【作者】 赵致广；

【导师】 邵国兴；

【作者基本信息】 第四军医大学 ， 外科学， 2004， 博士

【摘要】 肾癌是人类较常见的恶性肿瘤，发病率占所有人类恶性肿瘤的3％，是病死率最高的泌尿系肿瘤。多型核粒细胞弹性蛋白酶是一个强有力的丝氨酸蛋白酶，它能够降解组成细胞外基质的大多数蛋白成分，部份人类肿瘤细胞也可以分泌该酶并且在肿瘤进展的过程中起重要的作用。我们采用分子生物学免疫学和病理学的方法，检测PMNE在肾癌组织中的表达，肾癌细胞对肾血管的侵袭，肾癌组织中PMNE启动子重复序列的测定，将野生型和突变型PMNE启动子插入pGL3-Basic来比较两者对构建载体表达荧光素酶的研究，探讨PMNE在肾癌发生发展过程中的作用。 研究结果如下： 1．PMNE在肾癌组织中高水平表达，与TNM分期关系密切，T3a、T3b期肾癌组织抽提物中PMNE的浓度比T1、T2组明显增高。 2．PMNE在肾癌组织中的表达与癌细胞对微血管的侵袭关系密切。 3．发现1例肾癌患者PMNE启动子第3区-879位、第6区-726位碱基处分别发生单核苷酸置换突变，由原来的A突变为C。第.军医大母博士李位恰久4.插入突变PMNE启动子的荧光素酶表达载体转染的K562细胞中荧光素酶表达强度明显高于野生型P刚E启动子转染的细胞。结论二P删E在肾癌组织中高表达且与分期关系密切，PMNE启动子第3区一879位、第6区一716位碱基处分别发生单核昔酸置换突变，由原来的A突变为C，导致PMNE的表达明显增强，可能参与和促进了肾癌细胞对细胞外基质的侵袭。我们的实验结果证明PMNE在肾癌发生发展的过程中起了重要的作用。更多还原

【Abstract】 Renal cell carcinoma (RCC), which accounts for about 3% of all adult malignancy tumors, is the most lethal of the urologic cancers. Although tumor size, tumor grades and nuclear atypia of RCC have been reported as prognostic factors, the biological behavior of this tumor is still unpredictable. Polymorphonuclear leukocyte elastase (PMNE) is a powerful serine protease, since it degrads most protein components of the extracellular matrix, it is suggested to play a crucial role in the processes of some tumor cell invasion and metastasis, including non-small cell lung carcinoma and primary breast cancer. Because there is no published information on the relation between the level of PMNE and prognosis in patients of RCC, we, in this study, explored if PMNE level was involved in the prognosis of RCC patients. Using molecular, immunological and pathological methods, we measured levels of PMNE incytoplasmic extracts of 59 renal cell carcinomas and observed microvascular invasion of RCC cells and sequenced PMNE promoter gene of RCC, which showed single nucleic acid mutant whose promoter activity was also compared with wild type PMNE promoter through inserting both promoters into pGL3-Basic to construct luciferase expression vector. Results:1. The PMNE was over expressed in RCC cell and the level of PMNE in the tissue extract of T3a and T3b RCC was much more higher than those of T1 and T2, So we conclude that the level of PMNE expression in RCC cell is closely related to the TNM staging2. High level PMNE expression was detected in the specimen of RCC with microvascular invasion, which indicated that the PMNE over expression in RCC might be related to the tumor invasion into microvascular.3. One important finding of this study was that there were 1 cases of RCC whose PMNE promoter gene sequence had permutation mutant from C to A occurred in the -879th and -716th base of region3 6.4. Luciferase expressed in K562 cells transfected with mutant PMNE promoter luciferase expression vector was considerablely higher than those transfected with wild type PMNE promoter luciferase expression vector, which suggests that PMNE promoter occurred in RCC enhanced this enzyme activity.CONCLUSION:PMNE is over expressed in RCC and the level of its expression is closely related with the stage and the invasion abilities of the tumor. PMNE promoter gene permutation mutant in the region 3 6 may be the cause of higher activity of PMNE enzyme, which promoted the invasion of RCC cell within the matrix. Our data indicated that PMNE may play an important role in the development of RCC.更多还原