Liu Hongfei, Chen Xiaohong, Yang Xingjiu, Li Mengyuan, Zhang Wenlong, Zhang Guoxin, Zhan Xiangwen, Cao Lin, Li Weisha, Huang Zhigang, Gao Ran
Department of Otorhinolaryngology Head and Neck Surgery, Beijing Tongren Hospital, Capital Medical University, Beijing 100730, P.R. China.
NHC Key Laboratory of Human Disease Comparative Medicine, Institute of Laboratory Animal Sciences, Chinese Academy of Medical Sciences Peking Union Medical College, Beijing 100021, P.R. China.
Oncol Lett. 2020 Dec;20(6):394. doi: 10.3892/ol.2020.12257. Epub 2020 Oct 29.
The abnormal upregulation of heterogeneous nuclear ribonucleoprotein K (hnRNP K) expression levels were reported to be involved in the progression of various types of cancer. Therefore, it is hypothesized that hnRNP K may serve as a useful diagnostic marker and antitumor target; however, only a few studies to date have investigated the exact role of hnRNP K in head and neck squamous cell carcinoma (HNSCC) and the potential downstream signaling pathway involved. The present study aimed to identify the roles of hnRNP K in the proliferation and migration of HNSCC, and the possible signaling pathways hnRNP K may be associated with in HNSCC. hnRNP K expression levels in clinical HNSCC samples were analyzed using the Oncomine and UALCAN databases, and its association with the survival of patients with HNSCC was analyzed using the tumor-immune system interactions database. Short hairpin RNA targeting hnRNP K was transfected into the CAL-27 cell line to establish HNSCC cells with stable hnRNP K-knockdown. Cell viability was analyzed using a Cell Counting Kit-8 assay and an absolute count assay, and cell proliferation was measured using 5-ethynyl-2'-deoxyuridine incorporation and colony formation assays. Migratory ability of cells was analyzed using wound healing assay and transwell assay. The growth of xenografts derived from hnRNP K-knockdown cells was also evaluated, and bioinformatics analyses were performed using the Gene Ontology and Kyoto Encyclopedia for Genes and Genomes databases to determine the possible downstream signaling pathways of hnRNP K. Furthermore, the status of the Wnt/β-Catenin signaling pathway in hnRNP K-knockdown cells mediated by small interfering RNA was determined using reverse transcription-quantitative PCR and western blotting. The results revealed that the expression levels of hnRNP K were upregulated in HNSCC cell lines and tissues. Moreover, the upregulation of hnRNP K expression levels was associated with poor survival of patients with HNSCC. The knockdown of hnRNP K also decreased HNSCC cell proliferation and migration, and inhibited tumor growth in nude mice. Bioinformatics analyses identified the Wnt/β-Catenin signaling pathway as a possible downstream signaling pathway of hnRNP K. Knockdown of hnRNP K significantly downregulated the expression levels of Wnt/β-Catenin signaling pathway-related proteins; while with knockdown of hnRNP K and overexpression of β-Catenin, the expression levels of Wnt/β-Catenin signaling pathway-related proteins were partially rescued. In conclusion, the present findings indicated that hnRNP K may serve as a candidate diagnostic biomarker and a promising therapeutic target for HNSCC.
据报道,异质性核核糖核蛋白K(hnRNP K)表达水平的异常上调与多种癌症的进展有关。因此,有人推测hnRNP K可能是一种有用的诊断标志物和抗肿瘤靶点;然而,迄今为止,只有少数研究调查了hnRNP K在头颈部鳞状细胞癌(HNSCC)中的确切作用以及相关的潜在下游信号通路。本研究旨在确定hnRNP K在HNSCC增殖和迁移中的作用,以及hnRNP K在HNSCC中可能相关的信号通路。使用Oncomine和UALCAN数据库分析临床HNSCC样本中hnRNP K的表达水平,并使用肿瘤-免疫系统相互作用数据库分析其与HNSCC患者生存率的关系。将靶向hnRNP K的短发夹RNA转染到CAL-27细胞系中,以建立稳定敲低hnRNP K的HNSCC细胞。使用细胞计数试剂盒-8检测法和绝对计数检测法分析细胞活力,使用5-乙炔基-2'-脱氧尿苷掺入法和集落形成检测法测量细胞增殖。使用伤口愈合检测法和Transwell检测法分析细胞的迁移能力。还评估了源自hnRNP K敲低细胞的异种移植瘤的生长情况,并使用基因本体论和京都基因与基因组百科全书数据库进行生物信息学分析,以确定hnRNP K可能的下游信号通路。此外,使用逆转录-定量PCR和蛋白质印迹法确定小干扰RNA介导的hnRNP K敲低细胞中Wnt/β-连环蛋白信号通路的状态。结果显示,hnRNP K在HNSCC细胞系和组织中的表达水平上调。此外,hnRNP K表达水平的上调与HNSCC患者的不良生存相关。敲低hnRNP K也降低了HNSCC细胞的增殖和迁移,并抑制了裸鼠体内肿瘤的生长。生物信息学分析确定Wnt/β-连环蛋白信号通路是hnRNP K可能的下游信号通路。敲低hnRNP K显著下调了Wnt/β-连环蛋白信号通路相关蛋白的表达水平;而在敲低hnRNP K并过表达β-连环蛋白时,Wnt/β-连环蛋白信号通路相关蛋白的表达水平得到部分恢复。总之,本研究结果表明,hnRNP K可能是HNSCC的候选诊断生物标志物和有前景的治疗靶点。