抗癌药物索拉非尼体外抗多房棘球蚴的效果评价

中国寄生虫学与寄生虫病杂志 ›› 2017, Vol. 35 ›› Issue (5): 417-422.

抗癌药物索拉非尼体外抗多房棘球蚴的效果评价

党志胜¹, 奥祐三郎², 周晓农¹, 胡薇^1,³, 张颋¹, 陈军虎¹, 徐斌¹, 赵玉敏⁴, 肖宁^1,^*()

1 中国疾病预防控制中心寄生虫病预防控制所,世界卫生组织热带病合作中心,科技部国家级热带病国际联合研究中心,卫生部寄生虫病原与媒介生物学重点实验室,上海 200025
2 日本鸟取大学农学部兽医学院寄生虫学教研室,鸟取 680-8553,日本
3 复旦大学生命科学院微生物学与微生物工程系,上海 200433
4 桂林医学院生物技术学院生物医学工程系,桂林 541004

出版日期:2017-10-30 发布日期:2018-01-08
通讯作者: 肖宁
基金资助:
国家科技重大专项(No. 2012ZX10004-220);卫生部寄生虫与媒介生物学重点实验室开放课题(No. WSBKTKT201304);国家国际科技合作项目(No. 2014DFA31130)

In vitro inhibitive effect of the anticancer drug sorafenib on Echinococcus multilocularis larvae

Zhi-sheng DANG¹, Yuzaburo OKU², Xiao-nong ZHOU¹, Wei HU^1,³, Ting ZHANG¹, Jun-hu CHEN¹, Bin XU¹, Yu-min ZHAO⁴, Ning XIAO^1,^*()

1 National Institute of Parasitic Diseases, Chinese Center for Disease Control and Prevention; WHO Collaborating Centre for Tropical Diseases; National Center for International Research on Tropical Diseases, Ministry of Science and Technology; Key Laboratory of Parasite and Vector Biology, Ministry of Health, Shanghai 200025, China
2 Parasitology Laboratory, School of Veterinary Medicine, Faculty of Agriculture, Tottori University, Tottori, 680-8553, Japan
3 Department of Microbiology and Microbial Engineering, School of Life Sciences, Fudan University, Shanghai 200433, China
4 Department of Biomedical Engineering, College of Biotechnology, Guilin Medical University, Guilin 541004, China

Online:2017-10-30 Published:2018-01-08
Contact: Ning XIAO
Supported by:
Supported by the National Science and Technology Major Program(No. 2012ZX10004-220);Open Fund of Key Laboratory of Parasite and Vector Biology, Ministry of Health(No. WSBKTKT201304);International Science and Technology Cooperation Program of China (No. 2014DFA31130)

摘要/Abstract

摘要：

目的对抗癌药物索拉非尼体外抗多房棘球蚴的效果进行评价。方法从感染小鼠体内收集多房棘球蚴小囊泡,加入预先接种大鼠肝细胞的DMEM培养液（含10%胎牛血清、100 U/mL青霉素、100 mg/L链霉素）中,37 ℃、5% CO₂培养2~3个月后,选取直径2~4 mm的完整包囊用于药物效果评价实验。实验分阿苯达唑组（阳性对照）,阿苯达唑终浓度分别为10 μmol/L和30 μmol/L;索拉非尼组（实验组）,索拉非尼终浓度分别为10 μmol/L和30 μmol/L;DMSO组（溶剂对照组）,加DMSO（1︰9溶于PBS中）;PBS组（阴性对照）,仅加PBS。每孔取25~30个包囊,每个浓度设3个重复。药物处理48、72、96、120、144 和168 h后,光学显微镜和扫描电镜观察包囊形态结构变化,计算包囊抑制率,对其体外抗棘球蚴效果进行评价。结果 10、30 μmol/L索拉非尼处理48、72、96、120、144 和168 h后,包囊的抑制率分别为6.6%、42.4%、68.5%、77.4%、84.0% 、89.5%和7.1%、45.6%、70.9%、82.6%、84.0%、89.9%。10、30 μmol/L 阿苯达唑在每个时间点的包囊抑制率分别为2.6%、6.6%、21.4%、47.8%、59.9%、70.6%和3.8%、12.7%、27.0%、51.4%、54.0%、73.0%。DMSO和PBS对包囊没有明显的抑制效果。索拉非尼的抑制效果均高于阿苯达唑,从处理后72 h起,索拉非尼和阿苯达唑的抑制率差异有统计学意义（P < 0.05 ）。光学显微镜观察表明,索拉非尼处理后,包囊向内部陷塌,形成“日全食”状;扫描电镜观察发现,索拉非尼处理后,包囊内部生发层细胞脱落,完全失去活性。结论索拉非尼显示出较高的包囊抑制效果,是棘球蚴病治疗的潜在药物。

关键词: 多房棘球蚴病, 生发层, 索拉非尼

Abstract:

Objective To evaluate the effect of an anti-cancer drug sorafenib against alveolar echinococcosis (AE) in vitro. Methods Metacestode vesicles were obtained from infected mice and cultured with rat hepatocytes in Dulbecco’s modified Eagle’s medium (DMEM) containing 10% fetal calf serum (FCS), 100 U/ml penicillin G and 100 mg/L streptomycin sulphate at 37 ℃ with 5% CO₂. Morphologically intact metacestode vesicles with a minimal diameter of 2-4 mm were manually picked up from the co-culture and washed with PBS. They were then allocated (n = 25-30 cysts/well) into four groups: albendazole-treated group (positive group), sorafenib-treated group (experimental group), DMSO-treated group (solvent group), and PBS group (negative control), and incubated in 24-well plates in the presence of DMEM without phenol red. Albendazole and sorafenib were dissolved in DMSO (1 ∶ 9 dissolved in PBS) and used at 10 μmol/L and 30 μmol/L respectively. The experiment was performed in triplicates per sample. The integrity of the cysts was assessed with light microscopy and scanning electron microscopy (SEM). The inhibitory effects of albendazole and sorafenib on cysts were evaluated at 48, 72, 96, 120, 144 and 168 h post-treatment. Results The cyst inhibitory rate of sorafenib was 6.6%, 42.4%, 68.5%, 77.4%, 84.0% and 89.5% at 10 μmol/L, and 7.1%, 45.6%, 70.9%, 82.6%, 84.0% and 89.9% at 30 μmol/L, respectively, at 48, 72, 96, 120, 144 and 168 h post-treatment. While the cyst inhibitory rate of albendazole was 2.6%, 6.6%, 21.4%, 47.8%, 59.9% and 70.6% at 10 μmol/L, and 3.8%, 12.7%, 27.0%, 51.4%, 54.0% and 73.0% at 30 μmol/L, respectively. DMSO and PBS showed less toxic to the cysts. Sorafenib exerted a stronger inhibitory effect than albendazole at the corresponding time points, and the difference began to be statistically significant from 72 h post-treatment. Light microscopy and SEM showed observable collapse of the germinal layer and overall damage of germinal cells in the experiment groups. Conclusion Sorafenib shows a significant inhibitory effect on Echinococcus cysts, suggesting a potential drug candidate for echinococcosis treatment.

Key words: Alveolar echinococcosis, Germinal layer, Sorafenib

中图分类号:

R383.33

党志胜, 奥祐三郎, 周晓农, 胡薇, 张颋, 陈军虎, 徐斌, 赵玉敏, 肖宁. 抗癌药物索拉非尼体外抗多房棘球蚴的效果评价[J]. 中国寄生虫学与寄生虫病杂志, 2017, 35(5): 417-422.

Zhi-sheng DANG, Yuzaburo OKU, Xiao-nong ZHOU, Wei HU, Ting ZHANG, Jun-hu CHEN, Bin XU, Yu-min ZHAO, Ning XIAO. In vitro inhibitive effect of the anticancer drug sorafenib on Echinococcus multilocularis larvae[J]. Chinese Journal of Parasitology and Parasitic Diseases, 2017, 35(5): 417-422.

图/表 5

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