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中华妇幼临床医学杂志(电子版) ›› 2016, Vol. 12 ›› Issue (03) : 274 -279. doi: 10.3877/cma.j.issn.1673-5250.2016.03.005

所属专题： 文献；

论著

E6相关蛋白调节膜联蛋白A2的表达对三阴性乳腺癌裸鼠移植瘤的影响
侯令密1, 谢少利2, 陈茂山3, 李敬东4, Ajedichiga Aduah Emmanuel4, 杨宏伟3, 高砚春2, 幸天勇2, 邓世山5, 赵小波2,†()   
  1. 1. 637000　四川南充，川北医学院附属医院甲状腺乳腺外科；肝胆胰肠疾病研究所
    2. 637000　四川南充，川北医学院附属医院甲状腺乳腺外科
    3. 629000　四川省遂宁市中心医院乳腺甲状腺外科
    4. 肝胆胰肠疾病研究所
    5. 637000　四川南充，川北医学院解剖学教研室
  • 收稿日期:2015-11-01 修回日期:2016-05-10 出版日期:2016-06-01
  • 通信作者: 赵小波

Influence of annexin A2 expression regulated by E6-associated protein on trip-negative breast cancer in nude mouse with transplantation tumor

Lingmi Hou1, Shaoli Xie2, Maoshan Chen3, Jingdong Li4, Ajedichiga Aduah Emmanuel4, Hongwei Yang3, Yanchun Gao2, Tianyong Xing2, Shishan Deng5, Xiaobo Zhao2,†()   

  1. 1. Department of Thyroid and Breast Surgery; Institute of Hepatobiliary and Pancreatic-intestinal Diseases, Affiliated Hospital of North Sichuan Medical College, Nanchong 637000, Sichuan Province, China
    2. Department of Thyroid and Breast Surgery
    3. Department of Breast and Thyroid Surgery, Suining Central Hospital, Suining 629000, Sichuan Province, China
    4. Institute of Hepatobiliary and Pancreatic-intestinal Diseases, Affiliated Hospital of North Sichuan Medical College, Nanchong 637000, Sichuan Province, China
    5. Department of Anatomy, North Sichuan Medical College, Nanchong 637000, Sichuan Province, China
  • Received:2015-11-01 Revised:2016-05-10 Published:2016-06-01
  • Corresponding author: Xiaobo Zhao
  • About author:
    Corresponding author: Zhao Xiaobo, Email:
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引用本文：

侯令密, 谢少利, 陈茂山, 李敬东, Ajedichiga Aduah Emmanuel, 杨宏伟, 高砚春, 幸天勇, 邓世山, 赵小波. E6相关蛋白调节膜联蛋白A2的表达对三阴性乳腺癌裸鼠移植瘤的影响[J]. 中华妇幼临床医学杂志(电子版), 2016, 12(03): 274-279.

Lingmi Hou, Shaoli Xie, Maoshan Chen, Jingdong Li, Ajedichiga Aduah Emmanuel, Hongwei Yang, Yanchun Gao, Tianyong Xing, Shishan Deng, Xiaobo Zhao. Influence of annexin A2 expression regulated by E6-associated protein on trip-negative breast cancer in nude mouse with transplantation tumor[J]. Chinese Journal of Obstetrics & Gynecology and Pediatrics(Electronic Edition), 2016, 12(03): 274-279.

目的

探讨E6相关蛋白(E6-AP)调节膜联蛋白A2的表达，对三阴性乳腺癌(TNBC)裸鼠移植瘤的影响。

方法

于2014年4～10月，选择40只雌性BALB/C-nu裸鼠为研究对象。按照随机数字表法，将其分为实验组(n＝20)与对照组(n＝20)。两组TNBC裸鼠移植瘤模型构建：设计针对E6-AP-RNA干扰靶点序列，将小干扰RNA(siRNA)片段转染至TNBC细胞株MDA-MB-231，再将此细胞株皮下注射于实验组裸鼠；而对照组裸鼠则皮下注射等量未转染E6-AP siRNA的TNBC细胞株MDA-MB-231。观察两组裸鼠移植瘤成瘤情况。采用Western blotting法检测两组TNBC裸鼠移植瘤E6-AP及膜联蛋白A2的相对表达量，采用免疫组化法检测两组TNBC裸鼠移植瘤膜联蛋白A2阳性表达率，并对这些指标进行统计学分析。本研究实验过程中对动物处置符合动物伦理学标准。两组裸鼠的周龄及体重等资料比较，差异均无统计学意义(P>0.05)。

结果

①成功构建两组TNBC裸鼠移植瘤模型，成瘤率均为100%。与对照组相比，实验组裸鼠移植瘤生长较慢，成瘤体积较小。实验组裸鼠移植瘤重量为(0.562±0.032)g，显著低于对照组的(1.202±0.012)g，二者比较，差异有统计学意义(t＝96.792，P<0.001)。②Western blotting检测结果显示，实验组E6-AP相对表达量为11 168±441，显著低于对照组的24 284±636，二者比较，差异有统计学意义(t＝75.737，P<0.001)。实验组膜联蛋白A2相对表达量为8 631±651，亦显著低于对照组的17 976±510，二者比较，差异有统计学意义(t＝50.515，P<0.001)。③免疫组化检测结果显示，实验组膜联蛋白A2阳性表达率为33%，明显低于对照组的58%，二者比较，差异有统计学意义(χ2＝5.796，P＝0.016)。实验组TNBC裸鼠移植瘤膜联蛋白A2的积分光密度(IOD)值为17 976±239，显著低于对照组的26 565±329，二者比较，差异亦有统计学意义(t＝94.439，P<0.001)。

结论

在人体内，膜联蛋白A2可能通过E6-AP介导发挥生物学作用。膜联蛋白A2表达降低，可抑制TNBC细胞增殖生长，可能成为TNBC诊断和治疗的新靶标。

关键词: 膜联蛋白A2, E6相关蛋白, 裸鼠移植瘤, 印迹法，蛋白质, 免疫组化
Objective

To study the influence of annexin A2 expression regulated by E6-associated protein (E6-AP) on trip-negative breast cancer (TNBC) in nude mouse with transplantation tumor.

Methods

From April to November 2014, 40 female BALB/C-nu nude mice were selected as research subjects. They were divided into experiment group (n=20) and control group (n=20) according to random number table method. The construction of TNBC nude mice with transplantation tumor models in two groups: E6-AP-RNA interference target sequences were designed, and then small interfering RNA (siRNA) fragments were transfected into TNBC cell lines MDA-MB-231, and these TNBC cell lines were injected subcutaneously into nude mice in experimental group. While TNBC cell lines MDA-MB-231 which were not transfected with E6-AP siRNA were injected subcutaneously into control group. The transplanted tumor formations in two groups were observed. Relative expressions of E6-AP and annexin A2 in two groups were assayed through Western blotting method. And positive expression rate of annexin A2 in transplanted tumor of two groups were assayed by immunohistochemical method. And those indexes above were analyzed by statistical methods. The disposal of animals in this research was in accordance with the animal ethical standards. There were no significant differences between two groups in week of age and body weight (P>0.05).

Results

①The TNBC nude mice with transplanted tumor models were constructed successfully in both two groups, and the tumor formation rates both were 100%. The growth of transplantation tumor in experimental group was relatively slower and the tumor volume was smaller than those of control group. The weight of transplantation tumor in experimental group was (0.562±0.032) g, which was significantly heavier than that of (1.202±0.012) g in control group, and the difference was statistical significant (t=96.792, P<0.001). ②According to the results of Western blotting assay, the relative expression of E6-AP in experimental group was 11 168±441, which was significantly lower than that of (24 284±636) in control group, and the difference was statistical significant (t=75.737, P<0.001). However, the relative expressions of annexin A2 in experimental group and control group were 8 631±651 and 17 976±510, respectively, and the difference was statistical significant (t=50.515, P<0.001). ③Immunohistochemical assay results indicated that the positive expression rate of annexin A2 of experimental group (33%) was significantly lower than that of control group (58%), and the difference was statistically significant (χ2=5.796, P=0.016). The integral optical density (IOD) value of annexin A2 of experimental group (17 976±239) was notably lower than that of control group (26 565±329), and the difference was statistically significant (t=94.439, P<0.001).

Conclusions

Annexin A2 could inhibit biological behavior of breast cancer in vivo maybe through the mediation of E6-AP. If the expression of annexin A2 decreases, it can inhibit the proliferation and growth of breast cancer cells which may become the new target of diagnosis and treatment for TNBC.

Key words: Annexin A2, E6-associated protein, Transplanted tumor in nude mice, Blotting, Western, Immunohistochemisty
图4 免疫组化法检测两组三阴性乳腺癌裸鼠移植瘤组织膜联蛋白A2表达情况(图4A：对照组；图4B：实验组)(SP染色，高倍镜)
1
Dhillon S. Trastuzumab emtansine: a review of its use in patients with HER2-positive advanced breast cancer previously treated with trastuzumab-based therapy[J]. Drugs, 2014, 74(6):675-686.
2
Hagan CR, Lange CA. Molecular determinants of context-dependent progesterone receptor action in breast cancer[J]. BMC Med, 2014, 12:32.
3
幸天勇，侯令密，邓世山．泛素化调节膜联蛋白A2表达异常在乳腺癌中的作用研究[J/CD]．中华妇幼临床医学杂志：电子版，2013，9(2)：130-134.
4
侯令密，幸天勇，邓世山．Annexin A2与乳腺癌关系的研究进展[J/CD]．中华临床医师杂志：电子版，2011，5(14)：4206-4208.
5
幸天勇，侯令密，汪建国，等．膜联蛋白A2在人乳腺癌组织中的差异表达及意义[J/CD]．中华临床医师杂志：电子版，2011，5(18)：5310-5313.
6
Mirnezami R, Nicholson J, Darzi A. Preparing for precision medicine[J]. N Engl J Med, 2012, 366(6):489-491.
7
Mok TS, Wu YL, Thongprasert S, et al. Gefitinib or carboplatin-paclitaxel in pulmonary adenocarcinoma[J]. N Engl J Med, 2009, 361(10):947-957.
8
Knutson TP, Lange CA. Tracking progesterone receptor-mediated actions in breast cancer[J]. Pharmacol Ther, 2014, 142(1):114-125.
9
Sachdev D. Targeting the type Ⅰ insulin-like growth factor system for breast cancer therapy[J]. Curr Drug Targets, 2010, 11(9):1121-1132.
10
Shimizu A, Nakayama H, Wang P, et al. Netrin-1 promotes glioblastoma cell invasiveness and angiogenesis by multiple pathways including activation of RhoA, cathepsin B, and cAMP-response element-binding protein[J]. J Biol Chem, 2013, 288(4):2210-2222.
11
Zhang Q, Zhao Z, Ma Y, et al. Combined expression of S100A4 and Annexin A2 predicts disease progression and overall survival in patients with urothelial carcinoma[J]. Urol Oncol, 2014, 32(6):798-805.
12
Bydoun M, Waisman DM. On the contribution of S100A10 and annexin A2 to plasminogen activation and oncogenesis: an enduring ambiguity[J]. Future Oncol, 2014, 10(15):2469-2479.
13
Deng S, Zhou H, Xiong R, et al. Over-expression of genes and proteins of ubiquitin specific peptidases (USPs) and proteasome subunits (PSs) in breast cancer tissue observed by the methods of RFDD-PCR and proteomics[J]. Breast Cancer Res Treat, 2007, 104(1):21-30.
14
Zhang PW, Chen L, Huang T, et al. Classifying ten types of major cancers based on reverse phase protein array profiles[J]. PLoS One, 2015, 10(3):e0123147.
15
Cox E, Uzoma I, Guzzo C, et al. Identification of SUMO E3 ligase-specific substrates using the HuProt human proteome microarray[J]. Methods Mol Biol, 2015, 1295:455-463.
16
Smith SE, Koegl M, Jentsch S. Role of the ubiquitin/proteasome system in regulated protein degradation in Saccharomyces cerevisiae[J]. Biol Chem, 1996, 377(7-8):437-446.
17
Murakami Y, Matsufuji S, Kameji T, et al. Ornithine decarboxylase is degraded by the 26S proteasome without ubiquitination[J]. Nature, 1992, 360(6404):597-599.
18
Bird LM. Angelman syndrome: review of clinical and molecular aspects[J]. Appl Clin Genet, 2014, 7:93-104.
19
Słopień A, Rajewski A. Genetic studies in autistic disorders [J]. Psychiatr Pol, 2000, 34(3):435-446.
20
Zheng L, Foley K, Huang L, et al. Tyrosine 23 phosphorylation-dependent cell-surface localization of annexin A2 is required for invasion and metastases of pancreatic cancer[J]. PLoS One, 2011, 6(4):e19390.
21
Gao S, Fang L, Phan LM, et al. COP9 signalosome subunit 6 (CSN6) regulates E6AP/UBE3A in cervical cancer[J]. Oncotarget, 2015, 6(29):28026-28041.
22
Malecka KA, Fera D, Schultz DC, et al. Identification and characterization of small molecule human papillomavirus E6 inhibitors[J]. ACS Chem Biol, 2014, 9(7):1603-1612.
23
Santarosa M, Del Col L, Viel A, et al. BRCA1 modulates the expression of hnRNPA2B1 and KHSRP[J]. Cell Cycle, 2010, 9(23):4666-4673.
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