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目的:建立尿液中8种微小RNA(micro RNA,miRNA)的检测方法 ,明确其分布规律及其在膀胱癌中的变化。方法:选取15名健康人的晨尿及随机尿液,以及15例确诊膀胱癌患者的随机尿液,进行总RNA提取,然后反转录得到cDNA,用荧光实时PCR法扩增8种膀胱癌相关miRNA,分别为miR-126、miR-200a、miR-200b、miR-200c、miR-182、miR-183、miR-429和miR-141,探讨晨尿与随机尿、尿液浓缩前与浓缩后对尿液miRNA浓度的影响。比较正常对照组与膀胱癌患者尿液中上述8种miRNA的表达差异,并采用受试者工作特征曲线分析尿液miRNA检测在膀胱癌诊断中的价值。结果:浓缩尿液组的6种miRNA(miR-126、miR-200a、miR-200b、miR-200c、miR-429和miR-141)浓度显著高于未浓缩组(P<0.05),而浓缩晨尿组的8种miRNA浓度与浓缩随机尿组间比较无统计学差异(P>0.05)。与正常对照相比较,miR-126、miR-182、miR-183及miR-141的浓度在膀胱癌组中显著升高(P<0.05),而miR-200a浓度在膀胱癌组中显著降低(P<0.05)。受试者工作特征曲线分析显示,单独检测尿液miRNA对于膀胱癌的诊断缺乏有效性,而对5种尿液miRNA(miR-126、miR-200a、miR-182、miR-183和miR-141)进行联合检测则可显著提高膀胱癌诊断的灵敏度和特异度。结论:尿液中稳定存在多种miRNA,尿液中多项miRNA的联合检测可能有望成为诊断膀胱癌的有效指标。
OBJECTIVE: To establish a method for the detection of 8 microRNAs (miRNAs) in urine and clarify its distribution and its changes in bladder cancer. Methods: The morning urine and random urine of 15 healthy people were selected, and the random urine of 15 patients diagnosed with bladder cancer were selected for total RNA extraction. CDNA was reverse transcribed, and 8 kinds of bladder cancer were amplified by real-time fluorescent PCR Related miRNAs were used to evaluate the effects of morning urine and random urine on the expression of miR-126, miR-200a, miR-200b, miR-200c, miR-182, miR-183, miR-429 and miR- After the impact on the concentration of urine miRNA. The difference of expression of the above eight kinds of miRNAs in the urine of normal control group and bladder cancer patients was compared and the value of urine miRNA detection in diagnosis of bladder cancer was analyzed by using the working characteristic curve of the subjects. Results: The concentrations of six miRNAs (miR-126, miR-200a, miR-200b, miR-200c, miR-429 and miR-141) in the concentrated urine group were significantly higher than those in the non-concentrated group No significant difference was found between the concentration of 8 miRNAs in morning urine group and the concentration of randomized urine group (P> 0.05). Compared with the normal control, the concentrations of miR-126, miR-182, miR-183 and miR-141 were significantly increased in bladder cancer group (P <0.05), while the miR-200a concentration was significantly lower in bladder cancer group P <0.05). Analysis of receiver operating characteristic curves showed that urine miRNAs alone were not effective in the diagnosis of bladder cancer, whereas five urine miRNAs (miR-126, miR-200a, miR-182, miR-183 and miR-141 ) Joint detection can significantly improve the sensitivity and specificity of bladder cancer diagnosis. Conclusion: A variety of miRNAs are stable in urine. The combined detection of multiple miRNAs in urine may be an effective indicator for the diagnosis of bladder cancer.