目的探讨miR-191/425基因簇及其靶基因肾上腺髓质素2(Recombinant adrenomedullin 2,ADM2)、异连蛋白α(Dystrobrevin alpha,DTNA)、小整合膜蛋白11B(Homo sapiens small integral membrane protein 11B,SMIM11B)在食管癌组织中的表达,...目的探讨miR-191/425基因簇及其靶基因肾上腺髓质素2(Recombinant adrenomedullin 2,ADM2)、异连蛋白α(Dystrobrevin alpha,DTNA)、小整合膜蛋白11B(Homo sapiens small integral membrane protein 11B,SMIM11B)在食管癌组织中的表达,及与食管癌预后风险的关系。方法应用肿瘤基因组图谱(TCGA)数据库生物信息学工具分析食管癌miR-191/425基因簇及其靶基因对食管癌预后的影响;RT-qPCR验证36例食管癌组织miR-191/425基因簇及其靶基因ADM2、DTNA、SMIM11B在肿瘤组织中的表达,分析miR-191/425异常表达对食管癌发病风险的影响,利用受试者特征曲线(ROC)评价miR-191/425基因簇对食管癌的诊断效力。结果(1)生物信息学分析显示,ADM2、DTNA、SMIM11B低表达与食管癌患者的预后差有关。(2)RT-qPCR结果显示,食管癌组织中miR-191、miR-425表达显著上调,肿瘤组织中miR-191的表达上调与肿瘤T3+T4临床分期相关,miR-191和miR-425的表达呈显著正相关(r=0.639,P=0.000)。(3)Logistic回归分析显示,miR-191表达水平与食管癌的发病风险呈正相关[OR(95%CI)为:1.199(1.036~1.386),P<0.05];ROC分析显示,miR-191曲线下面积(AUC)为0.650(P<0.05)。(4)临床病理因素分析显示,在T3+T4期、淋巴结转移的肿瘤组织中DTNA mRNA的表达水平显著升高,ADM2 mRNA、SMIM11B mRNA无显著差异。(5)36例食管癌组织中DTNA mRNA的表达与miR-191和miR-425的表达呈显著正相关。结论在食管癌发生发展中,miR-191/425基因簇共同高表达增强了肿瘤细胞的侵袭,其机制可能与共同促进DTNA靶基因的表达有关,miR-191可能作为一个主要因素对食管癌具有诊断效力。展开更多
BACKGROUNDMultiple exosomal microRNAs were reported to have a significant role incolorectal cancer (CRC) cells. The function and mechanism of exosomal miR-191in CRC have not been clearly elucidated.AIMTo explore the r...BACKGROUNDMultiple exosomal microRNAs were reported to have a significant role incolorectal cancer (CRC) cells. The function and mechanism of exosomal miR-191in CRC have not been clearly elucidated.AIMTo explore the roles of miR-191 in CRC.METHODSSupernatant exosomes from CRC cells were extracted and identified. After coculture,macrophage polarization was determined using flow cytometry for themarkers cluster of differentiation (CD) 68 and CD163, enzyme-linked immunosorbentassay for the cytokines interleukin (IL)-4 and IL-10, western blotting forchitinase-like protein 3 and arginase-1 expression, and immunofluorescentstaining for 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindocarbocyanine perchlorate.Reactive oxygen species (ROS) level, ferroptosis-related proteins (SLC7A11 andGPX4), and apoptosis were determined with flow cytometry, western blotting,and TUNEL staining. We performed in vivo experiments to determine the functionof exosomal miR-191 and M2 macrophage polarization.RESULTSWe successfully isolated exosomes from CRC cells. Inhibition of miR-191 in CRCcells suppressed M2 polarization of macrophages. After coculture of macrophages,inhibition of miR-191 induced ROS production, ferroptosis, and apoptosisof CRC cells. Silencing of exosomal miR-191 from CRC cells prevented M2polarization of macrophages, and weakened CRC development by inducingferroptosis. Exosomal miR-191 accelerated cancer progression in CRC nude miceby promoting M2 polarization of macrophages. CONCLUSIONInhibition of exosomal miR-191 attenuated CRC progression by inducing ferroptosis in macrophages. This studyrevealed a novel mechanism by which exosomal miR-191 modulates the tumor microenvironment.展开更多
文摘目的探讨miR-191/425基因簇及其靶基因肾上腺髓质素2(Recombinant adrenomedullin 2,ADM2)、异连蛋白α(Dystrobrevin alpha,DTNA)、小整合膜蛋白11B(Homo sapiens small integral membrane protein 11B,SMIM11B)在食管癌组织中的表达,及与食管癌预后风险的关系。方法应用肿瘤基因组图谱(TCGA)数据库生物信息学工具分析食管癌miR-191/425基因簇及其靶基因对食管癌预后的影响;RT-qPCR验证36例食管癌组织miR-191/425基因簇及其靶基因ADM2、DTNA、SMIM11B在肿瘤组织中的表达,分析miR-191/425异常表达对食管癌发病风险的影响,利用受试者特征曲线(ROC)评价miR-191/425基因簇对食管癌的诊断效力。结果(1)生物信息学分析显示,ADM2、DTNA、SMIM11B低表达与食管癌患者的预后差有关。(2)RT-qPCR结果显示,食管癌组织中miR-191、miR-425表达显著上调,肿瘤组织中miR-191的表达上调与肿瘤T3+T4临床分期相关,miR-191和miR-425的表达呈显著正相关(r=0.639,P=0.000)。(3)Logistic回归分析显示,miR-191表达水平与食管癌的发病风险呈正相关[OR(95%CI)为:1.199(1.036~1.386),P<0.05];ROC分析显示,miR-191曲线下面积(AUC)为0.650(P<0.05)。(4)临床病理因素分析显示,在T3+T4期、淋巴结转移的肿瘤组织中DTNA mRNA的表达水平显著升高,ADM2 mRNA、SMIM11B mRNA无显著差异。(5)36例食管癌组织中DTNA mRNA的表达与miR-191和miR-425的表达呈显著正相关。结论在食管癌发生发展中,miR-191/425基因簇共同高表达增强了肿瘤细胞的侵袭,其机制可能与共同促进DTNA靶基因的表达有关,miR-191可能作为一个主要因素对食管癌具有诊断效力。
文摘BACKGROUNDMultiple exosomal microRNAs were reported to have a significant role incolorectal cancer (CRC) cells. The function and mechanism of exosomal miR-191in CRC have not been clearly elucidated.AIMTo explore the roles of miR-191 in CRC.METHODSSupernatant exosomes from CRC cells were extracted and identified. After coculture,macrophage polarization was determined using flow cytometry for themarkers cluster of differentiation (CD) 68 and CD163, enzyme-linked immunosorbentassay for the cytokines interleukin (IL)-4 and IL-10, western blotting forchitinase-like protein 3 and arginase-1 expression, and immunofluorescentstaining for 1,1’-dioctadecyl-3,3,3’,3’-tetramethylindocarbocyanine perchlorate.Reactive oxygen species (ROS) level, ferroptosis-related proteins (SLC7A11 andGPX4), and apoptosis were determined with flow cytometry, western blotting,and TUNEL staining. We performed in vivo experiments to determine the functionof exosomal miR-191 and M2 macrophage polarization.RESULTSWe successfully isolated exosomes from CRC cells. Inhibition of miR-191 in CRCcells suppressed M2 polarization of macrophages. After coculture of macrophages,inhibition of miR-191 induced ROS production, ferroptosis, and apoptosisof CRC cells. Silencing of exosomal miR-191 from CRC cells prevented M2polarization of macrophages, and weakened CRC development by inducingferroptosis. Exosomal miR-191 accelerated cancer progression in CRC nude miceby promoting M2 polarization of macrophages. CONCLUSIONInhibition of exosomal miR-191 attenuated CRC progression by inducing ferroptosis in macrophages. This studyrevealed a novel mechanism by which exosomal miR-191 modulates the tumor microenvironment.
文摘对实验室前期Solexa结果进行深入挖掘,结合生物信息学分析从不同发育阶段猪皮下脂肪组织差异表达的miRNAs中筛选出高丰度差异极显著的候选miR-191.采用腺病毒过表达miR-191,实时定量PCR、Western blot及双荧光素酶报告基因检测等技术方法,初步研究miR-191对猪前体脂肪细胞分化的影响.结果发现,miR-191随着猪前体脂肪细胞的分化表达量逐渐增加.与对照组相比,过表达miR-191的猪前体脂肪细胞中miR-191转录本显著增加,并引起CCAAT增强子结合蛋白β(C/EBPβ)、PPARγ和aP2的mRNA水平降低,抑制了猪前体脂肪细胞分化.同时,Western blot结果显示,与对照组相比过表达miR-191的猪前体脂肪细胞在48 h C/EBPβ蛋白水平下降了55%.更重要的是,通过TargetScan等算法正向筛选以及MicroInspector反向筛选联合获得miR-191候选靶基因,经双荧光素酶报告基因检测结果证实,miR-191可直接作用于C/EBPβ3′UTR,从而降低萤火虫荧光素酶活性.综上所述,miR-191可能通过抑制脂肪细胞分化早期标志基因C/EBPβ的表达,从而抑制了猪前体脂肪细胞的分化.
