肾透明细胞癌血清差异性蛋白质表达及临床意义研究

Expression and Clinical Significance of Serum Differentiated Protein among Patients with Clear-cell Renal Cell Carcinoma

背景与目的利用蛋白质芯片技术寻找肿瘤标志物仍是热点,但目前局限于仅对肾癌患者进行术前、术后的差异性蛋白质检测。为深入检测差异性蛋白质的变化规律,本研究探讨了肾透明细胞癌血清差异性蛋白质表达及临床意义。方法选取2013年11月—2014年4月在新疆医科大学第一附属医院泌尿外科手术治疗并且术后经病理学专家诊断为肾透明细胞癌患者40例为肾癌组,选择同期在本院体检健康志愿者以及非肾癌患者53例为对照组。运用表面增强激光解析电离化飞行时间质谱系统(SELDI-TOF-MS)和CM10弱阳离子交换蛋白芯片技术系统检测肾癌组及对照组血清差异性蛋白质。应用线性支持向量机(SVM)方法建立蛋白质指纹图诊断模型,留一法交叉验证模型判别效果。通过ZUCI-PDAS蛋白质谱数据分析系统软件对血清差异性蛋白质进行分析。结果最终5种蛋白质作为潜在标志蛋白,质荷比(M/Z)分别为15 953、7 987、9 304、8 948、5 911,所对应的蛋白质依次为Bcl-2家族细胞凋亡调节蛋白、WAP二硫化四物核心蛋白、Krueppel样因子8、单核细胞趋化蛋白1、血清β-淀粉样蛋白4。肾癌组术前与对照组5种蛋白质表达水平比较,差异有统计学意义(P<0.05)。肾癌组术前与术后1周、1个月、3个月、6个月5种蛋白质表达水平比较,差异均有统计学意义(P<0.05)。术前5种蛋白质表达水平均高于术后1周、1个月、3个月、6个月(P<0.05);术后3、6个月15 953、7 987蛋白质表达水平与术后1周比较,差异有统计学意义(P<0.05);术后6个月15 953、7 987蛋白质表达水平与术后3个月比较,差异有统计学意义(P<0.05)。术后1、3、6个月9 304、8 948、5 911蛋白质表达水平与术后1周比较,差异有统计学意义(P<0.05);术后6个月9 304、8 948、5 911蛋白质表达水平与术后1个月比较,差异有统计学意义(P<0.05);术后6个月5 911蛋白质表达水平与术后3个月比较,差异有统计学意义(P<0.05)。5种蛋白质作为生物标志物,预测肾透明细胞癌的灵敏度为87.5%(35/40),特异度为86.8%(46/53)。结论特异性蛋白质可能依次为Bcl-2家族细胞凋亡调节蛋白、WAP二硫化四物核心蛋白、Krueppel样因子8、单核细胞趋化蛋白、血清β-淀粉样蛋白4,共有望成为肾癌肿瘤标志物,对肾透明细胞癌的预测价值较高,在今后肾透明细胞癌的疗效评价、预后评估以及靶向治疗方面有一定参考价值。

Background and Objective Looking for tumor markers by using protein chip technology is one of the hot topics,but many studies are still limited to detection of differentiated protein before and after surgery among patients with kidney cancer. In order to detect the variability of differentiated protein,the expression and clinical significance of serum differentiated protein among patients with clear - cell renal cell carcinoma were discussed in this study. Methods 40 patients with clear - cell renal cell carcinoma who underwent surgery in Department of Urology Surgery in the First Affiliated Hospital of Xinjiang Medical University from November 2013 to April 2014,all cases were confirmed by postoperative pathology and were included in kidney cancer group,53 healthy volunteers who underwent medical examination in the same hospital and patients without kidney cancer were included in control group. The surface - enhanced laser desorption ionization time of flight mass spectrometry（SELDI - TOF - MS）and weak cation exchange protein chip（CM10）technology systems were used to identify the serum differentiated protein in kidney cancer group and in control group. The linear support vector machine（ SVM）method was used to establish the diagnostic model of protein fingerprints,and the leave - one - out cross validation was used to identify model discrimination effect. The differentially expressed proteins were analyzed by ZUCI - PDAS protein spectral data analysis system software. Results Five kinds of proteins were identified as potential marker protein,the M/ Z of these proteins was 15 953,7 987,9 304, 8 948,5 911,respectively,the corresponding protein was Bcl - 2 family apoptosis regulatory proteins,WAP four - disulfide core protein,Krueppel - like factor 8,monocyte chemotactic protein - 1,serum amyloid β - protein - 4,respectively. There were significant differences in expression levels of five kinds of proteins between kidney cancer group befrore surgery and control group（P ﹤ 0. 05）. There were significant differences in expression levels of five kinds of proteins in kidney cancer group among before surgery,1 week after surgery,1 month after surgery,3 months after surgery,6 months after surgery（P ﹤ 0. 05）. The expression levels of five kinds of proteins before surgery were significantly higher than those 1 week after surgery,1 month after surgery,3 months after surgery,6 months after surgery,respectively（ P ﹤ 0. 05 ） . There were significant differences in expression levels of protein 15 953 and 7 987 between 3,6 months after surgery and 1 week after surgery,respectively（P ﹤0. 05）. There were significant differences in expression levels of protein 15 953 and 7 987 between 3 months after surgery and 6 months after surgery（ P ﹤ 0. 05）. There were significant differences in expression levels of protein 9 304,8 948 and 5 911 between 1,3,6 months after surgery and 1 week after surgery,respectively（P ﹤ 0. 05）. There was significant difference in expression levels of protein 5 911 between 3 months after surgery and 6 months after surgery（ P ﹤ 0. 05）. The sensitivity and specificity of five kinds of proteins in predicting clear - cell renal cell carcinoma was 87. 5 % （35 / 40）and 86. 8% （46 / 53）, respectively. Conclusion The specific proteins may include Bcl - 2 family apoptosis regulatory proteins,WAP four - disulfide core protein,Krueppel - like factor 8,monocyte chemotactic protein - 1 and serum amyloid β - protein - 4,respectively, which may be used as tumor markers of kidney cancer,these proteins have high predictive value for clear - cell renal cell carcinoma,and may contribute to therapeutic evaluation,prognosis and targeted therapy for clear - cell renal cell carcinoma.