四次跨膜蛋白1在乳腺癌中的表达及其促进乳腺癌进展的作用机制

doi:10.3969/j.issn.1674-8115.2023.03.004

摘要/Abstract

摘要：

目的·探究四次跨膜蛋白1（tetraspanin 1，TSPAN1）在乳腺癌中的表达，及其对乳腺癌细胞迁移、侵袭能力的影响。方法·使用免疫组织化学染色（immunohistochemistry staining，IHC）检测106例乳腺癌患者的癌组织及癌旁组织中TSPAN1的表达，并分析其与患者临床特征的相关性。利用癌症基因组图谱（The Cancer Genome Atlas，TCGA）数据库分析TSPAN1 mRNA在乳腺癌及癌旁组织中的表达，及其与患者临床特征的相关性。在乳腺癌细胞MDA-MB-231、SUM159PT中下调TSPAN1后，通过细胞划痕实验、细胞侵袭实验检测TSPAN1对上述细胞迁移、侵袭的影响，并采用蛋白质印迹法（Western blotting）检测细胞上皮-间质转化（epithelial-mesenchymal transition，EMT）相关蛋白（E-钙黏蛋白、N-钙黏蛋白和波形蛋白）的表达。结果·IHC的结果显示，TSPAN1在乳腺癌组织中的表达高于癌旁组织（P=0.000），且其表达水平与肿瘤的总TNM分期、M分期、N分期和病理学分级显著相关（均P<0.05）。对来自TCGA数据库的转录组测序数据分析的结果显示，TSPAN1 mRNA在乳腺癌组织中的表达高于癌旁组织（P=0.000），其在TNM Ⅲ~Ⅳ期的乳腺癌组织中的表达高于TNMⅠ~Ⅱ期（P=0.007）。与转染Control siRNA相比，转染TSPAN1 siRNA的MDA-MB-231和SUM159PT细胞的迁移、侵袭能力均有所下降，E-钙黏蛋白的表达增加、N-钙黏蛋白及波形蛋白的表达减少（均P<0.05）。结论·TSPAN1在乳腺癌组织中表达较高，可促进乳腺癌细胞的迁移和侵袭。

关键词: 乳腺癌, 四次跨膜蛋白1, 上皮-间质转化

Abstract:

Objective ·To investigate the expression of tetraspanin 1 (TSPAN1) in breast cancer and its effect on the migration and invasion of breast cancer cells. Methods ·Immunohistochemistry staining (IHC) was used to detect the expression of TSPAN1 in breast cancer tissues and para-tumor tissues from 106 clinical patients, and to analyze its correlation with the clinical characteristics of patients. The Cancer Genome Atlas (TCGA) database was used to analyze the expression of TSPAN1 mRNA in breast cancer tissues and para-tumor tissues, and its correlation with the clinical characteristics of patients. After down-regulation of TSPAN1 in breast cancer cells MDA-MB-231 and SUM159PT, the effects of TSPAN1 on migration and invasion of cells were detected by wound healing assay and transwell assay, and the expression of epithelial-mesenchymal transition (EMT)-related proteins (E-cadherin, N-cadherin and vimentin) were detected by Western blotting. Results ·IHC results showed that the expression of TSPAN1 in breast cancer tissues was higher than that in para-tumor tissues (P=0.000), and the expression of TSPAN1 in total TNM stages, M stages, N stages and pathological grades of breast cancer tissues showed statistically significant differences (all P<0.05). The results of transcriptome sequencing data from TCGA database showed that the expression of TSPAN1 mRNA in breast cancer tissues was higher than that in para-tumor tissues (P=0.000), and its expression in TNM Ⅲ?Ⅳ stages was higher than that in TNMⅠ?Ⅱ stages (P=0.007). After down-regulation of TSPAN1, the migration and invasion ability of MDA-MB-231 and SUM159PT cells were decreased, the expression of E-cadherin was increased and the expressions of N-cadherin and vimentin were decreased (all P<0.05). Conclusion ·TSPAN1 is highly expressed in breast cancer tissues, which can promote the migration and invasion of breast cancer cells.

Key words: breast cancer, tetraspanin 1 (TSPAN1), epithelial-mesenchymal transition (EMT)

中图分类号:

R737.9

曹源, 王红霞, 朱灜, 李军建. 四次跨膜蛋白1在乳腺癌中的表达及其促进乳腺癌进展的作用机制[J]. 上海交通大学学报（医学版）, 2023, 43(3): 293-300.

CAO Yuan, WANG Hongxia, ZHU Ying, LI Junjian. Expression of tetraspanin 1 in breast cancer and its mechanism in promoting the progression of breast cancer[J]. Journal of Shanghai Jiao Tong University (Medical Science), 2023, 43(3): 293-300.

图/表 5

表1 siRNA 序列

Tab 1 Sequences of siRNA

siRNA	Forward (5′→3′)	Reverse (5′→3′)
Control siRNA	UUCUCCGAACGUGUCACGUTT	ACGUGACACGUUCGGAGAATT
TSPAN1 siRNA 1	CGUUUCGUCAAAGAGAAUATT	UAUUCUCUUUGACGAAACGTT
TSPAN1 siRNA 2	GGCUCACGACCAAAAAGUATT	UACUUUUUGGUCGUGAGCCTT

图1 TSPAN1在乳腺癌组织及癌旁组织中的表达及其与患者临床特征的关系Note： A. Detection of TSPAN1 expression in breast cancer tissues and para-tumor tissues by IHC. B. Statistical analysis of TSPAN1 expression in breast cancer tissues and para-tumor tissues. C. Statistical analysis of TSPAN1 expression in breast cancer tissues of TNM stages. D. Statistical analysis of TSPAN1 expression in breast cancer tissues of M stages. E. Statistical analysis of TSPAN1 expression in breast cancer tissues of N stages. F. Statistical analysis of TSPAN1 expression in breast cancer tissues of different pathological grades. Due to missing data of some cases, the total case number in fig C and F is less than 106 cases.

Fig 1 Expression of TSPAN1 in breast cancer tissues and para-tumor tissues and its relationship with clinical characteristics of patients

图2 TCGA数据库中 TSPAN1 mRNA在乳腺癌组织及癌旁组织中的表达及其与患者临床特征的相关性Note： A. Relative expression of TSPAN1 mRNA in breast cancer tissues and para-tumor tissues in TCGA database. B. Relative expression of TSPAN1 mRNA in breast cancer tissues of TNM stages in TCGA database. C. Relative expression of TSPAN1 mRNA in breast cancer tissues of N0-N3 stages. In TCGA database, due to missing data of some cases, the total case number in fig B and C is less than 1 101 cases.

Fig 2 Expression of TSPAN1 mRNA in breast cancer tissues and para-tumor tissues and its correlation with patients' clinical characteristics in TCGA database

图3 沉默 TSPAN1 对MDA-MB-231、SUM159PT细胞迁移、侵袭能力的影响Note： A. Detection of the expression level of TSPAN1 in different breast cancer cells by Western blotting. B. Detection of the expression level of TSPAN1 in MDA-MB-231 and SUM159PT cells transfected with Control siRNA or TSPAN1 siRNA by Western blotting. C/D. Effects of TSPAN1 siRNA on the migration (C) and invasion (D) ability of MDA-MB-231 and SUM159PT cells.

Fig 3 Effects of silencing TSPAN1 on the migration and invasion ability of MDA-MB-231 and SUM159PT cells

图4 沉默 TSPAN1 后MDA-MB-231、SUM159PT细胞中EMT相关蛋白的表达Note： A. MDA-MB-231 cells. B. SUM159PT cells.

Fig 4 Detection of EMT-related proteins expression in MDA-MB-231 and SUM159PT cells after silencing TSPAN1

参考文献 15

1	SIEGEL R L, MILLER K D, JEMAL A. Cancer statistics, 2015[J]. CA Cancer J Clin, 2015, 65(1):5-29.
2	STEEG P S. Targeting metastasis[J]. Nat Rev Cancer, 2016, 16(4): 201-218.
3	ZHU R X, GIRES O, ZHU L Q, et al. TSPAN8 promotes cancer cell stemness via activation of sonic Hedgehog signaling[J]. Nat Commun, 2019, 10(1): 2863.
4	HUANG Y W, LI J J, DU W Q, et al. Nuclear translocation of the 4-pass transmembrane protein Tspan8[J]. Cell Res, 2021, 31(11): 1218-1221.
5	LU X Q, AN L W, FAN G J, et al. EGFR signaling promotes nuclear translocation of plasma membrane protein TSPAN8 to enhance tumor progression via STAT3-mediated transcription[J]. Cell Res, 2022, 32(4): 359-374.
6	HEMLER M E. Tetraspanin proteins mediate cellular penetration, invasion, and fusion events and define a novel type of membrane microdomain[J]. Annu Rev Cell Dev Biol, 2003, 19: 397-422.
7	CHEN L, YUAN D Y, WANG G L, et al. Clinicopathological significance of expression of Tspan-1, Jab1 and p27 in human hepatocellular carcinoma[J]. J Korean Med Sci, 2010, 25(10): 1438-1442.
8	WANG S S, LIU X H, KHAN A A, et al. miR-216a-mediated upregulation of TSPAN1 contributes to pancreatic cancer progression via transcriptional regulation of ITGA2[J]. Am J Cancer Res, 2020, 10(4): 1115-1129.
9	CHEN L, ZHU Y Y, ZHANG X J, et al. TSPAN1 protein expression: a significant prognostic indicator for patients with colorectal adenocarcinoma[J]. World J Gastroenterol, 2009, 15(18): 2270-2276.
10	SCHOLZ C J, KURZEDER C, KORETZ K, et al. Tspan-1 is a tetraspanin preferentially expressed by mucinous and endometrioid subtypes of human ovarian carcinomas[J]. Cancer Lett, 2009, 275(2): 198-203.
11	WU Y G, CHEN W X, GONG Y F, et al. Tetraspanin 1 (TSPAN1) promotes growth and transferation of breast cancer cells via mediating PI3K/Akt pathway[J]. Bioengineered, 2021, 12(2): 10761-10770.
12	MENKE C H, POHLMANN P R, BACKES A, et al. Tumor size as a surrogate end point for the detection of early breast cancer: a 30-year (1972‒2002), single-center experience in southern Brazil[J]. Breast J, 2007, 13(5): 448-456.
13	LOIBL S, POORTMANS P, MORROW M, et al. Breast cancer[J]. Lancet, 2021, 397(10286): 1750-1769.
14	ESTEVA F J, HUBBARD-LUCEY V M, TANG J, et al. Immunotherapy and targeted therapy combinations in metastatic breast cancer[J]. Lancet Oncol, 2019, 20(3): e175-e186.
15	LI J J, CHEN X L, ZHU L Q, et al. SOX9 is a critical regulator of TSPAN8-mediated metastasis in pancreatic cancer[J]. Oncogene, 2021, 40(30): 4884-4893.

[1]	杨笑萱, 朱珊, 钱程, 储晓英. 术中使用小剂量右美托咪定对乳腺癌手术患者预后的影响[J]. 上海交通大学学报（医学版）, 2023, 43(2): 194-200.
[2]	夏坤健, 邓林林, 王琳. 乳腺癌化学治疗致肝损伤预测模型的构建及其评价[J]. 上海交通大学学报（医学版）, 2022, 42(4): 502-509.
[3]	张郁瑭, 金奕婕, 张凤春, 徐迎春. 乳腺癌合并新型冠状病毒肺炎感染患者的抗肿瘤合理化诊疗方案探讨[J]. 上海交通大学学报（医学版）, 2022, 42(12): 1745-1750.
[4]	苏俊澄, 王禹铮, 唐雷, 徐迎春, 张凤春. 乳腺癌脑转移患者的临床病理特征及预后的影响因素分析[J]. 上海交通大学学报（医学版）, 2022, 42(11): 1562-1568.
[5]	孙亚蒙, 马晔, 郭润生. 乳腺癌组织样本中 HER2基因拷贝数的检测分析[J]. 上海交通大学学报（医学版）, 2022, 42(11): 1589-1597.
[6]	王禹铮, 苏俊澄, 唐雷, 徐迎春, 张凤春. 单周紫杉醇联合顺铂一线治疗转移性乳腺癌的效果和安全性的回顾性研究[J]. 上海交通大学学报（医学版）, 2022, 42(10): 1420-1427.
[7]	陈翠, 金叶, 王琳, 李红丽, 万财凤, 姜立新. 30例乳腺化生性癌的多种影像学对比分析[J]. 上海交通大学学报(医学版), 2022, 42(1): 70-76.
[8]	周天浩, 辛肇晨, 杜少倩, 曹源, 许静轩, 劳曾红, 王红霞. 乳腺癌类器官共培养技术的建立和优化[J]. 上海交通大学学报(医学版), 2021, 41(8): 1017-1024.
[9]	季艳杰, 罗浩, 蔡海燕, 刘欣宇, 金诗佳, 粟深月, 徐含章, 雷虎, 吴英理. CDDO-Me对三阴性乳腺癌细胞泛素特异性蛋白酶2a活性及细胞增殖的抑制作用[J]. 上海交通大学学报(医学版), 2021, 41(8): 1025-1032.
[10]	王成志, 邓华云, 庞智, 黄雷. MUC1在HER2阳性乳腺癌发病中的作用及机制研究[J]. 上海交通大学学报(医学版), 2021, 41(7): 839-848.
[11]	张海燕, 储晓英. 非阿片类镇痛药应用在全身麻醉喉罩置入的乳腺癌保乳手术中的可行性[J]. 上海交通大学学报(医学版), 2021, 41(5): 637-641.
[12]	张佳玲, 张凤春, 徐迎春. 乳腺癌脑转移系统治疗的研究进展[J]. 上海交通大学学报(医学版), 2021, 41(5): 671-677.
[13]	郝艳云, 俞思慧, 陆静, 顾湘, 张帆, 程金科, 王田实. SIRT3去SUMO化修饰调节乳腺癌细胞MCF7增殖及化疗药物敏感性的研究[J]. 上海交通大学学报(医学版), 2021, 41(12): 1557-1563.
[14]	甘露, 金玉翡, 任清, 董晓晶, 张男. 乳房重建术后乳腺癌患者性生活及情感体验的质性研究[J]. 上海交通大学学报(医学版), 2021, 41(12): 1597-1601.
[15]	柴烨子, 马珺, 江雯珑, 刘启明, 卢启帆, 陶政宇, 姜萌, 卜军. 乳腺癌患者新辅助化疗早期的心肺功能和相关影响因素研究[J]. 上海交通大学学报(医学版), 2021, 41(12): 1643-1648.