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The vascular delta-like ligand-4 (DLL4)-Notch4 signaling correlates with angiogenesis in primary glioblastoma: an immunohistochemical study

  • Original Article
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Tumor Biology

Abstract

Delta-like ligand-4 (DLL4)-Notch signaling is known to play a pivotal role in the regulation of tumor angiogenesis. We had previously found that DLL4 was overexpressed, while Notch1 receptor, which binds to DLL4 during angiogenesis, was absent in the majority of human primary glioblastomas. Thus, DLL4-Notch signaling pathway in the regulation of tumor angiogenesis in primary glioblastoma remains unknown. Tumor tissues from 70 patients with primary glioblastoma were analyzed by immunohistochemistry for expression of components of DLL4-Notch signaling, vascular endothelial growth factor (VEGF), and microvessel density (MVD). Immunohistochemistry results showed that the positive staining of DLL4 and Notch4 was primarily distributed in tumor vascular endothelial cells but rarely detected in tumor cells. However, VEGF, hairy/enhancer of split-1 (HES1; a target gene of Notch signaling), and Notch1–3 expression was seen in both tumor vascular endothelial cells and tumor cells. Univariate analysis showed that the expression levels of VEGF and DLL4, HES1, and Notch4 in tumor endothelial cells were significantly associated with MVD in primary glioblastoma (P < 0.001). Binary logistic regression analysis showed that high expression levels of DLL4, HES1, and Notch4 in tumor endothelial cells were associated with a decrease of MVD in primary glioblastoma, while MVD increased with elevated VEGF expression in contrast. In addition, DLL4, Notch4, and HES1 expression were positively correlated in tumor vascular endothelial cells (P < 0.05). We conclude that the vascular DLL4-Notch4 signaling and VEGF signaling complementing each other plays an important role in the progression of tumor angiogenesis in primary glioblastoma.

A, positive staining of DLL4 in human kidney; B, positive staining of VEGF in human breast cancer; C, positive staining of CD34 in human lung cancer; D, positive staining of HES1 in human breast cancer; E-H, positive staining of Notch1-4: E-F in human lung cancer; G-H in human kidney

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Abbreviations

MVD:

Microvessel density

VEGF:

Vascular endothelial growth factor

DLL4:

Delta-like ligand-4

NICD:

Notch intracellular domain

WHO:

World Health Organization

PBS:

Phosphate buffer solution

DAB:

Diaminobenzidine

EC:

Endothelial cells

TC:

Tumor cell

GSC:

Glioma stem cell

PI3k:

Phosphatidylinositol 3-kinase

EGFR:

Epidermal growth factor receptor

MMP-9:

Matrix metalloprotein-9

References

  1. Wen PY, Kesari S. Malignant gliomas in adults. N Engl J Med. 2008;359:492–507.

    Article  CAS  PubMed  Google Scholar 

  2. Gerstner ER, Batchelor TT. Antiangiogenic therapy for glioblastoma. Cancer J. 2012;18:45–50.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Carmeliet P. Angiogenesis in life, disease and medicine. Nature. 2005;438:932–6.

    Article  CAS  PubMed  Google Scholar 

  4. Yamamoto S, Schulze KL, Bellen HJ. Introduction to notch signaling. Methods Mol Biol. 2014;1187:1–14.

    Article  CAS  PubMed  Google Scholar 

  5. Artavanis-Tsakonas S, Rand MD, Lake RJ. Notch signaling: cell fate control and signal integration in development. Science. 1999;284:770–6.

    Article  CAS  PubMed  Google Scholar 

  6. Fleming RJ. Structural conservation of notch receptors and ligands. Semin Cell Dev Biol. 1998;9:599–607.

    Article  CAS  PubMed  Google Scholar 

  7. Iso T, Kedes L, Hamamori Y. HES and HERP families: multiple effectors of the notch signaling pathway. J Cell Physiol. 2003;194:237–55.

    Article  CAS  PubMed  Google Scholar 

  8. Kume T. Novel insights into the differential functions of notch ligands in vascular formation. J Angiogenes Res. 2009;1:8.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Lobov IB, Renard RA, Papadopoulos N, Gale NW, Thurston G, Yancopoulos GD, et al. Delta-like ligand 4 (Dll4) is induced by VEGF as a negative regulator of angiogenic sprouting. Proc Natl Acad Sci U S A. 2007;104:3219–24.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Tung JJ, Tattersall IW, Kitajewski J. Tips, stalks, tubes: notch-mediated cell fate determination and mechanisms of tubulogenesis during angiogenesis. Cold Spring Harb Perspect Med. 2012;2:a006601.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Bolos V, Grego-Bessa J, de la Pompa JL. Notch signaling in development and cancer. Endocr Rev. 2007;28:339–63.

    Article  CAS  PubMed  Google Scholar 

  12. South AP, Cho RJ, Aster JC. The double-edged sword of Notch signaling in cancer. Semin Cell Dev Biol. 2012;23:458–64.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Thurston G, Noguera-Troise I, Yancopoulos GD. The delta paradox: Dll4 blockade leads to more tumour vessels but less tumour growth. Nat Rev Cancer. 2007;7:327–31.

    Article  CAS  PubMed  Google Scholar 

  14. Stockhausen MT, Kristoffersen K, Poulsen HS. The functional role of notch signaling in human gliomas. Neuro-Oncology. 2010;12:199–211.

    Article  CAS  PubMed  Google Scholar 

  15. Poellinger L, Lendahl U. Modulating Notch signaling by pathway-intrinsic and pathway-extrinsic mechanisms. Curr Opin Genet Dev. 2008;18:449–54.

    Article  CAS  PubMed  Google Scholar 

  16. Kangsamaksin T, Murtomaki A, Kofler NM, Cuervo H, Chaudhri RA, Tattersall IW, et al. NOTCH decoys that selectively block DLL/NOTCH or JAG/NOTCH disrupt angiogenesis by unique mechanisms to inhibit tumor growth. Cancer Discov. 2015;5:182–97.

    Article  CAS  PubMed  Google Scholar 

  17. Pedrosa AR, Trindade A, Fernandes AC, Carvalho C, Gigante J, Tavares AT, et al. Endothelial Jagged1 antagonizes Dll4 regulation of endothelial branching and promotes vascular maturation downstream of Dll4/Notch1. Arterioscler Thromb Vasc Biol. 2015;35:1134–46.

    Article  CAS  PubMed  Google Scholar 

  18. Qiu XX, Chen L, Wang CH, Lin ZX, Zhou CF, Liu SY, et al. High delta-like ligand 4 (DLL4) is correlated with peritumoral brain edema and predicts poor prognosis in primary glioblastoma. Medicine. 2014;93:e57.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Li ZQ, Gong LL, Wen ZH, Wang J, Xu CS, Huang XD. Delta-like ligand 4 correlates with endothelial proliferation and vessel maturation in human malignant glioma. Onkologie. 2012;35:763–8.

    Article  CAS  PubMed  Google Scholar 

  20. Jubb AM, Browning L, Campo L, Turley H, Steers G, Thurston G, et al. Expression of vascular Notch ligands Delta-like 4 and Jagged-1 in glioblastoma. Histopathology. 2012;60:740–7.

    Article  PubMed  Google Scholar 

  21. Li Z, Wang J, Gong L, Wen Z, Xu C, Huang X. Correlation of Delta-like ligand 4 (DLL4) with VEGF and HIF-1alpha expression in human glioma. Asian Pac J Cancer Prev. 2011;12:215–8.

    PubMed  Google Scholar 

  22. Cheung HC, Corley LJ, Fuller GN, McCutcheon IE, Cote GJ. Polypyrimidine tract binding protein and Notch1 are independently re-expressed in glioma. Mod Pathol. 2006;19:1034–41.

    CAS  PubMed  Google Scholar 

  23. Dell’albani P, Rodolico M, Pellitteri R, Tricarichi E, Torrisi SA, D’Antoni S, et al. Differential patterns of NOTCH1-4 receptor expression are markers of glioma cell differentiation. Neuro-Oncology. 2014;16:204–16.

    Article  CAS  PubMed  Google Scholar 

  24. Li J, Cui Y, Gao G, Zhao Z, Zhang H, Wang X. Notch1 is an independent prognostic factor for patients with glioma. J Surg Oncol. 2011;103:813–7.

    Article  CAS  PubMed  Google Scholar 

  25. Louis DN, Ohgaki H, Wiestler OD, Cavenee WK, Burger PC, Jouvet A, et al. The 2007 who classification of tumours of the central nervous system. Acta Neuropathol. 2007;114:97–109.

    Article  PubMed  PubMed Central  Google Scholar 

  26. Piperi C, Samaras V, Levidou G, Kavantzas N, Boviatsis E, Petraki K, et al. Prognostic significance of IL-8-STAT-3 pathway in astrocytomas: correlation with IL-6, VEGF and microvessel morphometry. Cytokine. 2011;55:387–95.

    Article  CAS  PubMed  Google Scholar 

  27. Weidner N. Current pathologic methods for measuring intratumoral microvessel density within breast carcinoma and other solid tumors. Breast Cancer Res Treat. 1995;36:169–80.

    Article  CAS  PubMed  Google Scholar 

  28. Xu P, Qiu M, Zhang Z, Kang C, Jiang R, Jia Z, et al. The oncogenic roles of Notch1 in astrocytic gliomas in vitro and in vivo. J Neuro-Oncol. 2010;97:41–51.

    Article  CAS  Google Scholar 

  29. Purow BW, Haque RM, Noel MW, Su Q, Burdick MJ, Lee J, et al. Expression of Notch-1 and its ligands, Delta-like-1 and Jagged-1, is critical for glioma cell survival and proliferation. Cancer Res. 2005;65:2353–63.

    Article  CAS  PubMed  Google Scholar 

  30. Chu J, Jeffries S, Norton JE, Capobianco AJ, Bresnick EH. Repression of activator protein-1-mediated transcriptional activation by the Notch-1 intracellular domain. J Biol Chem. 2002;277:7587–97.

    Article  CAS  PubMed  Google Scholar 

  31. Albig AR, Becenti DJ, Roy TG, Schiemann WP. Microfibril-associate glycoprotein-2 (MAGP-2) promotes angiogenic cell sprouting by blocking notch signaling in endothelial cells. Microvasc Res. 2008;76:7–14.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Liu ZJ, Xiao M, Balint K, Soma A, Pinnix CC, Capobianco AJ, et al. Inhibition of endothelial cell proliferation by Notch1 signaling is mediated by repressing MAPK and PI3K/Akt pathways and requires MAML1. FASEB J. 2006;20:1009–11.

    Article  CAS  PubMed  Google Scholar 

  33. Indulekha CL, Divya TS, Divya MS, Sanalkumar R, Rasheed VA, Dhanesh SB, et al. Hes-1 regulates the excitatory fate of neural progenitors through modulation of Tlx3 (HOX11L2) expression. Cell Mol Life Sci. 2012;69:611–27.

    Article  CAS  PubMed  Google Scholar 

  34. Yao J, Zheng K, Li C, Liu H, Shan X. Interference of Notch1 inhibits the growth of glioma cancer cells by inducing cell autophagy and down-regulation of Notch1-Hes-1 signaling pathway. Med Oncol. 2015;32:610.

    PubMed  Google Scholar 

  35. Benedito R, Roca C, Sorensen I, Adams S, Gossler A, Fruttiger M, et al. The notch ligands Dll4 and Jagged1 have opposing effects on angiogenesis. Cell. 2009;137:1124–35.

    Article  CAS  PubMed  Google Scholar 

  36. Hellstrom M, Phng LK, Hofmann JJ, Wallgard E, Coultas L, Lindblom P, et al. Dll4 signalling through Notch1 regulates formation of tip cells during angiogenesis. Nature. 2007;445:776–80.

    Article  CAS  PubMed  Google Scholar 

  37. Hernandez SL, Banerjee D, Garcia A, Kangsamaksin T, Cheng WY, Anastassiou D, et al. Notch and VEGF pathways play distinct but complementary roles in tumor angiogenesis. Vasc Cell. 2013;5:17.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Krebs LT, Xue Y, Norton CR, Shutter JR, Maguire M, Sundberg JP, et al. Notch signaling is essential for vascular morphogenesis in mice. Genes Dev. 2000;14:1343–52.

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Suchting S, Freitas C, le Noble F, Benedito R, Breant C, Duarte A, et al. The Notch ligand Delta-like 4 negatively regulates endothelial tip cell formation and vessel branching. Proc Natl Acad Sci U S A. 2007;104:3225–30.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Zhu TS, Costello MA, Talsma CE, Flack CG, Crowley JG, Hamm LL, et al. Endothelial cells create a stem cell niche in glioblastoma by providing notch ligands that nurture self-renewal of cancer stem-like cells. Cancer Res. 2011;71:6061–72.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Kenig S, Alonso MB, Mueller MM, Lah TT. Glioblastoma and endothelial cells cross-talk, mediated by SDF-1, enhances tumour invasion and endothelial proliferation by increasing expression of cathepsins B, S, and MMP-9. Cancer Lett. 2010;289:53–61.

    Article  CAS  PubMed  Google Scholar 

  42. Cheng L, Huang Z, Zhou W, Wu Q, Donnola S, Liu JK, et al. Glioblastoma stem cells generate vascular pericytes to support vessel function and tumor growth. Cell. 2013;153:139–52.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Li JL, Sainson RC, Shi W, Leek R, Harrington LS, Preusser M, et al. Delta-like 4 Notch ligand regulates tumor angiogenesis, improves tumor vascular function, and promotes tumor growth in vivo. Cancer Res. 2007;67:11244–53.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This study was supported by the National Natural Science Foundation of China (No. 21435002). The authors would like to thank the Department of Pathology, First Affiliated Hospital, Fujian Medical University, for their help in immunohistochemistry design and the Public Health School, Fujian Medical University, Fujian, China, for assistance in data processing and statistical analysis.

Authors’ contributions

JFZ and YC analyzed the data and drafted the manuscript. YC, JFZ, and XXQ performed the experiments. XFW and JDZ provided material support. WLT, GSL, and JHH contributed to data collection and helped in the analysis. ZXL conceived the study and participated in its design and coordination. All authors read and approved the final manuscript.

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    Authors and Affiliations

    1. The First Clinical Medical College, Fujian Medical University, Fuzhou, Fujian, 350005, China

      Jin-feng Zhang & Zhi-xiong Lin

    2. Department of Neurosurgery, The First Affiliated Hospital, Fujian Medical University, Fuzhou, Fujian, 350005, China

      Yao Chen, Wen-long Tang, Jian-dong Zhang, Jian-huang Huang & Guo-shi Lin

    3. Department of Neurosurgery, The Children’s Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, 310051, China

      Xian-xin Qiu

    4. Department of Pathology, The First Affiliated Hospital, Fujian Medical University, Fuzhou, Fujian, 350005, China

      Xing-fu Wang

    5. Department of Neurosurgery, Beijing Sanbo Brain Hospital, Capital Medical University, Beijing, 100093, China

      Zhi-xiong Lin

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    Jin-feng Zhang and Yao Chen contributed equally to this work.

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    Zhang, Jf., Chen, Y., Qiu, Xx. et al. The vascular delta-like ligand-4 (DLL4)-Notch4 signaling correlates with angiogenesis in primary glioblastoma: an immunohistochemical study. Tumor Biol. 37, 3797–3805 (2016). https://doi.org/10.1007/s13277-015-4202-8

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