大肠埃希菌外膜囊泡对乳腺癌细胞增殖及荷瘤小鼠肿瘤生长的影响  

Effects of Escherichia coli outer membrane vesicles on proliferation of breast cancer cells and tumor growth of tumor-bearing mice

作　　者：王斓茜 马官荣 姜咏竹 常秀林 方廖琼[1] 白晋[1,2] WANG Lanxi;MA Guanrong;JIANG Yongzhu;CHANG Xiulin;FANG Liaoqiong;BAI Jin(State Key Laboratory of Ultrasound in Medicine and Engineering,College of Biomedical Engineering,Chongqing Medical University,Chongqing 400016,China;Chongqing Key Laboratory of Biomedical Engineering,Chongqing Medical University,Chongqing 400016,China)

机构地区：[1]重庆医科大学生物医学工程学院,超声医学工程国家重点实验室,重庆400016 [2]重庆医科大学重庆市生物医学工程学重点实验室,重庆400016

出　　处：《上海交通大学学报（医学版）》2023年第10期1245-1254,共10页Journal of Shanghai Jiao tong University:Medical Science

摘　　要：目的·分析大肠埃希菌外膜囊泡(Escherichia coli outer membrane vesicle,E.coli-OMV)在体外对小鼠4T1乳腺癌细胞增殖的影响,在体内对BALB/c-4T1荷瘤小鼠肿瘤生长的抑制作用。方法·从E.coli的培养上清液中收集OMV并进行表征。采用荧光标记追踪法检测E.coli-OMV被4T1细胞摄入的情况。采用CCK-8法测定E.coli-OMV对4T1细胞增殖的影响。采用流式细胞术检测E.coli-OMV对4T1细胞周期的影响。利用皮下接种法建立BALB/c-4T1荷瘤小鼠模型,而后将小鼠分为E.coli-OMV组与对照(Control)组(每组10只);前者于每2 d注射0.25 mg/kg E.coli-OMV,后者则注射等剂量的PBS。观察2组小鼠的体质量、40 d生存率、肿瘤体积及质量变化,采用苏木精-伊红染色(hematoxylin-eosin staining,H-E染色)评价肿瘤组织的病理学形态,采用免疫组织化学染色法观察肿瘤组织中增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)与细胞周期蛋白D1 (CyclinD1)的表达情况。结果·E.coli-OMV是粒径为(216.00±18.30) nm的球形膜囊结构,可表达E.coli外膜蛋白A (outer membrane protein A,OmpA)与OmpC。荧光显微镜的观察结果显示4T1细胞可摄入E.coli-OMV。CCK-8法的结果显示,E.coli-OMV对4T1细胞的抑制作用与时间-剂量呈正相关。流式细胞术的结果显示,E.coli-OMV将4T1细胞的生长周期阻滞于G0/G1期。体内实验显示,与Control组相比,E.coli-OMV组小鼠的体质量在初期注射后稍有下降(P=0.031),而后出现回升;其40 d生存率增加(P=0.037);该组小鼠肿瘤的体积、质量的增长量均较少(P=0.041,P=0.004),其肿瘤体积抑制率达29.69%、肿瘤质量抑制率达49.81%。H-E染色的结果显示,E.coli-OMV组小鼠肿瘤组织细胞核分裂象较Control组减少(P=0.038)。免疫组织化学染色的结果显示,E.coli-OMV组小鼠肿瘤组织中的PCNA及CyclinD1阳性表达较Control组减少(P=0.031,P=0.002)。结论·体外研究与体内研究均表明,E.coli-OMV可对4T1细胞的增殖产生�Objective·To analyze the effect of Escherichia coli outer membrane vesicle(E.coli-OMV)on the proliferation of 4T1 breast cancer cells in vitro and the inhibition of tumor growth in BALB/c-4T1 tumor-bearing mice in vivo.Methods·OMVs were collected from the culture supernatant of E.coli and characterized.The uptake of E.coli-OMV by 4T1 cells was detected by fluorescent label tracking method.The effect of E.coli-OMV on 4T1 cell proliferation was detected by CCK-8 method.The effect of E.coli-OMV on 4T1 cell cycle was detected by flow cytometry.The BALB/c-4T1 tumor-bearing mouse models were established by subcutaneous inoculation,and the mice were divided into E.coli-OMV group and Control group,with 10 mice in each group.The mice in the E.coli-OMV group were injected with 0.25 mg/kg E.coli-OMV every 2 d,while the mice in the Control group were injected with equal doses of PBS.The changes in body weight,40 d survival rate,tumor volume and tumor weight of the two groups of tumor-bearing mice were observed.The pathological morphology of the tumor tissues was evaluated by hematoxylin-eosin staining(H-E staining).The expression of proliferating cell nuclear antigen(PCNA)and CyclinD1 in tumor tissues was observed by immunohistochemical staining.Results·E.coli-OMV was spherical membrane vesicle structure with a particle size of(216.00±18.30)nm,which expressed E.coli outer membrane protein A(OmpA)and OmpC.Fluorescence microscopy results showed that 4T1 cells could intake E.coli-OMV.CCK-8 results showed that the inhibitory effect of E.coli-OMV on 4T1 cells was positively correlated with time-dose.Flow cytometry results showed that E.coli-OMV arrested the growth cycle of 4T1 cells in G0/G1 phase.In vivo experiments showed that compared with the Control group,body weight of mice in the E.coli-OMV group decreased slightly after the initial injection(P=0.031),and then recovered,while 40 d survival rate increased(P=0.037).The growth of tumor volume and weight of mice in E.coli-OMV group were lower than those in the Control grou

关 键 词：细菌外膜囊泡 乳腺癌 细胞周期 抑瘤作用 

分 类 号：R737.9[医药卫生—肿瘤]