胡桃楸提取物抗弓形虫作用的体外实验和超微研究

In vitro effect and ultrastructural study of the extract of Juglans mandshurica maxim on Toxoplasma gondii

目的了解胡桃楸提取物(JMME)在体外杀灭弓形虫速殖子的作用效果及其机制。方法收集感染弓形虫小鼠腹水并稀释至含虫1×107个/ml,加入胡桃楸提取物使其终浓度为60μg/ml、30μg/ml、15μg/ml、7.5μg/ml、3.25μg/ml、1.625μg/ml;乙酰螺旋霉素为阳性对照组,终浓度25μg/ml,同时设培养液阴性对照组。用MTT法测定胡桃楸提取物及其含药血清对虫体增殖情况的影响。应用透射电镜观察药物作用24h虫体超微结构的变化并拍照。结果胡桃楸提取物可明显抑制弓形虫增殖,且具有剂量依赖性,浓度大于30μg/ml的抑虫效果优于乙酰螺旋霉素组(P<0.05)。胡桃楸提取物含药血清对弓形虫具有明显的增殖抑制作用,且随含药血清浓度的增加作用效果增强,JMME浓度为0.278g/(kg.d)的含药血清抑虫效果优于SM组(P<0.05)。透射电镜下可观察到虫体电子密度增高、大量空泡形成、细胞器被破坏、细胞膜破裂、内容物溢出。结论胡桃楸提取物对弓形虫具有明显的增殖抑制和破坏作用。

Objective To investigate the in vitro killing effect of Juglans mandshurica maxim extract （JMME） on Toxoplasma tachyzoites and its mechanism. Methods The peritoneal fluid of Toxoplasma gondii （ T. gondii） infected mice was collected. The number of the tachyzoite was adjusted to 1 × 10^7/ml. JMME was added to diluted tachyzoite - containing peritoneal fluid with the final concentration of 60 μg/ml, 30 μg/ ml, 15 μg/ml, 7.5 μg/ml, 3.25 μg/ml and 1. 625μg/ml respectively. 25 μg/ml of acetylspiramycin was added as a positive control and culture medium as negative control. MTT assay was used for the determination of the proliferation of Toxoplasma. The changes of the uhrastructure of the parasite 24h after JMME adding was observed with transmission electron microscopy. Results JMME significantly inhibited the proliferation of T. gondii and the inhibition was dose - dependent. When JMME concentration greater than 30μg/ml, the effect was better than that in acetylspiramycin group （ P 〈 0.05 ）. Tile JMME -containing serum also showed significant proliferation inhibition effect on T. gondii, and again the inhibition was dose- dependent. 0. 278 g/（kgod） of JMME -containing serum exhibited better inhibition effect than SM （ P 〈 0.05 ）. The observation of T. gondii under transmission electron microscopy showed increased electron density, vacuole formation, destroyed organelle, lnembranolysis and o- verflow of parasite contents. Conclusion JMME showes obvious proliferation inhibition and destruction effect on T. gondii tachyzoites.