Recently, several microRNAs (miRNAs) were reported to be involved in the modulation of glioma development. The aim of our study was to determine the effect of miR-181d on the growth of glioma and to investigate whether this growth is modulated by targeting K-ras and Bcl-2.
Real-time PCR was used to analyze the expression of miR-181d in human glioma samples and glioma cell lines. Apoptosis, cell cycle, and proliferation (MTT) assays were performed to assess the phenotypic changes in glioma cells. Immunohistochemistry was used to determine the expression of K-ras and Bcl-2 in glioma tissues, and a luciferase reporter assay was carried out to confirm whether K-ras and Bcl-2 are direct targets of miR-181d. Western blotting was used to identify the potential signaling pathways affected glioma cell growth by miR-181d. In vivo, xenograft tumors were examined for an anti-glioma effect of miR-181d.
MiR-181d was down-regulated in human glioma samples and up-regulated in transfected glioma cells. Ectopic expression of miR-181d suppressed proliferation and triggered cell cycle arrest and apoptosis in glioma cell lines. K-ras and Bcl-2 were identified as direct targets of miR-181d and were up-regulated in glioma samples. The results showed evidence linking the tumor suppressor activity of miR-181d in glioma cells with the K-ras-related PI3K/AKT and MAPK/ERK pathways. Furthermore, xenograft tumors from miR-181d-treated U251 cells were suppressed in vivo.
MiR-181d may act as a glioma suppressor by targeting K-ras and Bcl-2.
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Aguirre AJ, Bardeesy N, Sinha M, Lopez L, Tuveson DA, Horner J, Redston MS, DePinho RA (2003) Activated Kras and Ink4a/Arf deficiency cooperate to produce metastatic pancreatic ductal adenocarcinoma. Genes Dev 17:3112–3126
Almoguera C, Shibata D, Forrester K, Martin J, Arnheim N, Perucho M (1988) Most human carcinomas of the exocrine pancreas contain mutant c-K-ras genes. Cell 53:549–554
Bodemann BO, White MA (2008) Ral GTPases and cancer: linchpin support of the tumorigenic platform. Nat Rev Cancer 8:133–140
Calin GA, Croce CM (2006) MicroRNA signatures in human cancers. Nat Rev Cancer 6:857–866
Caulin C, Nguyen T, Longley MA, Zhou Z, Wang XJ, Roop DR (2004) Inducible activation of oncogenic K-ras results in tumor formation in the oral cavity. Cancer Res 64:5054–5058
Chan JA, Krichevsky AM, Kosik KS (2005) MicroRNA-21 is an antiapoptotic factor in human glioblastoma cells. Cancer Res 65:6029–6033
Chen CZ (2005) MicroRNAs as oncogenes and tumor suppressors. N Engl J Med 353:1768–1771
Chen CZ, Li L, Lodish HF, Bartel DP (2004) MicroRNAs modulate hematopoietic lineage differentiation. Science 303:83–86
Davies H, Bignell GR, Cox C, Stephens P, Edkins S, Clegg S, Teague J, Woffendin H, Garnett MJ, Bottomley W, Davis N, Dicks E, Ewing R, Floyd Y, Gray K, Hall S, Hawes R, Hughes J, Kosmidou V, Menzies A, Mould C, Parker A, Stevens C, Watt S, Hooper S, Wilson R, Jayatilake H, Gusterson BA, Cooper C, Shipley J, Hargrave D, Pritchard-Jones K, Maitland N, Chenevix-Trench G, Riggins GJ, Bigner DD, Palmieri G, Cossu A, Flanagan A, Nicholson A, Ho JW, Leung SY, Yuen ST, Weber BL, Seigler HF, Darrow TL, Paterson H, Marais R, Marshall CJ, Wooster R, Stratton MR, Futreal PA (2002) Mutations of the BRAF gene in human cancer. Nature 417:949–954
Deininger MH, Weller M, Streffer J, Meyermann R (1999) Antiapoptotic Bcl-2 family protein expression increases with progression of oligodendroglioma. Cancer 86:1832–1839
Engelman JA, Chen L, Tan X, Crosby K, Guimaraes AR, Upadhyay R, Maira M, McNamara K, Perera SA, Song Y, Chirieac LR, Kaur R, Lightbown A, Simendinger J, Li T, Padera RF, Garcia-Echeverria C, Weissleder R, Mahmood U, Cantley LC, Wong KK (2008) Effective use of PI3K and MEK inhibitors to treat mutant Kras G12D and PIK3CA H1047R murine lung cancers. Nat Med 14:1351–1356
Esquela-Kerscher A, Slack FJ (2006) Oncomirs—microRNAs with a role in cancer. Nat Rev Cancer 6:259–269
Forbes S, Clements J, Dawson E, Bamford S, Webb T, Dogan A, Flanagan A, Teague J, Wooster R, Futreal PA, Stratton MR (2006) Cosmic 2005. Br J Cancer 94:318–322
Gabriely G, Wurdinger T, Kesari S, Esau CC, Burchard J, Linsley PS, Krichevsky AM (2008) MicroRNA 21 promotes glioma invasion by targeting matrix metalloproteinase regulators. Mol Cell Biol 28:5369–5380
Gabriely G, Yi M, Narayan RS, Niers JM, Wurdinger T, Imitola J, Ligon KL, Kesari S, Esau C, Stephens RM, Tannous BA, Krichevsky AM (2011) Human glioma growth is controlled by microRNA-10b. Cancer Res 71:3563–3572
Godlewski J, Nowicki MO, Bronisz A, Nuovo G, Palatini J, De Lay M, Van Brocklyn J, Ostrowski MC, Chiocca EA, Lawler SE (2010) MicroRNA-451 regulates LKB1/AMPK signaling and allows adaptation to metabolic stress in glioma cells. Mol Cell 37:620–632
Hall A (1990) The cellular functions of small GTP-binding proteins. Science 249:635–640
Hingorani SR, Petricoin EF, Maitra A, Rajapakse V, King C, Jacobetz MA, Ross S, Conrads TP, Veenstra TD, Hitt BA, Kawaguchi Y, Johann D, Liotta LA, Crawford HC, Putt ME, Jacks T, Wright CV, Hruban RH, Lowy AM, Tuveson DA (2003) Preinvasive and invasive ductal pancreatic cancer and its early detection in the mouse. Cancer Cell 4:437–450
Howell RE, Wong FS, Fenwick RG (1990) A transforming Kirsten ras oncogene in an oral squamous carcinoma. J Oral Pathol Med 19:301–305
Janssen KP, Alberici P, Fsihi H, Gaspar C, Breukel C, Franken P, Rosty C, Abal M, El Marjou F, Smits R, Louvard D, Fodde R, Robine S (2006) APC and oncogenic KRAS are synergistic in enhancing Wnt signaling in intestinal tumor formation and progression. Gastroenterology 131:1096–1109
Janzarik WG, Kratz CP, Loges NT, Olbrich H, Klein C, Schafer T, Scheurlen W, Roggendorf W, Weiller C, Niemeyer C, Korinthenberg R, Pfister S, Omran H (2007) Further evidence for a somatic KRAS mutation in a pilocytic astrocytoma. Neuropediatrics 38:61–63
Ji J, Yamashita T, Budhu A, Forgues M, Jia HL, Li C, Deng C, Wauthier E, Reid LM, Ye QH, Qin LX, Yang W, Wang HY, Tang ZY, Croce CM, Wang XW (2009) Identification of microRNA-181 by genome-wide screening as a critical player in EpCAM-positive hepatic cancer stem cells. Hepatology 50:472–480
Jiang Z, Zheng X, Rich KM (2003) Down-regulation of Bcl-2 and Bcl-xL expression with bispecific antisense treatment in glioblastoma cell lines induce cell death. J Neurochem 84:273–281
Julien T, Frankel B, Longo S, Kyle M, Gibson S, Shillitoe E, Ryken T (2000) Antisense-mediated inhibition of the bcl-2 gene induces apoptosis in human malignant glioma. Surg Neurol 53:360–368 discussion 368–369
Kefas B, Godlewski J, Comeau L, Li Y, Abounader R, Hawkinson M, Lee J, Fine H, Chiocca EA, Lawler S, Purow B (2008) microRNA-7 inhibits the epidermal growth factor receptor and the Akt pathway and is down-regulated in glioblastoma. Cancer Res 68:3566–3572
Krajewski S, Krajewska M, Ehrmann J, Sikorska M, Lach B, Chatten J, Reed JC (1997) Immunohistochemical analysis of Bcl-2, Bcl-X, Mcl-1, and Bax in tumors of central and peripheral nervous system origin. Am J Pathol 150:805–814
Kwak HJ, Kim YJ, Chun KR, Woo YM, Park SJ, Jeong JA, Jo SH, Kim TH, Min HS, Chae JS, Choi EJ, Kim G, Shin SH, Gwak HS, Kim SK, Hong EK, Lee GK, Choi KH, Kim JH, Yoo H, Park JB, Lee SH (2011) Downregulation of Spry2 by miR-21 triggers malignancy in human gliomas. Oncogene 30:2433–2442
McDonald FE, Ironside JW, Gregor A, Wyatt B, Stewart M, Rye R, Adams J, Potts HW (2002) The prognostic influence of bcl-2 in malignant glioma. Br J Cancer 86:1899–1904
Qi L, Bart J, Tan LP, Platteel I, Sluis T, Huitema S, Harms G, Fu L, Hollema H, Berg A (2009) Expression of miR-21 and its targets (PTEN, PDCD4, TM1) in flat epithelial atypia of the breast in relation to ductal carcinoma in situ and invasive carcinoma. BMC Cancer 9:163
Qian B, Katsaros D, Lu L, Preti M, Durando A, Arisio R, Mu L, Yu H (2009) High miR-21 expression in breast cancer associated with poor disease-free survival in early stage disease and high TGF-beta1. Breast Cancer Res Treat 117:131–140
Rieger L, Weller M, Bornemann A, Schabet M, Dichgans J, Meyermann R (1998) BCL-2 family protein expression in human malignant glioma: a clinical-pathological correlative study. J Neurol Sci 155:68–75
Rozenblum E, Schutte M, Goggins M, Hahn SA, Panzer S, Zahurak M, Goodman SN, Sohn TA, Hruban RH, Yeo CJ, Kern SE (1997) Tumor-suppressive pathways in pancreatic carcinoma. Cancer Res 57:1731–1734
Santos E, Martin-Zanca D, Reddy EP, Pierotti MA, Della Porta G, Barbacid M (1984) Malignant activation of a K-ras oncogene in lung carcinoma but not in normal tissue of the same patient. Science 223:661–664
Schubbert S, Shannon K, Bollag G (2007) Hyperactive Ras in developmental disorders and cancer. Nat Rev Cancer 7:295–308
Sharma MK, Zehnbauer BA, Watson MA, Gutmann DH (2005) RAS pathway activation and an oncogenic RAS mutation in sporadic pilocytic astrocytoma. Neurology 65:1335–1336
Shi L, Cheng Z, Zhang J, Li R, Zhao P, Fu Z, You Y (2008) hsa-mir-181a and hsa-mir-181b function as tumor suppressors in human glioma cells. Brain Res 1236:185–193
Shin KH, Bae SD, Hong HS, Kim RH, Kang MK, Park NH (2011) miR-181a shows tumor suppressive effect against oral squamous cell carcinoma cells by downregulating K-ras. Biochem Biophys Res Commun 404:896–902
Strik H, Deininger M, Streffer J, Grote E, Wickboldt J, Dichgans J, Weller M, Meyermann R (1999) BCL-2 family protein expression in initial and recurrent glioblastomas: modulation by radiochemotherapy. J Neurol Neurosurg Psychiatry 67:763–768
Sun L, Yan W, Wang Y, Sun G, Luo H, Zhang J, Wang X, You Y, Yang Z, Liu N (2011) MicroRNA-10b induces glioma cell invasion by modulating MMP-14 and uPAR expression via HOXD10. Brain Res 1389:9–18
Tang H, Liu X, Wang Z, She X, Zeng X, Deng M, Liao Q, Guo X, Wang R, Li X, Zeng F, Wu M, Li G (2011) Interaction of hsa-miR-381 and glioma suppressor LRRC4 is involved in glioma growth. Brain Res 1390:21–32
Todd R, Donoff RB, Wong DT (1997) The molecular biology of oral carcinogenesis: toward a tumor progression model. J Oral Maxillofac Surg 55:613–623 discussion 623–615
Wang B, Hsu SH, Majumder S, Kutay H, Huang W, Jacob ST, Ghoshal K (2010) TGFbeta-mediated upregulation of hepatic miR-181b promotes hepatocarcinogenesis by targeting TIMP3. Oncogene 29:1787–1797
Wee S, Jagani Z, Xiang KX, Loo A, Dorsch M, Yao YM, Sellers WR, Lengauer C, Stegmeier F (2009) PI3K pathway activation mediates resistance to MEK inhibitors in KRAS mutant cancers. Cancer Res 69:4286–4293
Yip KW, Reed JC (2008) Bcl-2 family proteins and cancer. Oncogene 27:6398–6406
Yu K, Toral-Barza L, Shi C, Zhang WG, Zask A (2008) Response and determinants of cancer cell susceptibility to PI3K inhibitors: combined targeting of PI3K and Mek1 as an effective anticancer strategy. Cancer Biol Ther 7:307–315
Zhang Y, Chao T, Li R, Liu W, Chen Y, Yan X, Gong Y, Yin B, Liu W, Qiang B, Zhao J, Yuan J, Peng X (2009) MicroRNA-128 inhibits glioma cells proliferation by targeting transcription factor E2F3a. J Mol Med 87:43–51
Zhang C, Hao L, Wang L, Xiao Y, Ge H, Zhu Z, Luo Y, Zhang Y, Zhang Y (2010) Elevated IGFIR expression regulating VEGF and VEGF-C predicts lymph node metastasis in human colorectal cancer. BMC Cancer 10:184
Zhu S, Wu H, Wu F, Nie D, Sheng S, Mo YY (2008) MicroRNA-21 targets tumor suppressor genes in invasion and metastasis. Cell Res 18:350–359
This work was supported by China Natural Science Foundation (30872657, 30971136 and 81072078), Natural Science Foundation of Jiangsu Province (2008475 and 2010580), Scientific Program of Ministry of Health (W2011BX009), Program for Development of Innovative Research Team in the First Affiliated Hospital of NJMU, and A Project Funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions.
Conflict of interest
No potential conflicts of interest were disclosed.
Xie-Feng Wang, Zhu-Mei Shi, Xi-Rui Wang contributed equally to this work.
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Wang, X., Shi, Z., Wang, X. et al. MiR-181d acts as a tumor suppressor in glioma by targeting K-ras and Bcl-2. J Cancer Res Clin Oncol 138, 573–584 (2012). https://doi.org/10.1007/s00432-011-1114-x