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中华普外科手术学杂志(电子版) ›› 2023, Vol. 17 ›› Issue (02) : 139 -144. doi: 10.3877/cma.j.issn.1674-3946.2023.02.008

论著

基因相似序列家族成员126A靶向调控波形蛋白促进胰腺癌细胞侵袭和迁移及其机制
李永宁1, 付雪芹2, 李英3, 刘鹏1, 刘松柏3, 潘耀振4,†()   
  1. 1. 550001　贵阳，贵州医科大学附属医院肝胆外科；550025　贵阳，贵州医科大学临床医学院
    2. 550001　贵阳，贵州省人民医院乳腺外科
    3. 550025　贵阳，贵州医科大学临床医学院
    4. 550004　贵阳，贵州医科大学附属肿瘤医院肝胆外科
  • 收稿日期:2022-06-09 出版日期:2023-04-26
  • 通信作者: 潘耀振

Gene similar sequence family member 126A targets vimentin to promote invasion and migration of pancreatic cancer cells and its mechanism

Yongning Li1, Xueqin Fu2, Ying Li3, Peng Liu1, Songbai Liu3, Yaozhen Pan4,†()   

  1. 1. Department of Hepatobiliary Surgery, The Affiliated Hospital of Guizhou Medical University, Guiyang Guizhou Province 550001, China; Clinical Medical College of Guizhou Medical University, Guiyang Guizhou Province 550025, China
    2. Department of Breast Surgery, Guizhou Provincial People’s Hospital, Guiyang Guizhou Province 550001, China
    3. Clinical Medical College of Guizhou Medical University, Guiyang Guizhou Province 550025, China
    4. Department of Hepatobiliary Surgery, The Affiliated Cancer Hospital of Guizhou Medical University, Guiyang Guizhou Province 550004, China
  • Received:2022-06-09 Published:2023-04-26
  • Corresponding author: Yaozhen Pan
  • About author:
    Co-first author：Li Yongning；Fu Xueqin
  • Supported by:
    Guizhou Science and Technology Plan Project(QKH［2021］General 100); Guiyang Science and Technology Plan Project(Zhuke Contract［2018］1-86)
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李永宁, 付雪芹, 李英, 刘鹏, 刘松柏, 潘耀振. 基因相似序列家族成员126A靶向调控波形蛋白促进胰腺癌细胞侵袭和迁移及其机制[J]. 中华普外科手术学杂志(电子版), 2023, 17(02): 139-144.

Yongning Li, Xueqin Fu, Ying Li, Peng Liu, Songbai Liu, Yaozhen Pan. Gene similar sequence family member 126A targets vimentin to promote invasion and migration of pancreatic cancer cells and its mechanism[J]. Chinese Journal of Operative Procedures of General Surgery(Electronic Edition), 2023, 17(02): 139-144.

目的

探讨基因相似序列家族成员126A（FAM126A）靶向波形蛋白（Vimentin）对胰腺癌侵袭和迁移能力的影响。

方法

利用基因表达谱交互分析数据库（GEPIA2）和泛癌长期预后和基因表达谱数据库（LOGpc）中的数据，分析FAM126A在胰腺癌和癌旁组织中的表达差异以及对胰腺癌患者生存期的影响；选取2020年1月至2022年1月收集的胰腺导管癌肿瘤组织和对应的癌旁组织样本共16对作为研究对象，采用蛋白印迹法（Western blot）分析胰腺癌组织和癌旁组织中FAM126A表达水平；在胰腺癌PANC-1细胞系构建2个小干扰RNA（siRNA）序列并建立si-FAM126A#1、si-FAM126A#2沉默组和si-con对照组，采用细胞划痕和Transwell实验检测沉默FAM126A后对胰腺癌细胞侵袭和迁移能力的影响；Western blot检测上皮-间充质转化（EMT）相关蛋白E-钙黏蛋白（E-candberin）、波形蛋白（Vimentin）和Snail蛋白的表达量；采用生物信息学和蛋白质谱检测报告分析FAM126A的靶基因，并通过共定位和共沉淀的方法进一步证实FAM126A与Vimentin的相互作用关系。计量数据资料采用均值±标准差（Mean±SD）进行描述，组间比较采用t检验。P<0.05为差异有统计学意义。

结果

在胰腺癌组织中FAM126A表达明显高于癌旁组织（t=12.880，P<0.05）；沉默组si-FAM126A#1、si-FAM126A#2胰腺癌细胞侵袭细胞数（66.11±8.68，91.11±8.68）和迁移率［（22.56±5.36）%，（39.11±6.91）%］较对照组si-con［（146.00±9.15）%，（75.01±10.36）%］显著降低（P<0.05）；si-FAM126A#1和si-FAM126A#2组E-candberin蛋白相对表达量（1.20±0.06，0.97±0.14）较si-con组（0.71±0.08）显著升高（P<0.05），而Vimentin、Snail蛋白表达量为（0.92±0.09，1.01±0.15；0.49±0.07，0.55±0.08）均显著高于对照组（1.68±0.11；1.21±0.08）（P<0.05）；生物信息学和蛋白质谱检测报告提示FAM126A与Vimentin存在相互作用关系，共定位和免疫共沉淀证实了Vimentin是FAM126A的靶蛋白。

结论

在胰腺癌组织中FAM126A的表达高于癌旁组织，而FAM126A可通过靶向调控Vimentin的表达诱导胰腺癌细胞EMT进程，最终促进胰腺癌细胞的侵袭和迁移能力。

关键词: 胰腺肿瘤, 波形蛋白, 肿瘤侵润, 迁移, 上皮-间质转化
Objective

To investigate the effects of 126A（FAM126A）targeting Vimentin on invasion and migration of pancreatic cancer

Methods

The expression difference of FAM126A in pancreatic and paracancer tissues and its effect on survival of pancreatic cancer patients were analyzed using data from the gene expression profile Interactive analysis database（GEPIA2）and the pancarcinoma Long-Term Prognosis and Gene expression profile database（LOGpc）. A total of 16 pairs of pancreatic ductal carcinoma tumor tissues and corresponding paracancer tissue samples collected from January 2020 to January 2022 were selected as research objects. Western blot was used to analyze the expression level of FAM126A in pancreatic and paracancer tissues. Two small interfering RNA（siRNA）sequences were constructed in PANC-1 cell line of pancreatic cancer and si-FAM126A#1，si-FAM126A#2 silencing groups and si-con control group were established. The effects of silencing FAM126A on the invasion and migration of pancreatic cancer cells were detected by cell scratch and Transwell assay. The expression levels of EMT-related proteins E-candberin，Vimentin and Snail were detected by Western blot. The target gene of FAM126A was analyzed by bioinformatics and protein spectrogram reports，and the interaction between FAM126A and Vimentin was further confirmed by co-localization and co-precipitation methods. Measurement data were described by Mean±SD，and t test was used for comparison between groups.

Results

The expression of FAM126A in pancreatic cancer tissues was significantly higher than that in adjacent tissues（t=12.880，P<0.05）. The number of pancreatic cancer cells invaded by si-FAM126A#1 and si-FAM126A#2 groups（66.11±8.68，91.11±8.68）and mobility［（22.56±5.36）%，（39.11±6.91）%］was significantly lower than that of control group si-con［（146.00±9.15）%，（75.01±10.36）%］（P<0.05）. The relative expression level of E-candberin protein in si-FAM126A#1 and si-FAM126A#2 groups was significantly increased（1.20±0.06，0.97±0.14）compared with that in si-con group（0.71±0.08）（P<0.05）. Vimentin Snail protein expression levels were（0.92±0.09，1.01±0.15；0.49±0.07，0.55±0.08）were significantly higher than the control group（1.68±0.11；1.21±0.08）（P<0.05）；Bioinformatics and protein spectrogram reports suggested that there was an interaction between FAM126A and Vimentin. Co-localization and co-immunoprecipitation confirmed that Vimentin was the target protein of FAM126A.

Conclusion

The expression of FAM126A in pancreatic cancer tissues is higher than that in adjacent tissues，and FAM126A can induce the EMT process of pancreatic cancer cells through targeted regulation of Vimentin expression，and finally promote the invasion and migration ability of pancreatic cancer cells.

Key words: Pancreatic Neoplasms, Vimentin, Neoplasm Invasiveness, Metastasis, Epithelial-Mesenchymal Transition
图1 FAM126A在胰腺癌组织中的表达与胰腺癌患者预后的关系注：A=GEPIA2数据集中FAM126A在胰腺癌组织高表达；B= FAM126A对胰腺癌患者预后的影响；C=FAM126A在胰腺癌组织中的表达。
表1 16例胰腺癌患者术前临床特征和实验室指标（
xˉ
±s）
病理类型

年龄

(岁)

BMI

(kg/m2)

 性别 肿瘤分期
男/女 ⅠA ⅠB ⅡA ⅡB
胰腺导管癌 49.4±6.3 23.2±2.8 9/7 1 9 4 2
病理类型 肿瘤大小(cm) 淋巴结转移(例) Ca199(U/ml)
胰腺导管癌 3.9±1.2 2 93.2±54.9
图2 Western blot验证FAM126A对胰腺癌细胞EMT相关标志物的影响
图3 沉默FAM126A后对胰腺癌细胞侵袭和转移能力的影响注：A=通过Transwell验证FAM126A对胰腺癌细胞侵袭能力的影响；B=通过细胞划痕实验验证FAM126A对胰腺癌细胞迁移能力的影响。
图4 通过GEPIA2对FAM126A与Vimentin进行Spearman等级相关性分析
图5 共聚焦显微镜观察FAM126A与Vimentin的共定位情况（×20）注：A=FAM126A；B=Vimentin；C=DAPI染色的细胞核；D=合并图（FAM126A+Vimentin+DAPI）。
图6 免疫共沉淀检测FAM126A与Vimentin的相互作用注：左侧=FAM126A沉淀Vimentin；右侧=Vimentin沉淀FAM126A。
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