贾建平,孙晓慧
JIA Jianping,SUN Xiaohui

• 1:中国人民解放军北部战区空军医院

摘要(Abstract)：

目的:探讨ECRG4对人喉鳞癌细胞Hep-2侵袭转移能力的影响及机制。方法:设计并构建ECRG4过表达载体、并转染Hep-2细胞,Real-time PCR及Western Blot检测Hep-2细胞中ECRG4 mRNA和蛋白的表达;通过细胞划痕实验检测细胞迁移能力,Transwell实验检测细胞迁移和侵袭能力,应用Western Blot检测NF-κB信号通路相关蛋白NF-κB p65在胞浆和胞核中的表达及转录因子Snail、EMT标志性分子E-cadherin,N-cadherin、Vinmentin蛋白表达。结果:在Hep-2细胞中过表达ECRG4后,ECRG4 mRNA与蛋白表达水平显著升高,划痕和Tranwell实验表明过表达ECRG4可抑制细胞的迁移和侵袭;同时过表达ECRG4可下调细胞核中NF-κB p65蛋白表达,上调细胞浆中NF-κB p65蛋白表达;过表达ECRG4可上调EMT上皮标志物E-cadherin蛋白,下调转录因子Snail及EMT间质标记物Vimentin和N-cadherin蛋白表达。结论:ECRG4可能通过抑制NF-κB/Snail信号通路来抑制EMT的发生,进而抑制Hep-2细胞的侵袭转移。
Objective: To investigate effects of ECRG4 on invasion and metastasis of human laryngeal squamous cell carcinoma Hep-2 and its molecular mechanism. Methods: ECRG4 overexpression vectors were designed and constructed,and transfected into Hep-2 cells. Then the expression of ECRG4 mRNA and protein by Real-time PCR and Western blot were detected. The cell migration through the scratched wound healing assay were detected; the cell migration and invasion were detected by Transwell assay. In addition,the western blot was used to detect the protein expression of NF-κB signaling pathway associated protein NF-κB p65 in cytoplasm and nuclei. The protein expression of transcription factor Snail and EMT( epithelial to mesenchymal transition) molecule markers E-cadherin,N-cadherin,and Vinmentin were also detected. Results: The mRNA and protein expression level of ECRG4 increased significantly after overexpression of ECRG4 in Hep-2 cells. The results of Scratched wound healing assay and Tranwell assay showed that the overexpression of ECRG4 inhibited the cell migration and invasion. At the same time,overexpression of ECRG4 could down-regulate the protein expression of NF-κB p65 in nucleus and up-regulate the protein expression of NF-κB p65 in cytoplasm. In addition,overexpression of ECRG4 could up-regulate the expression of E-cadherin protein of EMT epithelial marker and down-regulate the expression of Vimentin and N-cadherin protein of Snail and EMT interstitial markers. Conclusion: ECRG4 may inhibit the EMT occurrence by inhibiting the activation of NF-κB/Snail signaling pathway,thereby inhibiting the invasion and metastasis of Hep-2 cell.

关键词(KeyWords)： ECRG4;喉肿瘤;癌,鳞状细胞;上皮间质转化;细胞迁移;细胞侵袭;核因子κB
ECRG4;laryngeal neoplasm;carcinoma,squamous cell;epithelial to mesenchymal transition;cell migration;cell invasion;nuclear factor-κB

Abstract:

Keywords:

基金项目(Foundation): 辽宁省自然科学基金指导计划(201602757)

作者(Author): 贾建平,孙晓慧
JIA Jianping,SUN Xiaohui

DOI: 10.19367/j.cnki.1000-2707.2019.02.011

参考文献(References)：

• [1] CHU E A,KIM Y J. Laryngeal cancer:diagnosis andpreoperative work-up[J]. Otolaryngol Clin North Am,2008,41(4):673-695.
• [2]SIEGEL R,NAISHADHAM D,JEMAL A. Cancer statis-tics,2013[J]. Ca A Cancer Journal for Clinicians,2013,63(1):11-15.
• [3] HERMANS R. Staging of laryngeal and hypopharyngealcancer:value of imaging studies[J]. European Radiolo-gy,2007,31(2):2386-2400.
• [4]SABATIER R,FINETTI P,ADELAIDE J,et al. Down-regulation of ECRG4,a candidate tumor suppressor gene,in human breast cancer[J]. Plo S One,2011,6(11):e27656.
• [5]WANG Y B,BA C F. Promoter methylation of esophagealcancer-related gene 4 in gastric cancer tissue and its clini-cal significance[J]. Hepato-gastroenterology,2012,59(118):1696-1698.
• [6]GOTZE S,FELDHAUS V,TRASKA T,et al. ECRG4 isa candidate tumor suppressor gene frequently hypermethy-lated in colorectal carcinoma and glioma[J]. BMC Canc-er,2009,9:447.
• [7]XU T,XIAO D,ZHANG X. ECRG4 inhibits growth andinvasiveness of squamous cell carcinoma of the head andneck in vitro and in vivo[J]. Oncology Letters,2013,5(6):1921-1926.
• [8]GE S,XU Y,WANG H,et al. Downregulation of esoph-ageal cancer-related gene 4 promotes proliferation and mi-gration of hepatocellular carcinoma[J]. Oncology Let-ters,2017,14(3):3689-3696.
• [9]JIA J,DAI S,SUN X,et al. A preliminary study of theeffect of ECRG4 overexpression on the proliferation andapoptosis of human laryngeal cancer cells and the underly-ing mechanisms[J]. Molecular Medicine Reports,2015,12(4):5058-5064.
• [10]贾建平.食管癌相关基因4(ECRG4)在喉癌中的表达和临床意义以及过表达ECRG4对喉癌细胞增殖和凋亡的影响[D].沈阳:中国医科大学,2014.
• [11]PUISIEUX A,BRABLETZ T,CARAMEL J. Oncogenicroles of EMT-inducing transcription factors[J]. NatureCell Biology,2014,16(6):488-494.
• [12]TIWARI N,GHELDOF A,TATARI M,et al. EMT asthe ultimate survival mechanism of cancer cells[J].Seminars in Cancer Biology,2012,22(3):194-207.
• [13]ZHANG K,ZHAOS J,LIU X,et al. Activation of NF-Bupregulates Snail and consequent repression of E-cadherinin cholangiocarcinoma cell invasion[J]. Hepato-gastroen-terology,2011,58(105):1-7.
• [14]WANG X,LI L,GUAN R,et al. Emodin Inhibits ATP-Induced Proliferation and Migration by Suppressing P2YReceptors in Human Lung Adenocarcinoma Cells[J]. In-ternational Journal of Experimental Cellular Physiology,Biochemistry,and Pharmacology,2017,44(4):1337-1351.
• [15]LIN S X,JIANG H,XIANG G Z,et al. Up-regulation oflong non-coding RNA SNHG1 contributes to proliferationand metastasis in laryngeal squamous cell carcinoma[J].European Review for Medical and Pharmacological Sci-ences,2018,22(5):1333-1341.

文章评论(Comment)：