Tumor Biology

, Volume 35, Issue 10, pp 10177–10184 | Cite as

MiR-7-5p is frequently downregulated in glioblastoma microvasculature and inhibits vascular endothelial cell proliferation by targeting RAF1

  • Zhiguo Liu
  • Yuguang Liu
  • Lianling Li
  • Zhenkuan Xu
  • Baibin Bi
  • Yunyan Wang
  • Jian Yi Li
Research Article


The aberrant expression of microRNAs (miRNAs) is always associated with tumor development and progression. Microvascular proliferation is one of the unique pathologic features of glioblastoma (GBM) . In this study, the microvasculature from GBM or normal brain tissue derived from neurosurgeries was purified and total RNA was isolated from purified microvasculature. The difference of miRNA expression profiles between glioblastoma microvasculature and normal brain capillaries was investigated. It was found that miR-7-5p in GBM microvessels was significantly reduced compared with that in normal brain capillaries. In the in vitro experiments, overexpression of miR-7-5p significantly inhibited human umbilical vein endothelial cell proliferation. Forced expression of miR-7-5p in human umbilical vein endothelial cells in vitro significantly reduced the protein level of RAF1 and repressed the activity of the luciferase, a reporter vector carrying the 3′-untranslated region of RAF1. These findings indicate that RAF1 is one of the miR-7-5p target genes. Furthermore, a significant inverse correlation between miR-7-5p expression and RAF1 protein level in GBM microvasculature was found. These data suggest that miR-7-5p functions as a tumor suppressor gene to regulate GBM microvascular endothelial cell proliferation potentially by targeting the RAF1 oncogene, implicating an important role for miR-7-5p in the pathogenesis of GBM. It may serve as a guide for the antitumor angiogenesis drug development.


Microvasculature MiR-7-5p Glioblastoma RAF1 Proliferation 







Real-time quantitative reverse transcriptase PCR


3′-Untranslated region


Wild type


Mutant type



We thank Yan Song, Meng Zhang, and Yubao Zhang for the administrative and operational support; Drs. Guiyan Xu, Guangming Qu, Deze Jia, and Donghai Wang for their helpful discussions and critical reading of the manuscript.

Ethics approval

Ethics approval was provided by the National Hospital for Neurology and Neurosurgery Research Ethics Committee.

Conflicts of interest

None, there are no conflict of interest.


This work was supported by the National Natural Scientific Foundation of China (NO.81141088) and by the Promotive Research Fund for Excellent Young and Middle-aged Scientists of Shandong Province. (NO. 2004BS02010). JYL was supported by North Shore-LIJ Cancer Institute.


  1. 1.
    Ambros V. The functions of animal microRNAs. Nature. 2004;431(7006):350–5.CrossRefPubMedGoogle Scholar
  2. 2.
    Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell. 2004;116(2):281–97.CrossRefPubMedGoogle Scholar
  3. 3.
    Pasquinelli AE, Hunter S, Bracht J. MicroRNAs: a developing story. Curr Opin Genet Dev. 2005;15(2):200–5.CrossRefPubMedGoogle Scholar
  4. 4.
    Ahmed FE. Role of miRNA in carcinogenesis and biomarker selection: a methodological view. Expert Rev Mol Diagn. 2007;7(5):569–603.CrossRefPubMedGoogle Scholar
  5. 5.
    Croce CM, Calin GA. miRNAs, cancer, and stem cell division. Cell 2005; 122 (1) :6–7.Google Scholar
  6. 6.
    Gregory RI, Shiekhattar R. MicroRNA biogenesis and cancer. Cancer Res. 2005;65(9):3509–12.CrossRefPubMedGoogle Scholar
  7. 7.
    Croce CM. Causes and consequences of microRNA dysregulation in cancer. Nat Rev Genet. 2009;10(10):704–14.CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Van Meir EG, Hadjipanayis CG, Norden AD, Shu HK, Wen PY, Olson JJ. Exciting new advances in neuro-oncology: the avenue to a cure for malignant glioma. CA Cancer J Clin. 2010;60(3):166–93.Google Scholar
  9. 9.
    Kraus JA, Lamszus K, Glesmann N, Beck M, Wolter M, Sabel M, et al. Molecular genetic alterations in glioblastomas with oligodendroglial component. Acta Neuropathol. 2001;101(4):311–20.Google Scholar
  10. 10.
    Fuller GN, The WHO. Classification of tumours of the central nervous system, 4th edition. Arch Pathol Lab Med. 2008;132(6):906.PubMedGoogle Scholar
  11. 11.
    Cancer Genome Atlas Research Network. Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature. 2008;455(7216):1061–8.CrossRefGoogle Scholar
  12. 12.
    Zhang C, Li C, Li J, Han J, Shang D, Zhang Y, et al. Identification of miRNA-mediated core gene module for glioma patient prediction by integrating high-throughput miRNA, mRNA expression and pathway structure. PLoS One. 2014;9(5):e96908.Google Scholar
  13. 13.
    Li JY, Boado RJ, Pardridge WM. Blood–brain barrier genomics. J Cereb Blood Flow Metab. 2001;21(1):61–8.CrossRefPubMedGoogle Scholar
  14. 14.
    Kefas B, Godlewski J, Comeau L, Li Y, Abounader R, Hawkinson M, Lee J, Fine H, Chiocca EA, Lawler S, Purow B. microRNA-7 inhibits the epidermal growth factor receptor and the Akt pathway and is down-regulated in glioblastoma. Cancer Res 2008; 68 (10) :3566–3572Google Scholar
  15. 15.
    Kyriakis JM. The integration of signaling by multiprotein complexes containing Raf kinases. Biochim Biophys Acta. 2007;1773(8):1238–47.CrossRefPubMedGoogle Scholar
  16. 16.
    Leicht DT, Balan V, Kaplun A, Singh-Gupta V, Kaplun L, Dobson M, et al. Raf kinases: function, regulation and role in human cancer. Biochim Biophys Acta. 2007;1773(8):1196–212.Google Scholar
  17. 17.
    McPhillips F, Mullen P, MacLeod KG, Sewell JM, Monia BP, Cameron DA, et al. Raf-1 is the predominant Raf isoform that mediates growth factor-stimulated growth in ovarian cancer cells. Carcinogenesis. 2006;27(4):729–39.Google Scholar
  18. 18.
    Zebisch A, Troppmair J. Back to the roots: the remarkable RAF oncogene story. Cell Mol Life Sci. 2006;63(11):1314–30.CrossRefPubMedGoogle Scholar
  19. 19.
    Pal A, Ahmad A, Khan S, Sakabe I, Zhang C, Kasid UN, et al. Systemic delivery of RafsiRNA using cationic cardiolipin liposomes silences Raf-1 expression and inhibits tumor growth in xenograft model of human prostate cancer. Int J Oncol. 2005;26(4):1087–91.Google Scholar
  20. 20.
    Fushimi K, Nakashima S, You F, Takigawa M, Shimizu K. Prostaglandin E2 downregulates TNF-alpha-induced production of matrix metalloproteinase-1 in HCS-2/8 chondrocytes by inhibiting Raf-1/MEK/ERK cascade through EP4 prostanoid receptor activation. J Cell Biochem. 2007;100(3):783–93.Google Scholar
  21. 21.
    von Kriegsheim A, Pitt A, Grindlay GJ, Kolch W, Dhillon AS. Regulation of the Raf-MEK-ERK pathway by protein phosphatase 5. Nat Cell Biol. 2006;8(9):1011–6.CrossRefGoogle Scholar
  22. 22.
    von B, V, Dubben S, Engelhardt G, Hebel S, Plumakers B, Heine H, Rink L, Haase H. Zinc-dependent suppression of TNF-alpha production is mediated by protein kinase A-induced inhibition of Raf-1 , I kappa B kinase beta, and NF-kappa B. J Immunol 2007; 179 (6) :4180–4186Google Scholar
  23. 23.
    Li Y, Levesque LO, Anand-Srivastava MB. Epidermal growth factor receptor transactivation by endogenous vasoactive peptides contributes to hyperproliferation of vascular smooth muscle cells of SHR. Am J Physiol Heart Circ Physiol. 2010;299(6):H1959–67.CrossRefPubMedGoogle Scholar
  24. 24.
    Maretzky T, Evers A, Zhou W, Swendeman SL, Wong PM, Rafii S, et al. Migration of growth factor-stimulated epithelial and endothelial cells depends on EGFR transactivation by ADAM17. Nat Commun. 2011;2:229.Google Scholar
  25. 25.
    Kyriakakis E, Cavallari M, Pfaff D, Fabbro D, Mestan J, Philippova M, et al. IL-8-mediated angiogenic responses of endothelial cells to lipid antigen activation of iNKT cells depend on EGFR transactivation. J Leukoc Biol. 2011;90(5):929–39.Google Scholar
  26. 26.
    Guang-Wu H, Sunagawa M, Jie-En L, Shimada S, Gang Z, Tokeshi Y, et al. The relationship between microvessel density, the expression of vascular endothelial growth factor (VEGF), and the extension of nasopharyngeal carcinoma. Laryngoscope. 2000;110(12):2066–9.Google Scholar
  27. 27.
    Goel HL, Mercurio AM. VEGF targets the tumour cell. Nat Rev Cancer. 2013;13(12):871–82.CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Chen Y, Rabson AB, Gorski DH. MEOX2 regulates nuclear factor-kappaB activity in vascular endothelial cells through interactions with p65 and IkappaBbeta. Cardiovasc Res. 2010;87(4):723–31.CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Chiplunkar AR, Curtis BC, Eades GL, Kane MS, Fox SJ, Haar JL, et al. The Kruppel-like factor 2 and Kruppel-like factor 4 genes interact to maintain endothelial integrity in mouse embryonic vasculogenesis. BMC Dev Biol. 2013;13:40.Google Scholar
  30. 30.
    Zhang Y, Chen N, Zhang J, Tong Y. Hsa-let-7 g miRNA targets caspase-3 and inhibits the apoptosis induced by ox-LDL in endothelial cells. Int J Mol Sci. 2013;14(11):22708–20.CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Lee JW, Soung YH, Kim SY, Nam SW, Park WS, Lee JY, et al. Absence of JAK2 V617F mutation in gastric cancers. Acta Oncol. 2006;45(2):222–3.Google Scholar
  32. 32.
    Orom UA, Kauppinen S, Lund AH. LNA-modified oligonucleotides mediate specific inhibition of microRNA function. Gene. 2006;372:137–41.CrossRefPubMedGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2014

Authors and Affiliations

  • Zhiguo Liu
    • 1
  • Yuguang Liu
    • 1
  • Lianling Li
    • 1
  • Zhenkuan Xu
    • 1
  • Baibin Bi
    • 1
  • Yunyan Wang
    • 1
  • Jian Yi Li
    • 2
  1. 1.Department of Neurosurgery, Qilu Hospital of Shandong UniversityBrain Science Research Institute of Shandong UniversityJinanPeople’s Republic of China
  2. 2.Department of Pathology and Laboratory Medicine, North Shore-Long Island Jewish Health System, Lake SuccessHofstra North Shore-LIJ School of MedicineNew YorkUSA

Personalised recommendations