摘要
目的 探讨外泌体来源的miR-1246对食管鳞状细胞癌(esophageal squamous cell carcinoma,ESCC)转移和自噬的影响。方法 用外泌体提取试剂盒提取ESCC细胞外泌体,电镜观察形态,纳米粒径分析仪检测粒径,Western blot法检测外泌体标志物TSG101和Calnexin的表达情况。对ESCC细胞与正常食管上皮细胞外泌体进行RNA测序,分析差异表达微小核糖核酸(microRNA,miRNA),并对差异表达miRNA进行KEGG Pathway富集分析。通过转录组测序评价155例ESCC患者的癌组织和癌旁组织中miR-1246的表达水平。Transwell小室试验检测miR-1246对ESCC细胞迁移能力的影响;Western blot法检测miR-1246对ESCC细胞自噬的影响。结果 ESCC细胞来源的外泌体形状完整,近似球形囊泡样结构;粒径为50~80 nm;外泌体标志物TSG101蛋白呈阳性,Calnexin蛋白呈阴性。ESCC细胞与正常食管上皮细胞外泌体有59个共同差异表达miRNA,外泌体miR-1246在ESCC细胞中呈高表达,差异表达miRNA主要富集在自噬代谢通路上。外泌体miR-1246在ESCC患者的癌组织中呈高表达。过表达miR-1246可显著促进ESCC细胞的迁移和自噬(t分别为4.119和48.150,P分别<0.05和<0.001),抑制表达miR-1246可抑制ESCC细胞的迁移和自噬(t分别为9.067和51.270,P分别<0.01和<0.001)。结论 外泌体来源的miR-1246可显著影响ESCC细胞的转移和自噬。
Objective To investigate the effects of exosome-derived miR-1246 on metastasis and autophagy of esophageal squamous cell carcinoma(ESCC).Methods The exosomes of ESCC cells were extracted by exosome extraction kit,the morphology was observed under electron microscope,the particle size was detected by nano-particle size analyzer,and the expression of exosome markers TSG101 and Calnexin was detected by Western blot.The RNA of exosomes of ESCC cells and normal esophageal epithelial cells were sequenced to analyze the differentially expressed miRNAs,which were analyzed by KEGG Pathway enrichment.The expression of miR-1246 in tumor tissues and paracancer tissues of 155 patients with ESCC was evaluated by transcriptome sequencing.The effect of miR-1246 on the migration ability of ESCC cells was detected by Transwell assay and the effect on autophagy was detected by Western blot.Results The exosomes derived from ESCC cells were intact in shape and similar to spherical vesicle-like structure with the particle size of 50~80 nm.The exosome marker TSG101 protein was positive and Calnexin protein was negative.There were 59 common differentially expressed miRNAs between exosomes of ESCC cells and normal esophageal epithelial cells.Exosome miR-1246 was highly expressed in ESCC cells,and the differentially expressed miRNAs were mainly enriched in autophagy metabolic pathway.Exosome miR-1246 was highly expressed in cancer tissues of patients with ESCC.Overexpression of miR-1246 significantly promoted the migration and autophagy of ESCC cells(t=4.119 and 48.150,P<0.05 and<0.001,respectively).Inhibition of miR-1246 expression inhibited the migration and autophagy of ESCC cells(t=9.067 and 51.270,P<0.01 and<0.001,respectively).Conclusion miR-1246 derived from exosomes can significantly affect the metastasis and autophagy of ESCC cells.
作者
彭美兰
王璐
刘丽丽
闫振鹏
阎婷
PENG Meilan;WANG Lu;LIU Lili;YAN Zhenpeng;YAN Ting(Translational Medicine Research Center,Shanxi Medical University,Taiyuan 030001,Shanxi Province,China)
出处
《中国生物制品学杂志》
CAS
CSCD
北大核心
2023年第10期1158-1165,共8页
Chinese Journal of Biologicals
基金
国家自然科学基金青年项目(81702449)。
