双氢青蒿素与甲硝唑体外抗阴道毛滴虫的电镜观察

In Vitro Anti-Trichomonas vaginalis Effects of a Mixture of Dihydroartemisinin and Metronidazole

目的观察双氢青蒿素与甲硝唑单用和联合用药体外对阴道毛滴虫超微结构的影响。方法在2.5×10~6个/ml滴虫培养液中加入两药合剂（双氢青蒿素0.5 mg/ml和甲硝唑0.002mg/ml）,同时设不加药的阴性对照组和单药对照组,即甲硝唑组（5 mg/ml）、双氢青蒿素组（1 mg/ml）,37℃培养3.5～5 h,扫描电镜和透射电镜观察阴道毛滴虫超微结构结果扫描电镜观察结果表明,仅用双氢青蒿素作用3.5 h,阴道毛滴虫细胞膜被破坏,部分表膜脱落;仅用甲硝唑作用3.5 h虫体表面结构无变化,作用5 h虫体表面出现许多小泡和小凹,但表膜未见破坏。两药联合作用3.5～4.2 h,阴道毛滴虫表面凹凸不平,出现皱褶、裂隙;细胞膜破损、脱落明显,表面粗糙,呈蜂窝状;细胞内含物从细胞膜裂口处溢出,虫体破裂,其内氢化酶体、核、轴柱和盾等裸露。透射电镜观察结果表明,仅用双氢青蒿素作用3.5 h,阴道毛滴虫膜系统破坏明显,破坏的滴虫细胞质从细胞膜破损处溢出;仅用甲硝唑作用3.5～5 h,细胞质破坏严重,细胞中可见许多空泡、裂隙和无结构区两药联合作用3.5～4.5 h,滴虫表膜破损,细胞内含物破坏严重,出现许多空泡、裂隙,内质网肿胀,氢化酶体膜破损、变形,细胞器大多消失;细胞核变形,核内出现裂隙,核膜受损,甚至消失。结论双氢青蒿素和甲硝唑对阴道毛滴虫的作用部位不同,两药联用可增进对虫体的破坏力。

Objective To observe the effect of a mixture of dihydroartemisinin and metronidazole on ultrastructure of Trichomonas vaginalis trophozoites in vitro for exploring trichomonacidal mechanism of the drug mixture.Methods The trophozoites were cultivated with liver extract solution medium that contained 2.5×10~6 parasites/ml.There were dihydroartemisinin 0.5 mg/ml and metronidazole 0.002 mg/ml in the experimental tubes of the drug mixture group.Groups of control （without drug）,dihydroartemisinin （1 mg/ml）and metronidazole （5 mg/ml）were established and performed in the same experimental conditions.The parasites were observed by scanning and transmission electron microscopes after having treated with the drugs at 37℃for 3.5-5 h.Results Under scanning electron microscope,the cell membrane of T.vaginalis treated only with dihydroartemisinin for 3.5 h was damaged,part of pellicle peeled off.Although the surface of the trophozoites treated only with metronidazole for 5 h showed many small bubbles and hollows,the cell membrane looked integral.However,surface of the parasite exposed to the drug mixture for 3.5-4.2 h showed deep folds and cracks,the cell membrane was damaged and even peeled off.When the cell ruptured,the nucleus,axostyle,pelta and hydrogenosomes were exposed,and the cytoplasm spilled out.Transmission electron microscopy showed that the membrane system of the trophozoites treated only with dihydroartemisinin for 3.5 h was damaged considerably.The cytoplasm of damaged parasite spilled out.The cytoplasm of the parasite treated only with metronidazole for 3.5-5 h was damaged seriously.Vacuoles and crevices were visible in the cytoplasm.The cell membrane and the content of the parasites treated with the drug mixture for 3.5-4.5 h were damaged seriously.There were some vacuoles and crevices, dilated endoplasmic reticulum,injured and deformed hydrogenosomes in the cytoplasm.The cell organelles mostly disappeared. Crevices also existed in the nucleus.The nuclear membrane fractured and even disappeared.Conclusion The acting targets of dihydroartemisinin and metronidazole to T.vaginalis trophozoite were different,and a combination of the two drugs shows stronger effect in killing the parasites.