Lentivirus-mediated siRNA Targeting Cyclooxygenase-2 Gene Inhibits Human Laryngocarcinoma Cells Proliferation and Invasion_West China Medical Journal

Authors：

WANG Xiaoxia ^1,2 ,  YAO Xiaobao ² , ZHANG Shaoqiang. ¹

1. Department of Otolaryngology, the First Affiliated Hospital of Medical College, Xi’an Jiaotong University, Xi’an, Shaanxi 710061, P. R. China; 2. Department of Otolaryngology, 451 Hospital of PLA, Xi’an, Shaanxi 710054, P. R. China;

Corresponding?author：

YAO Xiaobao, Email: xiaoxiawang9139@sina.com.cn

Keywords：

喉癌; 環氧化酶-2基因; 慢病毒; 基因沉默

DOI：

10.7507/1002-0179.20130063

Video：

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目的　構建沉默環氧化酶-2（COX-2）基因重組慢病毒，觀察其體外侵襲的抑制作用，從而探討干擾COX-2抑制喉癌細胞增殖的作用機理，為喉癌的治療提供新的思路。方法　逆轉錄聚合酶鏈反應（RT-PCR）檢測COX-2基因在人表皮樣喉癌細胞（Hep-2）中的表達情況。利用上海吉凱公司RNA干擾（RNAi）慢病毒表達載體系統，構建針對COX-2基因慢病毒RNAi表達載體。轉染Hep-2細胞，干擾COX-2基因的表達，實時定量PCR檢測干擾前后基因表達變化。利用生長曲線測定干擾載體轉染前后細胞生長速度變化。流式細胞儀檢測細胞的生長周期。Boyden侵襲小室法測定體外侵襲力。結果　成功構建了COX-2慢病毒RNAi表達載體，并建立了干擾COX-2基因的Hep-2細胞系。實時定量PCR檢測COX-2基因在Hep-2細胞系中過表達被顯著抑制。生長曲線測定，COX-2基因干擾后細胞增殖明顯變慢。流式細胞儀檢測細胞的生長周期可見干擾組誘導Hep-2細胞凋亡，轉染G0～G1期細胞數量明顯上升，S期細胞減少，表明siRNA干擾Hep-2細胞后，細胞由G0～G1期進入到S期受到阻滯，細胞增殖速度下降。體外侵襲實驗中，Hep-2-AS侵襲細胞數（31.0 ± 1.8）顯著低于Hep-2細胞（104.0 ± 2.6）及Hep-2-P細胞（99.0 ± 2.7），差異有統計學意義（P＜0.05）。結論　喉癌中過表達的COX-2基因被干擾后表達明顯降低并顯著抑制細胞的生長速度和侵襲能力。同時驗證了COX-2基因RNA干擾在進行抗腫瘤的治療中潛在的應用前景。

Citation： WANG Xiaoxia,YAO Xiaobao,ZHANG Shaoqiang.. Lentivirus-mediated siRNA Targeting Cyclooxygenase-2 Gene Inhibits Human Laryngocarcinoma Cells Proliferation and Invasion. West China Medical Journal, 2013, 28(2): 195-199. doi: 10.7507/1002-0179.20130063 Copy

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1. 　Das D, Arber N, Jankowski JA. Chemoprevention of colorectal cancer[J]. Digestion, 2007, 76(1): 51-67.
2. 　Wang MT, Honn KV, Nie D. Cyclooxygenases, prostanoids, and tumor progression[J]. Cancer Metastasis Rev, 2007, 26(3-4): 525-534.
3. 　姚小寶, 王曉俠, 張少強, 等. 抑制COX-2表達對逆轉喉癌惡性生物學行為的影響[J]. 西安交通大學學報(醫學版), 2009, 19(7): 671-673.
4. 　楊健, 胡為民, 任碧軒, 等. RNA干擾體外抑制HBeAg的表達[J]. 細胞與分子免疫學雜志, 2006, 22(5): 670-671, 673.
5. 　Brummelkamp TR, Bernards R, Agami R. Stable suppression of tumorigenicity by virus-mediated RNA interference[J]. Cancer Cell, 2002, 2(3): 243-247.
6. 　Feng J, Funk WD, Wang SS, et al. The RNA component of human telomerase[J]. Science, 1995, 269(5228): 1236-1241.
7. 　Gwyn K, Sinicrope F. Chemoprevention of colorectal cancer[J]. Am J Gastroenterol, 2002, 97(1): 13-21.
8. 　王磊, 陳衛昌, 謝學順, 等. 塞來昔布對實驗性結腸癌原位移植瘤生長及血管形成的影響[J]. 中華腫瘤防治雜志, 2006, 13(17): 1295-1300.
9. 　Chen Z, Zhang X, Li M, et al. Simultaneously targeting epidermal growth factor receptor tyrosine kinase and cyclooxyg-enase-2, an efficient approach to inhibition of squamous cell carcinoma of the head and neck[J]. Clin Cancer Res, 2004, 10(17): 5930-5939.
10. 　王晨, 費小凡. 饒丹, 等. 環氧化酶-2抑制劑致心血管系統的作用機制[J]. 華西醫學, 2011, 26(10): 1581-1584.
11. 　Brummelkamp TR, Bernards R. New tools for functional mammalian cancer genetics[J]. Nat Rev Cancer, 2003, 10(10): 781-789.
12. 　Barańska M, Skretkowicz J. Prospects of gene therapy[J]. Wiad Lek, 2007, 60(7-8): 305-311.
13. 　達明緒, 張明鳴, 羅婷, 等. 胃癌COX-2的表達與微血管密度及相關臨床病理因素的關系[J]. 華西醫學, 2005, 20(1): 12-14.
14. 　宋繼榮, 董穎, 潘月娜. 宮頸癌組織中Survivin和Cox-2的表達及相關性[J]. 黑龍江醫藥科學, 2012, 35(5): 36-38.
15. 　Masferrer JL, Leahy KM, Koki AT, et al. Antiangiogenic and antitumor activities of cyclooxygenase-2 inhibitors[J]. Cancer Res, 2000, 60(5): 1306-1311.
16. 　Gallo O, Franchi A, Magnelli L, et al. Cyclooxygenase-2 pathway correlates with VEGF expression in head and neck cancer. Implications for tumor angiogenesis and metastasis[J]. Neoplasia, 2001, 3(1): 53-61.
17. 　趙暉, 王樹成. RNA干擾沉默食管癌COX-2基因的實驗研究[J]. 山東大學學報:醫學版, 2006, 44(3): 256-259.
18. 　Suzuki O, Abe M. Cell surface N-glycosylation and sialylation regulate galectin-3-induced apoptosis in human diffuse large B cell lymphoma[J]. Oncol Rep, 2008, 19(3): 743-748.
19. 　Takai N, Ueda T, Nishida M, et al. Histone deacetylase inhibitors induce growth inhibition, cell cycle arrest and apoptosis in human choriocarcinoma cells[J]. Int J Mol Med, 2008, 21(1): 109-115.
20. 　Folkman J. Angiogenesis and apoptosis[J]. Cancer Biol, 2003, 13(2): 159-167.

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Lentivirus-mediated siRNA Targeting Cyclooxygenase-2 Gene Inhibits Human Laryngocarcinoma Cells Proliferation and Invasion

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