敲减TULP3对头颈鳞癌细胞生物学行为的影响

doi:10.13591/j.cnki.kqyx.2022.06.003

口腔医学 ›› 2022, Vol. 42 ›› Issue (6): 494-500.doi: 10.13591/j.cnki.kqyx.2022.06.003

敲减TULP3对头颈鳞癌细胞生物学行为的影响

江丰^1,2, 何垚^1,2, 朱载瓯², 丁旭², 叶金海^1,2, 吴煜农^1,2, 宋晓萌^1,2

1 南京医科大学口腔疾病研究江苏省重点实验室,江苏南京(210029);
2 南京医科大学附属口腔医院口腔颌面外科,江苏南京(210029)

修回日期:2022-03-18 出版日期:2022-06-28 发布日期:2022-07-21
通讯作者: 宋晓萌 E-mail:Xiaomengsong@njmu.edu.cn
基金资助:
国家自然科学基金(81772877);江苏省医学创新团队资助项目(CXTDA2017036);江苏省高校优势学科建设工程(2018-87)

Effects of knocking down TULP3 on biological behaviors of head and neck squamous cell carcinoma cells

JIANG Feng, HE Yao, ZHU Zaiou, DING Xu, YE Jinhai, WU Yunong, SONG Xiaomeng

Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University,Nanjing 210029,China

Revised:2022-03-18 Online:2022-06-28 Published:2022-07-21

摘要/Abstract

摘要： 目的探讨敲减TULP3对头颈鳞状细胞癌(HNSCC)细胞生物学行为的影响。方法应用TCGA数据比较头颈鳞癌组织与癌旁组织中TULP3表达水平;体外培养人HNSCC细胞系HN4、HN6、CAL27、HSC3、SCC4及正常口腔上皮角质细胞HOK并应用Western Blot比较TULP3蛋白表达水平,免疫组化分析TULP3在HNSCC组织中表达情况。构建RNA干扰寡核苷酸si-TULP3及si-NC转染HN4、HN6,应用CCK-8实验、平板克隆形成实验、划痕愈合实验、Transwell侵袭实验检测敲减TULP3对HNSCC细胞增殖、侵袭、迁移能力的影响。实时定量逆转录PCR及Western Blot检测细胞周期及上皮间质转化(EMT)相关指标变化。构建HN6-shTULP3及HN6-shNC细胞株接种于裸鼠皮下,分析裸鼠移植瘤体积差异。结果 TCGA数据显示头颈鳞癌组织TULP3表达量显著高于癌旁组织 (P＜0.000 1) ;HN4、HN6、CAL27、HSC3细胞中TULP3蛋白表达量均高于HOK细胞,TULP3在HNSCC组织中呈阳性表达。HN4、HN6细胞转染si-TULP3后,增殖、侵袭、迁移能力明显弱于si-NC组。敲减TULP3后,HN4、HN6细胞CDK4、Cyclin D1、Cyclin D3、N-cadherin、ZEB2、Slug蛋白表达水平下降,其中CDK4、Slug基因水平也下降 (P＜0.001) ;此外,β-catenin蛋白表达水平上升。敲减TULP3后裸鼠皮下移植瘤体积明显小于对照组(P＜0.01)。结论敲减TULP3可以抑制HNSCC细胞系的增殖、侵袭及迁移能力,可能通过调控细胞周期及抑制EMT进程发挥作用。

关键词: TULP3, 头颈鳞状细胞癌, 上皮间质转化, 细胞周期

Abstract: Objective To investigate the effect of knocking down TULP3 on the biological behavior of head and neck squamous cell carcinoma (HNSCC) cells. Methods Differential expression analysis of TULP3 between HNSCC and pericarcinous tissues were conducted according to TCGA data. Human normal oral keratinocytes HOK and HNSCC cell lines HN4, HN6, CAL27, HSC3, SCC4 were cultured in vitro, and the protein levels of TULP3 were analyzed by Western Blot. Expression of TULP3 in HNSCC tissues was analyzed by immunohistochemistry. RNA interference was applied in constructed si-TULP3 and si-NC, and HN4 and HN6 were transfected. Effects of TUKP3 knockdown on proliferation, invasion and migration of HN4 and HN6 cells were investigated by CCK-8 assay, plate colony formation assay, scratch assay and Transwell invasion assay. Real-time qRT-PCR and Western Blot were used to detect the expression of cell cycle-related and EMT-related markers. HN6-shTULP3 and HN6-shNC were subcutaneously injected into nude mice to compare the volume of the xenografted tumor. Results TCGA data revealed that TULP3 expression was significantly higher in HNSCC tissues than pericarcinous tissues (P＜0.000 1). The expression of TULP3 protein in HN4, HN6, CAL27, HSC3 were significantly up-regulated compared with HOK. TULP3 was positively expressed in HNSCC tissues. Meanwhile, the proliferation, invasion and migration capacities of HN4 and HN6 cells were attenuated in HN4 and HN6 cells transfected with si-TULP3. The protein expressions of CDK4, Cyclin D1, Cyclin D3, N-cadherin, ZEB2, Slug were decreased after knockdown of TULP3 in HN4 and HN6 cells, as well as the mRNA expression of CDK4 and Slug (P＜0.001). Furthermore, the up-regulated expression of β-catenin protein was observed. The volume of xenograft tumor formed by HN6-shTULP3 was significantly lower than that in the control group (P＜0.01). Conclusion Knockdown of TULP3 can significantly inhibit the proliferation, invasion and migration of head and neck squamous cells, which may be achieved by regulating cell cycle and suppressing EMT process.

Key words: TULP3, head and neck squamous cell carcinoma, epithelial-mesenchymal transition, cell cycle

中图分类号:

R739.85

江丰, 何垚, 朱载瓯, 丁旭, 叶金海, 吴煜农, 宋晓萌. 敲减TULP3对头颈鳞癌细胞生物学行为的影响[J]. 口腔医学, 2022, 42(6): 494-500.

JIANG Feng, HE Yao, ZHU Zaiou, DING Xu, YE Jinhai, WU Yunong, SONG Xiaomeng. Effects of knocking down TULP3 on biological behaviors of head and neck squamous cell carcinoma cells[J]. Stomatology, 2022, 42(6): 494-500.

参考文献

[1] Sung H, Ferlay J, Siegel RL, et al. Global cancer statistics 2020:GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries[J]. CA Cancer J Clin, 2021, 71(3):209-249.
[2] Muzaffar J, Bari S, Kirtane K, et al. Recent advances and future directions in clinical management of head and neck squamous cell carcinoma[J]. Cancers, 2021, 13(2):338.
[3] Xing DT, Khor R, Gan H, et al. Recent research on combination of radiotherapy with targeted therapy or immunotherapy in head and neck squamous cell carcinoma:A review for radiation oncologists[J]. Cancers, 2021, 13(22):5716.
[4] Leemans CR, Snijders PJF, Brakenhoff RH. The molecular landscape of head and neck cancer[J]. Nat Rev Cancer, 2018, 18(5):269-282.
[5] Bos T, Ratti JA, Harada H. Targeting stress-response pathways and therapeutic resistance in head and neck cancer[J]. Front Oral Health, 2021, 2:676643.
[6] Kitamura N,Sento S, Yoshizawa Y, et al. Current trends and future prospects of molecular targeted therapy in head and neck squamous cell carcinoma[J]. Int J Mol Sci, 2020, 22(1):240.
[7] Wang M,Xu ZC, Kong YZ. The tubby-like proteins kingdom in animals and plants[J]. Gene, 2018, 642:16-25.
[8] Nishina PM, North MA, Ikeda A, et al. Molecular characteriza-tion of a novel tubby gene family member, TULP3, in mouse and humans[J]. Genomics, 1998, 54(2):215-220.
[9] Patterson VL, Damrau C, Paudyal A, et al. Mouse hitchhiker mutants have spina bifida, dorso-ventral patterning defects and polydactyly:Identification of Tulp3 as a novel negative regulator of the Sonic hedgehog pathway[J]. Hum Mol Genet, 2009, 18(10):1719-1739.
[10] Hirano T,Katoh Y, Nakayama K. Intraflagellar transport-A complex mediates ciliary entry and retrograde trafficking of ciliary G protein-coupled receptors[J]. Mol Biol Cell, 2017, 28(3):429-439.
[11] Sartor ITS, Recamonde-Mendoza M, Ashton-Prolla P. TULP3:A potential biomarker in colorectal cancer?[J]. PLoS One, 2019, 14(1):e0210762.
[12] Sartor ITS, Zeidán-Chuliá F, Albanus RD, et al. Computational analyses reveal a prognostic impact of TULP3 as a transcriptional master regulator in pancreatic ductal adenocarcinoma[J]. Mol Biosyst, 2014, 10(6):1461-1468.
[13] Scanlon CS, van Tubergen EA, Inglehart RC, et al. Biomarkers of epithelial-mesenchymal transition in squamous cell carcinoma[J]. J Dent Res, 2013, 92(2):114-121.
[14] Yeung KT, Yang J. Epithelial-mesenchymal transition in tumor metastasis[J]. Mol Oncol, 2017, 11(1):28-39.
[15] Smith A,Teknos TN, Pan Q. Epithelial to mesenchymal transition in head and neck squamous cell carcinoma[J]. Oral Oncol, 2013, 49(4):287-292.
[16] Baumeister P, Zhou JF, Canis M, et al. Epithelial-to-mesenchy-mal transition-derived heterogeneity in head and neck squamous cell carcinomas[J]. Cancers, 2021, 13(21):5355.
[17] Malumbres M. Cyclin-dependent kinases[J]. Genome Biol, 2014, 15(6):122.
[18] Bertoli C, Skotheim JM, de Bruin RAM. Control of cell cycle transcription during G1 and S phases[J]. Nat Rev Mol Cell Biol, 2013, 14(8):518-528.
[19] Malumbres M, Barbacid M. Cell cycle, CDKs and cancer:A changing paradigm[J]. Nat Rev Cancer, 2009, 9(3):153-166.
[20] Suski JM, Braun M, Strmiska V, et al. Targeting cell-cycle machinery in cancer[J]. Cancer Cell, 2021, 39(6):759-778.

敲减TULP3对头颈鳞癌细胞生物学行为的影响

Effects of knocking down TULP3 on biological behaviors of head and neck squamous cell carcinoma cells

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