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Targeted Oncology

, Volume 11, Issue 2, pp 197–207 | Cite as

RETRACTED ARTICLE: MicroRNA-216b is Down-Regulated in Human Gastric Adenocarcinoma and Inhibits Proliferation and Cell Cycle Progression by Targeting Oncogene HDAC8

  • Ying WangEmail author
  • Po Xu
  • Jun Yao
  • Ruina Yang
  • Zhenguo Shi
  • Xiaojuan Zhu
  • Xiaoshan FengEmail author
  • Shegan GaoEmail author
Original Research Article

Abstract

Purpose

Accumulating evidence indicates that micro (mi)RNAs play a critical role in carcinogenesis and cancer progression; however, their role in the tumorigenesis of gastric adenocarcinoma remains unclear so the present study investigated this in gastric cancer (GC) tissues and cell lines.

Methods

Human GC specimens (n = 57) and patient-paired non-cancerous specimens were obtained from patients at the First Affiliated Hospital, Henan University of Science and Technology. The AGS and GC9811 gastric cancer cell lines were also used. Expression levels of miR-216b and HDAC8 were examined by quantitative real-time PCR and the expression of HDAC8 was also examined by Western blotting and immunohistochemistry assay. The cell cycle progression was determined by FACS. MiR-216b inhibitor, mimics, and siRNA-HDAC8 transfections were performed to study the loss and gain of function.

Results

We reported a significantly decreased expression of miR-216b in GC clinical specimens compared with paired non-cancerous tissues. We also observed a significant down-regulation of miR-216b expression in GC cell lines AGS and GC9811 (p < 0.0001). The introduction of miR-216b suppressed GC cell proliferation and cell cycle progression by targeting HDAC8, an oncogene shown to promote malignant tumor development with a potential miR-216b binding site in its 3′ untranslated region. HDAC8 expression was shown to be significantly increased in AGS and GC9811 cell lines (p < 0.0001) and GC tissues compared with controls. Moreover, HDAC8 inhibition suppressed cell cycle progression compared with control groups (22 % ± 1.6 % vs 34 % ± 2.1), indicating that HDAC8 may function as an oncogene in the development of GC. Furthermore, HDAC8 expression was negatively correlated (p < 0.0001), while miR-216b expression was positively correlated with the clinical outcome of GC patients (p < 0.0001).

Discussion

Our data suggest that miR-216b functions as a tumor suppressor in human GC by, at least partially, targeting HDAC8.

Keywords

Gastric Cancer Gastric Cancer Cell Line Human Gastric Cancer HDAC8 Expression GC9811 Cell 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Author Contributions

Y.W. performed data analyses and wrote the manuscript, initiated the project, designed the experiments, and interpreted the data. P.X. and ZG.S. performed cell culture and cell transfection. SY.S. and XJ.Z. performed qRT-PCR analysis. XS.F. and SG.G performed the cell cycle and proliferation experiments in vitro.

Conflict of Interest

Dr. Ying Wang, Dr. Po Xu, Dr. Jun Yao, Dr. Ruina Yang, Dr. Zhenguo Shi, Dr. Xiaojuan Zhu, Dr. Xiaoshan Feng and Dr. Shegan Gao have no conflicts of interest to disclose.

Funding Statement

This study was supported by the National Natural Science Foundation of China (No. 81301763 and No. 81572849) and Henan provincial key scientific and technological projects (No. 142102310473).

Ethics Statement

Written informed consent was obtained from patients before obtaining tissue samples. The procedures used in this study were approved by the Institutional Review Board of the First Affiliated Hospital, Henan University of Science and Technology and conformed to the Helsinki Declaration and to local legislation.

Supplementary material

11523_2015_390_MOESM1_ESM.docx (16 kb)
Supplementary Table 1 Relationship between clinical parameters and miR-216b (mean ± SE) expression in primary gastric adenocarcinoma. One-way ANOVA analysis, n = 3 (DOCX 15 kb)
11523_2015_390_MOESM2_ESM.docx (20 kb)
Supplementary Table 2 Relationship between clinical parameters and HDAC8 (mean ± SE) expression in primary gastric adenocarcinoma. One-way ANOVA analysis, n = 3 (DOCX 20 kb)
11523_2015_390_MOESM3_ESM.docx (17 kb)
Supplementary Table 3 Antibodies used in Western blotting assay (DOCX 17 kb)
11523_2015_390_Fig7_ESM.gif (69 kb)
Supplementary Fig. 1

A. Schematic of Luc-HDAC8 3′UTR and Luc-HDAC8 3′MutUTR constructs. Luc-HDAC8 3′UTR and Luc-HDAC8 3′Mut UTR were cloned into a pmirGLO plasmid downstream of the firefly luciferase coding region between PmeI and XbaI sites. B. Wild-type and mutant HDAC8-3′UTR containing the putative binding site of miR-216b were cloned into psiCHECK-2 vector. HDAC8-3′UTR was amplified from genomic DNA of GC9811. C. Lane 3,4: Recombinant plasmids of HDAC8-1; lane 7,8: Results of enzyme digestion of recombinant plasmids of HDAC8-1. Results showed that HDAC8-1 have been successfully inserted into the vectors.(M: DL2000 DNA Marker; HDAC8-1 bands: 1283 bp; Vectors bands: 6.1 Kb). D. M: 1 kb DNA Ladder Marker. Lane 1: amplification of mutHDAC8F1/R1 (negative control without Taq enzyme). E. Sequencing results of WT-HDAC8 and MT –HDAC8 (WT: wild type; MT: mutated type). (GIF 69 kb)

11523_2015_390_MOESM4_ESM.tif (2.5 mb)
High resolution image (TIFF 2586 kb)
11523_2015_390_Fig8_ESM.gif (8 kb)
Supplementary Fig. 2

Expression of HDAC8 in GC cells and the normal gastric epithelial cells were examined by Western blotting and shown as mean ± SE (normalized to tubulin). (GIF 7 kb)

11523_2015_390_MOESM5_ESM.tif (482 kb)
High resolution image (TIFF 482 kb)

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Copyright information

© Springer International Publishing Switzerland 2015

Authors and Affiliations

  1. 1.Oncology Department of the First Affiliated Hospital of Henan University of Science and TechnologyLuoyangChina
  2. 2.Urology Surgery Department of the First Affiliated Hospital of Henan University of Science and TechnologyLuoyangChina

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