MicroRNA-101 is a potential prognostic indicator of laryngeal squamous cell carcinoma and modulates CDK8.
BACKGROUND
Various microRNAs (miRNAs) negatively modulate genes that are involved in cellular proliferation, differentiation, invasion, and apoptosis. In many types of cancer, the expression profiles of these miRNAs are altered. Recently, miR-101 was identified as a tumour suppressor and was found to be expressed at low levels in various types of tumours, including prostate, breast, endometrium, and bladder cancers. However, the function(s) of miR-101 in laryngeal carcinoma remain unknown.


METHODS
The expression levels of miR-101 in laryngeal squamous cell carcinoma (LSCC) tissues and cells were detected by qPCR. Cell proliferation, migration, cell cycle, and apoptosis assay were applied to assess the function(s) of miR-101 in vitro. Nude mice subcutaneous tumour model was used to perform in vivo study. Moreover, we identified Cyclin-dependent kinase 8 (CDK8) as the target of miR-101 by a luciferase assay. The possible downstream effectors of CDK8 were investigated in Wnt/β-catenin signaling pathway. Changes of CDK8, β-catenin, and cyclin D1 protein levels were analyzed by western blotting and immunohistochemical staining. The prognostic effect of miR-101 was evaluated using the Kaplan-Meier method.


RESULTS
Expression of miR-101 was down-regulated in the LSCC tissues compared with the adjacent normal tissues. Furthermore, downregulation of miR-101 correlated with T3-4 tumour grade, lymph node metastasis, and an advanced clinical stage in the LSCC patients examined (P < 0.05). The low level of miR-101 expression was associated with poor prognosis (P < 0.05). CDK8 was identified as the target gene of miR-101 by luciferase reporter assay. Moreover, we showed that up-regulation of miR-101 expression suppressed humen LSCC Hep-2 cells proliferation and migration, and induced cell-cycle arrest. Increased expression of miR-101 induced cells apoptosis both in vitro and in vivo. Correspondingly, exogenous expression of miR-101 significantly reduced the growth of tumour in a LSCC xenograft model. Furthermore, the miR-101 level was inversely correlated with levels of CDK8, β-catenin, and cyclin D1 in western blotting assay and immunohistochemical staining assay.


CONCLUSIONS
These results indicate that miR-101 is a potent tumour repressor that directly represses CDK8 expression. Thus, detection and targeting of miR-101 may represent a novel diagnostic and therapeutic strategy for LSCC patients.
Services of Head and Neck Surgery, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. lmh012345@163.com.
MingHua
M
Li
Services of Laryngology, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. linjingyi2005@qq.com.
LinLi
L
Tian
The First Clinical Hospital Affiliated to Harbin Medical University, Harbin, 150001, People's Republic of China. 475818992@qq.com.
Hui
H
Ren
Services of Head and Neck Surgery, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. dlbaxin@163.com.
XiaoXue
X
Chen
Services of Laryngology, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. xberry80@sina.com.
Yu
Y
Wang
Services of Laryngology, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. 1621070879@qq.com.
JingChun
J
Ge
The Human Anatomy and Histoembryology Department, Harbin Medical University, Harbin, 150081, People's Republic of China. 13936447287@163.com.
ShuLiang
S
Wu
Services of Head and Neck Surgery, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. jiaobo200508@163.com.
YaNan
Y
Sun
Services of Head and Neck Surgery, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. liumingorl@qq.com.
Ming
M
Liu
Services of Laryngology, Department of Otolaryngology-Head and Neck Surgery, The Second Affiliated Hospital of Harbin Medical University, No. 148, Bao jian Road, Harbin, 150081, People's Republic of China. ybbxuz2001@163.com.
Hui
H
Xiao
D000818:Animals; D017209:Apoptosis; D002294:Carcinoma, Squamous Cell / Q000378:metabolism*; D002453:Cell Cycle; D002460:Cell Line; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D056850:Cyclin-Dependent Kinase 8 / Q000378:metabolism*; D020869:Gene Expression Profiling; D015972:Gene Expression Regulation, Neoplastic; D016147:Genes, Tumor Suppressor; D006801:Humans; D007822:Laryngeal Neoplasms / Q000378:metabolism*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D035683:MicroRNAs / Q000378:metabolism*; D009361:Neoplasm Invasiveness; D016133:Polymerase Chain Reaction; D060449:Wnt Signaling Pathway; D023041:Xenograft Model Antitumor Assays; D051176:beta Catenin / Q000378:metabolism
C534072:MIRN101 microRNA, human; D035683:MicroRNAs; D051176:beta Catenin; C095686:CDK8 protein, human; D056850:Cyclin-Dependent Kinase 8