探讨乳腺癌组织中膜联蛋白(annexin)A2表达异常及其泛素化调节机制，annexin A2与乳腺癌TNM临床分期的关系，泛素化调节annexin A2表达异常在乳腺癌发生发展中的作用。

选择2008年9月至2009年9月在本院普外三科确诊为乳腺癌的10例患者手术切除的新鲜组织，包括组织病理学证实的乳腺癌组织10份及癌旁形态学相对正常组织10份(直径>2 cm)为研究对象。组织切除后，立即液氮保存待用。采用二维聚丙烯酰胺凝胶电泳(2-DE)技术，对乳腺癌组织及其癌旁正常组织的蛋白质进行分离，Western bloting法检测该蛋白质的泛素化(免疫蛋白质组学)情况，取稳定出现的差异阳性蛋白质点进行基质辅助激光解吸/电离时间飞行时间质谱(MALDI-TOF-MS)、质谱/质谱串联(MS-MS)及生物信息学分析。通过免疫组化染色技术，对annexin A2在乳腺癌组织及其癌旁正常组织中的表达水平，与乳腺癌TNM临床分期的关系进行观察，并对结果进行统计学分析(本研究遵循的程序符合本院人体试验委员会所制定的伦理学标准，得到该委员会批准，征得受试对象本人的知情同意，并与之签署临床研究知情同意书)。

对4个在乳腺癌组织标本及其癌旁正常组织中明显表达差异的蛋白质点，经质谱鉴定和数据库比对的结果为小泛素相关修饰因子3前体(SMT3A)、蛋白酶体亚单位a型1(PSMA1)、annexin A2及核糖体蛋白S12(RPS12)。免疫组化检测在TNM临床分期为0，Ⅰ，Ⅱ及Ⅲ期的乳腺癌组织标本中，annexin A2表达的阳性率及强阳性率分别为75.0%(6/8)与37.5%(3/8)，100.0%(14/14)与42.9%(6/14)，100.0%(10/10)与66.7%(4/6)及100.0%(6/6)与50.0%(5/10)。在TNM临床分期不同的乳腺癌组织中，annexin A2表达的阳性率及强阳性率比较，差异均有统计学意义(P<0.05)。annexin A2的表达水平与乳腺癌的TNM临床分期呈正相关(r＝0.508，P<0.05)。

annexin A2在乳腺癌组织中的表达受泛素化调节，且表达增高。乳腺癌TNM临床分期越晚，annexin A2表达水平越高。泛素化调节annexin A2的高表达，可能与乳腺癌的发生发展有关。

To study the abnormal expression of annexin A2 and the ubiquitniation regulatory mechanism in breast cancer tissue, relationship between annexin A2 and TNM clinical stages of breast cancer, effects of abnormal expression of ubiquitination regulation annexin A2 in the process of breast cancer development.

From September 2008 to September 2009, a total of 10 cases of women were diagnosed as breast cancer. The breast cancer tissue samples by pathology (n=10) and para-tumor relatively normal tissue based on morphology (n=10) (diameter>2 cm) which were surgically removed from those 10 cases patients. Samples of tissues were preserved with liquid nitrogen as soon as it had been excised. Two-D polyacrylamide gel eletrophresis (2-DE) was applied to separate protein of breast cancer tissue and para-tumor normal tissues, and confirmed ubiquitination (immunoproteomics) was detected by Western bolting with PDQuest software (7.1), then MALDI-TOF-TOF, MS-MS and bioinformatics analysis was performed to obtain differential proteins ubiquitinated. Different expression of annexin A2 in breast cancer tissue and para-tumor normal tissues, as well as their relationship with TNM clinical stages of breast cancer were observed and analyzed by applying the method of immunohistochemical staining and SPSS 13.0. The study protocol was approved by the Ethical Review Board of Investigation in Human Being of Affiliated Hospital of North Sichuan Medical College. Informed consent was obtained from each participates.

Four proteins with identified by immunoproteomics, they were, SMT3A, PSMA1, annexin A2 and RPS12. By immunohistochemical staining analysis, positive expression rates and strong positive expression rates of annexin A2 in breast cancer of 0, Ⅰ, Ⅱ and Ⅲ TNM clinical stages and the adjacent normal tissues were 75.0%(6/8)/37.5%(3/8), 100.0%(14/14)/42.9%(6/14), 100.0%(10/10)/66.7%(4/6), and 100.0%(6/6)/50.0%(5/10), respectively. There had statistically significant difference between expression positive rate and strong positive rates at different TNM clinical stages in breast cancer (P<0.05). The expression levels of annexin A2 had positive correlation with TNM clinical stages (r=0.508, P<0.05).

Expression of annexin A2 in breast cancer tissues is regulated by ubiquitination, and more later the TNM clinical stages in breast cancer, more higher the expression of annexin A2. Over-expression of annexin A2 regulated by ubiquitination may be involve in breast cancer.