摘要
目的研究大肠杆菌胞外囊泡(E.coli-OMVs)对人神经母细胞瘤SK-N-SH细胞增殖、凋亡和迁移的影响。方法采用差速离心法提取大肠杆菌胞外囊泡,观察囊泡形态并测定囊泡粒径范围。随机分为4组:正常对照组、胞外囊泡低剂量组(100μg/m L E.coli-OMVs)、胞外囊泡中剂量组(500μg/m L E.coli-OMVs)和胞外囊泡高剂量组(1000μg/m L E.coli-OMVs)。MTT检测各组细胞增殖能力;TUNEL和Western blot分别检测各组细胞凋亡情况及凋亡相关蛋白caspase3的表达;Transwell检测各组细胞迁移能力。结果透射电镜下E.coli-OMVs呈双层膜、中空、圆形或椭圆形,其内含低密度或高密度物质,具有明显的异质性,粒径分析结果表明其直径分布在30~450 nm。与对照组相比,经E.coli-OMVs干预后,SK-N-SH细胞增殖能力显著降低;细胞凋亡增加,caspase3蛋白表达水平明显上调;而迁移能力显著降低(P<0.05)。结论 E.coli-OMVs对SK-N-SH细胞具有直接杀伤作用,有望成为难治性神经母细胞瘤的治疗药物。
Objective To investigate the effect of outer membrane vesicles derived from Escherichia coli on proliferation,apoptosis and migration of human neuroblastoma SK-N-SH cells in vitro. Methods The outer membrane vesicles（OMVs） were obtained from wild-type Escherichia coli with ultracentrifugation method, and the morphology of the OMVs was observed by transmission electron microscopy and the vesicle diameter was determined using MALVERN ZEN3690. Human neuroblastoma SK-N-SH cells were treated with the OMVs at low（100 μg/m L）, moderate（500 μg/m L） and high（1000 μg/m L）doses, and 24, 48 and 72 h later, the cell proliferation activity was detected with MTT assay. The expressions of apoptosisrelated marker caspase-3 was detected using Western blotting, and TUNEL assay was performed to detect the cell apoptosis.The migration capacity of SK-N-SH cells was evaluated using Transwell migration assay. Results The isolated OMVs showed a circular or elliptical hollow structure with double-layer membrane and a diameter range of 30-450 nm. Compared with the control cells, SK-N-SH cells treated with the OMVs showed significantly lowered cell proliferation capacity with enhanced expression of caspase-3. Treatment of the cells with the OMVs resulted in increased cell apoptosis and significantly lowered migration capacity（P〈0.05）. Conclusion The OMVs derived from Escherichia coli can produce cytotoxicity against SK-N-SH cells and might serve as a therapeutic agent for refractory neuroblastoma.
作者
唐波
何大维
李典
郭文浩
张丹
魏光辉
TANG Bo;HE Dawei;LI Dian;GUO Wenhao;ZHANG Dan;WEI Guanghui(Department of Urinary Surgery, Chongqing Key Laboratory of Children Urogenital Development and Tissue Engineering, Children＇s Hospital of Chongqing Medical University, Ministry of Education Key Laboratory of Child Development and Disorders, China International Science and Technology Cooperation base of Child development and Critical Disorders, Chongqing Key Laboratory of Pediatrics, Chongqing 400014, China)
出处
《南方医科大学学报》
CAS
CSCD
北大核心
2018年第3期334-339,共6页
Journal of Southern Medical University
基金
国家自然科学基金(81370701)
重庆市社会事业与民生保障科技创新专项(cstc2017shms A130103)~~
关键词
SK-N-SH细胞
大肠杆菌胞外囊泡
增殖
凋亡
迁移
neuroblastoma
SK-N-SH cells
outer membrane vesicles
Escherichia coli
cell proliferation
apoptosis
migration
