Abstract
Gallbladder cancer (GBC) is a highly malignant cancer with poor prognosis. Although long noncoding RNA (lncRNA) H19 has been reported to play vital role in many human cancers, whether it is involved in GBC proliferation is still unknown. This study was designed to explore the effect of H19 in GBC cell proliferation. The expression of H19 and AKT2 were significantly elevated in GBC tissues, and the level of miR-194-5p is markedly decreased. Moreover, the RNA levels of H19 and AKT2 were positively correlated, and H19 elevation was significantly associated with tumor size. Cell proliferation decreased significantly after knockdown of H19 in GBC-SD and NOZ cells and after knockdown of AKT2 in NOZ cells. Results from cell cycle studies indicated that the S phase were significantly decreased after knockdown of H19 in NOZ cells but significantly elevated after overexpression of H19 in GBC-SD cells. Furthermore, knockdown of H19 upregulated miR-194-5p levels, yet significantly decreased miR-194-5p targeting AKT2 gene expression in NOZ cells. Inhibitor against miR-194-5p reversed these effects. In addition, overexpression of H19 in GBC-SD cells downregulated miR-194-5p and markedly increased AKT2 expression, and miR-194-5p mimic reversed these effects. Eventually, GBC cells were arrested in G0/G1-phase after H19 knockdown, inhibition of miR-194-5p markedly promoted cells into S-phase and co-transfection of siH19, and miR-194-5p inhibitor exerted mutually counter-regulated effects on cell cycle. These results suggested that H19/miR-194-5p/AKT2 axis regulatory network might modulate cell proliferation in GBC.
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References
Siegel RL, Miller KD, Jemal A. Cancer statistics, 2015. CA: Cancer J Clin. 2015;65(1):5–29. doi:10.3322/caac.21254.
Kanthan R, Senger JL, Ahmed S, Kanthan SC. Gallbladder Cancer in the 21st Century. J Oncol. 2015;2015:967472. doi:10.1155/2015/967472.
Zhu AX, Hong TS, Hezel AF, Kooby DA. Current management of gallbladder carcinoma. Oncologist. 2010;15(2):168–81. doi:10.1634/theoncologist.2009-0302.
Rakic M, Patrlj L, Kopljar M, Klicek R, Kolovrat M, Loncar B, et al. Gallbladder cancer. Hepatobiliary Surg Nutr. 2014;3(5):221–6. doi:10.3978/j.issn.2304-3881.2014.09.03.
Managadze D, Lobkovsky AE, Wolf YI, Shabalina SA, Rogozin IB, Koonin EV. The vast, conserved mammalian lincRNome. PLoS Comput Biol. 2013;9(2):e1002917. doi:10.1371/journal.pcbi.1002917.
Dey BK, Mueller AC, Dutta A. Long non-coding RNAs as emerging regulators of differentiation, development, and disease. Transcription. 2014;5(4):e944014. doi:10.4161/21541272.2014.944014.
Panzitt K, Tschernatsch MM, Guelly C, Moustafa T, Stradner M, Strohmaier HM, et al. Characterization of HULC, a novel gene with striking up-regulation in hepatocellular carcinoma, as noncoding RNA. Gastroenterology. 2007;132(1):330–42. doi:10.1053/j.gastro.2006.08.026.
Pandey GK, Mitra S, Subhash S, Hertwig F, Kanduri M, Mishra K, et al. The risk-associated long noncoding RNA NBAT-1 controls neuroblastoma progression by regulating cell proliferation and neuronal differentiation. Cancer Cell. 2014;26(5):722–37. doi:10.1016/j.ccell.2014.09.014.
Brunkow ME, Tilghman SM. Ectopic expression of the H19 gene in mice causes prenatal lethality. Genes Dev. 1991;5(6):1092–101.
Lottin S, Adriaenssens E, Dupressoir T, Berteaux N, Montpellier C, Coll J, et al. Overexpression of an ectopic H19 gene enhances the tumorigenic properties of breast cancer cells. Carcinogenesis. 2002;23(11):1885–95.
Zhang Y, Shields T, Crenshaw T, Hao Y, Moulton T, Tycko B. Imprinting of human H19: allele-specific CpG methylation, loss of the active allele in Wilms tumor, and potential for somatic allele switching. Am J Hum Genet. 1993;53(1):113–24.
Polytarchou C, Iliopoulos D, Hatziapostolou M, Kottakis F, Maroulakou I, Struhl K, et al. Akt2 regulates all Akt isoforms and promotes resistance to hypoxia through induction of miR-21 upon oxygen deprivation. Cancer Res. 2011;71(13):4720–31. doi:10.1158/0008-5472.CAN-11-0365.
Fohlin H, Perez-Tenorio G, Fornander T, Skoog L, Nordenskjold B, Carstensen J, et al. Akt2 expression is associated with good long-term prognosis in oestrogen receptor positive breast cancer. Eur J Cancer. 2013;49(6):1196–204. doi:10.1016/j.ejca.2012.12.006.
Nitsche C, Edderkaoui M, Moore RM, Eibl G, Kasahara N, Treger J, et al. The phosphatase PHLPP1 regulates Akt2, promotes pancreatic cancer cell death, and inhibits tumor formation. Gastroenterology. 2012;142(2):377–87 e1-5. doi:10.1053/j.gastro.2011.10.026.
Galicia VA, He L, Dang H, Kanel G, Vendryes C, French BA, et al. Expansion of hepatic tumor progenitor cells in Pten-null mice requires liver injury and is reversed by loss of AKT2. Gastroenterology. 2010;139(6):2170–82. doi:10.1053/j.gastro.2010.09.002.
Zhao HJ, Ren LL, Wang ZH, Sun TT, Yu YN, Wang YC, et al. MiR-194 deregulation contributes to colorectal carcinogenesis via targeting AKT2 pathway. Theranostics. 2014;4(12):1193–208. doi:10.7150/thno.8712.
Pachnis V, Belayew A, Tilghman SM. Locus unlinked to alpha-fetoprotein under the control of the murine raf and Rif genes. Proc Natl Acad Sci U S A. 1984;81(17):5523–7.
Zhou X, Yin C, Dang Y, Ye F, Zhang G. Identification of the long non-coding RNA H19 in plasma as a novel biomarker for diagnosis of gastric cancer. Sci Report. 2015;5:11516. doi:10.1038/srep11516.
Vennin C, Spruyt N, Dahmani F, Julien S, Bertucci F, Finetti P, et al. H19 non coding RNA-derived miR-675 enhances tumorigenesis and metastasis of breast cancer cells by downregulating c-Cbl and Cbl-b. Oncotarget. 2015;6(30):29209–23. doi:10.18632/oncotarget.4976.
Medrzycki M, Zhang Y, Zhang W, Cao K, Pan C, Lailler N, et al. Histone h1.3 suppresses h19 noncoding RNA expression and cell growth of ovarian cancer cells. Cancer Res. 2014;74(22):6463–73. doi:10.1158/0008-5472.CAN-13-2922.
Steenman MJ, Rainier S, Dobry CJ, Grundy P, Horon IL, Feinberg AP. Loss of imprinting of IGF2 is linked to reduced expression and abnormal methylation of H19 in Wilms’ tumour. Nat Genet. 1994;7(3):433–9. doi:10.1038/ng0794-433.
Honda S, Arai Y, Haruta M, Sasaki F, Ohira M, Yamaoka H, et al. Loss of imprinting of IGF2 correlates with hypermethylation of the H19 differentially methylated region in hepatoblastoma. Br J Cancer. 2008;99(11):1891–9. doi:10.1038/sj.bjc.6604754.
Zhu M, Chen Q, Liu X, Sun Q, Zhao X, Deng R, et al. lncRNA H19/miR-675 axis represses prostate cancer metastasis by targeting TGFBI. FEBS J. 2014;281(16):3766–75. doi:10.1111/febs.12902.
Zhang L, Yang F, Yuan JH, Yuan SX, Zhou WP, Huo XS, et al. Epigenetic activation of the MiR-200 family contributes to H19-mediated metastasis suppression in hepatocellular carcinoma. Carcinogenesis. 2013;34(3):577–86. doi:10.1093/carcin/bgs381.
Tsang WP, Ng EK, Ng SS, Jin H, Yu J, Sung JJ, et al. Oncofetal H19-derived miR-675 regulates tumor suppressor RB in human colorectal cancer. Carcinogenesis. 2010;31(3):350–8. doi:10.1093/carcin/bgp181.
Liang WC, Fu WM, Wong CW, Wang Y, Wang WM, Hu GX, et al. The lncRNA H19 promotes epithelial to mesenchymal transition by functioning as miRNA sponges in colorectal cancer. Oncotarget. 2015;6(26):22513–25. doi:10.18632/oncotarget.4154.
Yoshimizu T, Miroglio A, Ripoche MA, Gabory A, Vernucci M, Riccio A, et al. The H19 locus acts in vivo as a tumor suppressor. Proc Natl Acad Sci U S A. 2008;105(34):12417–22. doi:10.1073/pnas.0801540105.
Seitz H. Redefining microRNA targets. Curr Biol : CB. 2009;19(10):870–3. doi:10.1016/j.cub.2009.03.059.
Salmena L, Poliseno L, Tay Y, Kats L, Pandolfi PP. A ceRNA hypothesis: the Rosetta Stone of a hidden RNA language? Cell. 2011;146(3):353–8. doi:10.1016/j.cell.2011.07.014.
Kallen AN, Zhou XB, Xu J, Qiao C, Ma J, Yan L, et al. The imprinted H19 lncRNA antagonizes let-7 microRNAs. Mol Cell. 2013;52(1):101–12. doi:10.1016/j.molcel.2013.08.027.
Zhou X, Ye F, Yin C, Zhuang Y, Yue G, Zhang G. The Interaction Between MiR-141 and lncRNA-H19 in Regulating Cell Proliferation and Migration in Gastric Cancer. Cell Physiol Biochem : Int J Exp Cell Physiol Biochem Pharmacol. 2015;36(4):1440–52. doi:10.1159/000430309.
Imig J, Brunschweiger A, Brummer A, Guennewig B, Mittal N, Kishore S, et al. miR-CLIP capture of a miRNA targetome uncovers a lincRNA H19-miR-106a interaction. Nat Chem Biol. 2015;11(2):107–14. doi:10.1038/nchembio.1713.
Liu C, Chen Z, Fang J, Xu A, Zhang W, Wang Z. H19-derived miR-675 contributes to bladder cancer cell proliferation by regulating p53 activation. Tumour Biol : J Int Soc Oncodev Biol Med. 2015. doi:10.1007/s13277-015-3779-2.
Chiam K, Wang T, Watson DI, Mayne GC, Irvine TS, Bright T, et al. Circulating Serum Exosomal miRNAs As Potential Biomarkers for Esophageal Adenocarcinoma. J Gastrointest Surg : Off J Soc Surg Aliment Tract. 2015;19(7):1208–15. doi:10.1007/s11605-015-2829-9.
Franke TF, Hornik CP, Segev L, Shostak GA, Sugimoto C. PI3K/Akt and apoptosis: size matters. Oncogene. 2003;22(56):8983–98. doi:10.1038/sj.onc.1207115.
Lv J, Ma L, Chen XL, Huang XH, Wang Q. Downregulation of LncRNAH19 and MiR-675 promotes migration and invasion of human hepatocellular carcinoma cells through AKT/GSK-3beta/Cdc25A signaling pathway. J Huazhong Univ Sci Technol Med Sci = Hua zhong ke ji da xue xue bao Yi xue Ying De wen ban = Huazhong keji daxue xuebao Yixue Yingdewen ban. 2014;34(3):363–9. doi:10.1007/s11596-014-1284-2.
Zhang P, Guo Z, Wu Y, Hu R, Du J, He X, et al. Histone deacetylase inhibitors inhibit the proliferation of gallbladder carcinoma cells by suppressing AKT/mTOR signaling. PLoS One. 2015;10(8):e0136193. doi:10.1371/journal.pone.0136193.
Chin YR, Yuan X, Balk SP, Toker A. PTEN-deficient tumors depend on AKT2 for maintenance and survival. Cancer Discovery. 2014;4(8):942–55. doi:10.1158/2159-8290.CD-13-0873.
Xu X, Sakon M, Nagano H, Hiraoka N, Yamamoto H, Hayashi N, et al. Akt2 expression correlates with prognosis of human hepatocellular carcinoma. Oncol Rep. 2004;11(1):25–32.
Acknowledgments
We thank the Eastern Hepatobiliary Surgical Hospital and Institute, The Second Military University, Shanghai, for their help. This work was supported by the National Natural Science Foundation of China (Grant numbers 81272747and 81572297) and Doctorial innovation fund of Shanghai Jiao Tong University School of Medicine. We thank Dr. Qiu Lei, who helped us with language editing.
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Shou-Hua Wang and Xiao-Cai Wu contributed equally to this work.
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Wang, SH., Wu, XC., Zhang, MD. et al. Long noncoding RNA H19 contributes to gallbladder cancer cell proliferation by modulated miR-194-5p targeting AKT2. Tumor Biol. 37, 9721–9730 (2016). https://doi.org/10.1007/s13277-016-4852-1
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DOI: https://doi.org/10.1007/s13277-016-4852-1