In Vitro Parasiticidal Effects Of PIK-75 Against Echinococcus Granulosus Protoscoleces

Objective: The aims of this study were to investigate the role of the nuclear factor E2-related factor 2(Nrf2)signaling pathway in Echinococcus granulosus protoscoleces,to examine the effects of PIK-75 inhibition on Nrf2 activity,and to seek new drugs against cystic echinococcosis.Methods: Echinococcus granulosus protoscoleces were collected aseptically from cysts of infected sheep and In vitro cultured with RPMI 1640.we used the following final concentration of the drugs:0,0.4 ?M,0.8 ?M,1.2 ?M,1.6 ?M and 2.0 ?M).Nrf2 protein expression and localization in protoscoleces were examined via confocal immunofluorescence microscopy.The effects of PIK-75 on the levels and activity of heme oxygenase 1(HO-1)and NAD(P)H quinone oxidoreductase 1(NQO1)were characterized using ELISA.Evidence of parasite damage and death was observed by 0.1% eosin staining and scanning electron microscopy.Drawing dynamic curve after the experiment was independent repeated three times.In addition,,after 24 h and 48 h post-incubation,Caspase-3 activity treated with different concentrations of PIK-75 were detected using an assay kit.Results: Immunolocalization revealed that Nrf2 protein is primarily expressed in the cytoplasm,and its levels decrease during PIK-75-treatment in a dose-dependent manner.ELISA data showed that treating cultured protoscoleces with 0.8 ?M PIK-75 for either 2 or 5 days caused a significant decrease in both HO-1 and NQO-1 production in a time-and dose-dependent manner(P<0.05).It was observed that the morphological damage and reduced dynamic occurring in PIK-75-treated protoscoleces.Protoscoleces cultured with 2.0 ?M PIK-75 after 6 days has no longer viable.PIK-75 increased the mortality of protoscoleces in a dose-dependent manner.In addition,treatment with 0.8 ?M of PIK-75 reduced the number of viable protoscoleces within a period of 1 day and ~67.4 % after 3 days,as 32.1% of protoscoleces were still viable after 7 days of treatment.In the control group,no observed changes in morphology and dynamic was seen throughout the experimental period.SEM revealed many ultrastructural changes in PIK-75-treated protoscoleces,including numerous pits appearing at the tegument,rostellar deformation and irregular cavities,digitiform tegumental extensions in the tegument of the soma region,and protoscoleces contraction.Besides,All concentrations induced a significant increase of caspase-3activity at 24 h and 48 h compared with controls(P<0.05).Conclusion:1.PIK-75 efficiently reduced the expression of Nrf2 in E.granulosus protoscoleces and decreased the activities of Nrf2-dependent HO-1 and NQO-1.2.PIK-75 demonstrated clearly scolicidal activity against E.granulosus protoscoleces and could induce protoscoleces apoptosis.