Abstract:

Objective To evaluate the effects of tacrine on Echinococcus multilocularis in vitro and against E. multilocularis infection in mice. Methods Tacrine was added at 100.00, 50.00, 25.00, 12.50 and 6.25 μmol/L to each well of a 96-well plate containing 50-200 E. multilocularis protoscoleces. The methylene blue exclusion method was used to determine the viabilities of protoscoleces on days 1, 3, 5 and 7 after the addition. The cytotoxicity of tacrine on three noncarcinogenic cell types(L929 cells, HK-2 cells and Chang liver) and three tumor cell types(A172, A2058 and HCT-8 cells) was tested using the Cell Counting Kit-8（CCK-8） method. In another experiment, 40 BALB/c mice with 6 months of E. multilocularis infection were randomly divided into 4 groups: 30 mg/kg tacrine group, 15 mg/kg tacrine group, 25 mg/kg mebendazole group, and 1% tragacanth group as a control. All the mice received intragastric administration for 28 days. After 14 days of withdrawal, the mice were sacrificed, and cysts in the peritoneal cavity were isolated and weighed to calculate the reduction rate of cyst weight. Data were analyzed with the nonparametric test in SPSS 17.0.  Results In vitro, tacrine showed an obvious dose-dependent effect on E. multilocularis protoscoleces. The viability of protoscoleces decreased with increased drug concentration and incubation time. After 3 days of exposure to 100 μmol/L tacrine, all the protoscoleces were dead, with dramatic morphological alterations. Intact parasites were hardly seen. After 7 days of tarcine treatment, the percentage dead of protoscoleces was (100 ± 0)%, (91.2 ± 2.5)%, (80.3 ± 5.1)%, (71.6 ± 2.4)% and (51.7 ± 2.9)% in the 100.00, 50.00, 25.00, 12.50, and 6.25 μmol/L groups, respectively. The normal host cell types were less affected by tarcine at 250.0 μmol/L, as demonstrated by the (27.6 ± 4.7)%, (29.6 ± 3.9)% and (26.9 ± 2.1)% inhibitory rate for L929 cells, HK-2 cells, and Chang liver cells, respectively. In contrast, tarcine exerted high cytotoxicity to tumor cells, as demonstrated by Tox50 values of (178.2 ± 3.2), (133.2 ± 5.2) and (128.8 ± 4.0) μmol/L for A172 cells, A2058 cells and HCT-8 cells, respectively. The reduction rate of cyst weight was -3.4%, 9.4%, and 38.2% in 30 mg/kg tacrine, 15 mg/kg tacrine and 25 mg/kg mebendazole groups, respectively. However, none was significantly different from the 1% tragacanthin group(P > 0.05). During the experimental period, 3, 6, 2 and 5 mice died in the four groups, and 25 mg/kg mebendazole and 15 mg/kg tacrine increased the survival rate compared with other groups.  Conclusion Tacrine can inhibit the activities of E. multilocularis protocoleces in vitro and increase the survival rate of mice infected with E. multilocularis.

Key words:  Tacrine, Echinococcus multilocularis, Protoscolex, Cytotoxicity