循环外泌体抑制弥漫大B细胞淋巴瘤细胞增殖及其可能机制研究  

Circulating exosomes inhibits proliferation of diffuse large B-cell lymphoma and its possible mechanism

作　　者：梁艳丽[1] 李娟[2] 李凤[1] 魏容[1] 王欣[3] LIANG Yan-li;LI Juan;LI Feng;WEI Rong;WANG Xin(Department of Clinical Laboratory,An Affiliated Hospital of Chongqing Medical University,Suining 629000,Sichuan,China;Department of Hematology,Suining Central Hospital,An Affiliated Hospital of Chongqing Medical University,Suining 629000,Sichuan,China;Central Laboratory,West China Hospital,Sichuan University,Chengdu 610041,Sichuan,China)

机构地区：[1]遂宁市中心医院.重庆医科大学附属遂宁市中心医院检验科,四川遂宁629000 [2]四川大学华西医院中心实验室,四川成都610041 [3]遂宁市中心医院.重庆医科大学附属遂宁市中心医院血液科,四川遂宁629000

出　　处：《生物医学工程与临床》2019年第1期79-86,共8页Biomedical Engineering and Clinical Medicine

摘　　要：目的探讨循环外泌体对弥漫大B细胞淋巴瘤(DLBCL)细胞增殖的影响及其可能机制。方法选择2016年6月至2017年6月遂宁市中心医院血液科13例经病理诊断明确的DLBCL患者,其中男性8例,女性5例;年龄37~68岁,平均年龄55.6岁。同期14例体检中心的健康体检者,其中男性7例,女性7例;年龄44~69岁,平均年龄58.9岁。获得血浆外泌体。通过电子显微镜和qNano分析仪对体检者、DLBCL患者外泌体进行鉴定。设空白对照组(U2932细胞)、对照组(健康体检者外泌体+U2932细胞)、研究组(DLBCL患者外泌体+U2932细胞),用四甲基偶氮唑盐(MTT)法、流式细胞法和Western blot检测增殖相关蛋白增殖细胞核抗原(PCNA)、Ki-67,综合评估各组细胞的增殖情况;采用高通量技术检测4例DLBCL患者血浆和4例健康体检者血浆外泌体中miRNA表达谱;用实时荧光定量聚合酶链式反应(PCR)检测DLBCL患者和健康体检者血浆外泌体中miR-92b表达水平。结果电子显微镜观察到,健康体检者、DLBCL患者外泌体呈现典型的茶托样结构,qNano分析结果显示所提取的外泌体主要集中在20~100 nm;Western blot结果显示,相比于血浆,外泌体中CD63、CD9这两个外泌体标志蛋白均有明显的表达。流式细胞结果显示,相比于空白对照组,对照组细胞G0/G1期受到抑制(P <0.05);MTT结果显示,相比于空白对照组,对照组细胞增殖能力降低(t=17.3,P <0.05);Western blot结果显示,相比于空白对照组,对照组细胞中PCNA和Ki-67表达减少(t=33.5,P <0.05)。相比于对照组,miRNA芯片分析结果显示,研究组血浆外泌体中miR-92b表达减少,RT-PCR结果显示miR-92b表达减少(t=12.9,P <0.05)。结论循环外泌体通过介导miR-92b抑制DLBCL细胞增殖。Objective To investigate the effect of circulating exosomes on the proliferation of diffuse large B-cell lymphoma(DLBCL)and its possible mechanism.Methods From June 2016 to June 2017,13 patients with DLBCL confirmed by pathological diagnosis were enrolled,which included 8 males and 5 females,aged 37-68 years old with mean age of 55.6 years old.Forteen cases of healthy subjects were selected,which included 7 males and 7 females,aged 44-69 years old with mean age of 58.9 years old.The plasma exosomes were collected and identified by electron microscopy and qNano analyzer.The blank control group(U2932 cell),control group(healthy exosomes+U2932 cell)and study group(DLBCL exosomes+U2932 cell)were established,and methyl thiazolyl tetrazolium(MTT)assay,flow cytometry and Western blot were used to detect proliferation-associated proteins proliferating cell nuclear antigenc(PCNA)and Ki-67.The high-throughput technique was used to detect miRNA expression profiles in exosomes plasma of 4 DLBCL patients and 4 healthy controls.The real-time fluorescence quantitative PCR were used to detect miR-92 b expression levels in plasma exosomes of DLBCL patients and healthy controls.Results The results of electron microscopy observation showed that the exosome had typical saucer like structure,and qNano analysis showed that the extracted exosomes were mainly concentrated at 20-100 nm.Compared with plasma,the Western blot results showed that exosome marker proteins of exosome CD63,CD9 were significant expressed.Compared with blank control group,the results of flow cytometry showed that the G0/G1 phase of control group was inhibited(P<0.05).MTT results showed that the proliferative capacity of control group was decreased(t=17.3,P<0.05)and Western blot results showed that the expression of PCNA and Ki-67 of control group were decreased(t=33.5,P<0.05).Compared with control group,miRNA microarray analysis showed that the expression of miR-92 b in exosomes was decreased in study group,and the RT-PCR results showed that the expression of miR-

关 键 词：弥漫大B细胞淋巴瘤 外泌体 miR-92b 细胞增殖 

分 类 号：R733.4[医药卫生—肿瘤]