucyl-L-leucine methyl ester (leu- leu-OMe). Cytolysis was quantified using a spectrophotometric assay. C ocultures of amoeba and cells were also observed by transmission elect ron microscopy at 1, 2, and 3 hr. Results show that trophozoites forme d pseudopodia that made intimate contact with the target cell membrane . Neither amebostomes nor phagocytosis was seen. The calcium ionophore A23187 increased the cytopathic effect of the trophozoites on the cul tured OCM1. In contrast, cytochalasin D, Bepridil, and PMA reduced the cytopathic effect. Leu-leu-OMe did not result in killing of Acanthamo eba trophozoites. The results suggest that the cytopathic effect of Ac anthamoeba trophozoites involves calcium channels and cytoskeletal ele ments. There was no evidence of trogocytosis or phagocytosis as someti mes occurs in cytolysis by other free-living amoeba. Although Acantham oeba-mediated CPE in some ways resembles CPE produced by cytotoxic lym phocytes, the mechanisms are not identical.