胰腺癌患者血浆外泌体circ-BCAS3的水平和临床意义

Clinical value of circ-BCAS3 in plasmal exosomes of patients with pancreatic cancer

目的探讨胰腺癌患者血浆外泌体环状RNA-BCAS3(circ-BCAS3)的水平及临床意义。方法通过生物信息学网站exoRBase筛选胰腺癌患者血浆外泌体中异常表达的circRNAs。收集2018年1月至2020年2月年龄、性别相匹配的胰腺癌患者与健康体检者的血浆各20例,通过试剂盒分离、提取外泌体并通过电子显微镜检测验证分离效果。实时荧光定量PCR(qPCR)对血浆外泌体中的circRNAs定量检测并结合临床信息和随访数据分析circ-BCAS3水平与胰腺癌临床病理特征和预后的关系。使用circ-BCAS3小干扰RNA(si-circ-BCAS3)和无义核酸序列(si-NC)转染胰腺癌细胞PANC-1和BxPC3,采用CCK-8和流式细胞术检测细胞增殖和凋亡情况。结果通过肿瘤外泌体数据库exoRBase在胰腺癌患者血浆外泌体中筛选出3个高表达的circRNAs(circ-CMTR1、circ-SP1和circ-BCAS3),表达验证发现胰腺癌患者血清外泌体circ-BCAS3水平为16.98±2.33,高于健康对照的6.37±1.18(P<0.05)。circ-BCAS3水平与胰腺癌患者的TNM分期和淋巴结转移有关(P<0.05),而与性别、年龄、肿瘤大小和分化程度无关(P>0.05)。circ-BCAS3高水平患者中位总生存期为5.6个月,低于低表达患者的13.4个月(P<0.05)。细胞实验结果显示,与转染si-NC的细胞相比,转染si-circ-BCAS3后,PANC-1和BxPC3细胞株的增殖活力降低而凋亡率升高(P<0.05)。结论circ-BCAS3异常高表达于胰腺癌患者血浆外泌体中,影响胰腺癌细胞的增殖与凋亡,与胰腺癌患者的预后有关,可作为潜在的诊断以及预后标志物。

Objective To investigate the clinical value of circular RNA-BCAS3(circ-BCAS3)in plasmal exosomes of patients with pancreatic cancer.Methods The abnormally expressed exosomal circRNAs in pancreatic cancer patients were screened by bioinformatics website exoRBase.Twenty plasma samples from pancreatic cancer patients and 20 samples from age and sex matched healthy volunteers were collected from Jan 2018 to Feb 2020.Exosomes were separated by SBI ExoQuick kit and verified by electron microscope.Levels of circRNAs were quantified through real-time quantitative PCR(qPCR).Relationship of circ-BCAS3 levels with clinico-pathological features and prognosis of pancreatic cancer were analyzed.Nonsense nucleic acid sequence(si-NC)and small interfering RNAs targeting circ-BCAS3(si-circ-BCAS3)were transfected into pancreatic cancer cells(PANC-1 and BxPC3).CCK-8 assay and flow cytometry were used to detect proliferative activity and apoptotic rate,respectively.Results Three predicted circRNAs including circ-CMTR1,circ-SP1 and circ-BCAS3 were screened and qPCR analysis showed that the exosomal level of circ-BCAS3 in pancreatic cancer was 16.98±2.33,significantly higher than 6.37±1.18 in healthy volunteers(P<0.05).Circ-BCAS3 level was associated with TNM stage and lymph node metastasis in pancreatic cancer patients(P<0.05),but not related to gender,age,tumor size and differentiation degree(P>0.05).The median overall survival of patients with higher circ-BCAS3 level was 5.6 months,shorter than 13.4 months of patients with low circ-BCAS3 level(P<0.05).In addition,functional assay showed that compared with cells transfected with si-NC,the proliferation ability of cells transfected with si-circ-BCAS3 was suppressed and apoptotic rate was up-regulated in PANC-1 and BxPC3 cell lines(P<0.05).Conclusion Circ-BCAS3 was highly expressed in pancreatic cancer patients and might contribute to the tumor progression in pancreatic cancer.This circRNA may serve as a novel potential diagnostic biomarker and provide potential effective therapeutic target.