Expression of in vitro virulence by Entamoeba histolytica: effect of calmodulin inhibitors.

The possible role of calmodulin (CaM) in functions such as cytotoxicity, phagocytosis, and amoebic growth was studied. These are important factors for the in vitro pathogenicity of Entamoeba histolytica HM1 strain. The CaM inhibitors trifluoperazine (TFP), N-(6-aminohexyl)-5-chloro-1-naphthalene-sulfonamide (W-7) and N-(6-aminohexyl)-naphthalenesulfonamide (W-5) were used for these purposes. Cytotoxicity was determined as the ability of amoebae to kill and/or lyse nucleated mouse spleen cells. Phagocytosis by amoebae was assessed by calculating the number of endocytosed human red blood cells. Cytotoxicity by E. histolytica showed a decrease dependent upon the concentration of W-7 and TFP, whereas W-5 did not show a significant inhibiting effect. Phagocytosis was roughly sensitive to W-7; TFP and W-5 did not have any significant effect. Amoebic growth was sensitive to TFP and W-7 CaM inhibitors. These results suggest that CaM participates in the expression of in vitro cytotoxicity of E. histolytica.

Possible role of calmodulin in the secretion of Entamoeba histolytica electron-dense granules containing collagenase.

Entamoeba histolytica cells secrete electron-dense granules (EDGs) that have collagenase activity. To study the possible involvement of calmodulin (CaM) on EDG secretion, the effect of several CaM antagonists (TFP, R24571, W-7, W-5, dibucaine and DL-propranolol) was tested on this cellular function. Except for W-5 and dibucaine, the rest of these compounds inhibited EDG secretion. Transmission electron microscopy of collagen-activated trophozoites showed numerous EDGs located in or near the surface membrane. In contrast, trophozoites incubated with TFP showed no EDGs. Protein kinase C inhibitors (H-7, ML-9) had no effect on EDG secretion, suggesting that CaM antagonists acted by selectively inhibiting CaM. These results suggest that a CaM-dependent process is involved in EDG secretion.