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Involvement of integrin-linked kinase in capillary/tube-like network formation of human vascular endothelial cells

Biological Procedures Online volume 7, pages 41–47 (2005)Cite this article

Abstract

Angiogenesis is a complex process involving an ECM and vascular endothelial cells (EC), and is regulated by various angiogenic factors including VEGF. The ability to form a capillary/tube-like network is a specialized function of EC. Therefore, in vitro angiogenesis was assessed by a capillary/tube-like network formation assay. There are three angiogenic parameters: capillary length, number of capillaries, and relative capillary area per field. We evaluated capillary length per field in the assay. VEGF promoted capillary/tube-like network formation of EC in a type I collagen gel matrix in vitro. Moreover, we demonstrated the involvement of ILK in a VEGF signaling pathway mediating capillary/tube-like network formation of EC using dominant-negative, kinase deficient ILK. This is a straightforward assay to monitor responses of human vascular endothelial cells.

Abbreviations

ECM:

extracellular matrix

HUVEC:

human umbilical vein endothelial cells

ILK:

integrin-linked kinase

KD:

kinase deficient

PI 3-kinase:

phosphatidylinositol 3-kinase

PKB:

protein kinase B

VEGF:

vascular endothelial growth factor

References

  1. Hanahan D, Folkman J. Patterns of and emerging mechanisms of the angiogenesis switch during tumorgenesis. Cell 1996; 86:353–364.

    Article  PubMed  CAS  Google Scholar 

  2. Kumer R, Yoneda J, Bucana CD, Fidler IJ. Regulation of distinct steps of angiogenesis by different angiogenic molecules. Int J Oncol 1998; 12:749–757.

    Google Scholar 

  3. Vinals F, Pouyssegur J. Transforming growth factor beta1 (TGF-beta1) promotes endothelial cell survival during in vitro angiogenesis via an autocrine mechanism implicating TGF-alpha signaling. Mol Cell Biol 2001; 21:7218–7230.

    Article  PubMed  CAS  Google Scholar 

  4. Plate KH, Breier G, Millauer B, Ullrich A, Risau W. Up-regulation of vascular endothelial growth factor and its cognate receptors in a rat glioma model of tumor angiogenesis. Cancer Res 1993; 53:5822–5827.

    PubMed  CAS  Google Scholar 

  5. Takahashi A, Sasaki H, Kim SJ, Tobisu K, Kakizoe T, Tsukamoto T, Kumamoto Y, Sugimura T, Terada M. Markedly increased amounts of messenger RNAs for vascular endothelial growth factor and placenta growth factor in renal cell carcinoma associated with angiogenesis. Cancer Res 1994; 54:4233–4237.

    PubMed  CAS  Google Scholar 

  6. Anan K, Morisaki T, Katano M, Ikubo A, Kitsuki H, Uchiyama A, Kuroki S, Tanaka M, Torisu M. Vascular endothelial growth factor and platelet-derived growth factor are potential angiogenic and metastatic factors in human breast cancer. Surgery 1996; 119:333–339.

    Article  PubMed  CAS  Google Scholar 

  7. Albelda SM, Buck CA. Integrins and other cell adhesion molecules. FASEB J 1990; 4:2868–2880.

    PubMed  CAS  Google Scholar 

  8. Akiyama SK, Nagata K, Yamada KM. Cell surface receptors for extracellular matrix components. Biochim Biophys Acta 1990; 1031:91–110.

    Google Scholar 

  9. Schwartz MA, Schaller MD, Ginsberg MH. Integrins: emerging paradigms of signal transduction. Annu Rev Cell Dev Biol 1995; 11:549–599.

    Article  PubMed  CAS  Google Scholar 

  10. Hannigan GE, Leung-Hagesteijin C, Fitz-Gibbon L, Coppolino MG, Radeva G, Filmus J, Bell JC, Dedhar S. Regulation of cell adhesion and anchorage-dependent growth by a new beta 1-integrin-linked protein kinase. Nature 1996; 379:91–96.

    Article  PubMed  CAS  Google Scholar 

  11. Delcommenne M, Tan C, Gray V, Rue L, Woodgett J, Dedhar S. Phosphoinositide-3-OH kinase-dependent regulation of glycogen synthase kinase 3 and protein kinase B/Akt by the integrin linked kinase. Proc Natl Acad Sci USA 1998; 95:1121–1126.

    Article  Google Scholar 

  12. Kaneko Y, Kitazato K, Basaki Y. Integrin-linked kinase regulates vascular morphogenesis induced by vascular endothelial growth factor. J Cell Sci 2004;117:407–415.

    Article  PubMed  CAS  Google Scholar 

  13. Byzova TV, Goldman CK, Pampori N, Thomas KA, Bett A, Shattil SJ, Plow EF. A mechanism for modulation of cellular responses to VEGF: activation of the integrins. Mol Cell 2000; 6:851–860.

    PubMed  CAS  Google Scholar 

  14. Wu C, Keightley SY, Leung-Hagesteijin C, Radeva G, Coppolino M, Goicoechea S, McDonald JA, Dedhar S. Integrin-linked protein kinase regulates fibronectin matrix assembly, E-cadherin expression, and tumorigenicity. J Biol Chem 1998; 273:528–536.

    Article  PubMed  CAS  Google Scholar 

  15. Novak A, Hsu SC, Leung-Hagesteijn C, Radeva G, Papkoff J, Montesano R, Roskelley C, Grosschedl R, Dedhar S. Cell adhesion and the integrin-linked kinase regulate the LEF-1 and beta-catenin signaling pathways. Proc Natl Acad Sci USA 1998; 95:4374–4379.

    Article  PubMed  CAS  Google Scholar 

  16. Basaki Y, Chikahisa L, Aoyagi K, Miyadera K, Yonekura K, Hashimoto A, Okabe S, Wierzba K, Yamada Y. Gamma-hydroxybutyric acid and 5-fluorouracil, metabolites of UFT, inhibit the angiogenesis induced by vascular endothelial growth factor. Angiogenesis 2001; 4:163–173.

    Article  PubMed  CAS  Google Scholar 

  17. Orkin RW, Gehron P, McGoodwin EB, Martin GR, Valentine T, Swarm R. A murine tumor producing a matrix of basement membrane. J Exp Med 1977;145:204–220.

    Article  PubMed  CAS  Google Scholar 

  18. Vukicevic S, Kleinman HK, Luyten FP, Roberts AB, Roche NS, Reddi AH. Identification of multiple active growth factors in basement membrane Matrigel suggests caution in interpretation of cellular activity related to extracellular matrix components. Exp Cell Res 1992; 202:1–8.

    Article  PubMed  CAS  Google Scholar 

  19. Persad S, Attwell S, Gray V, Mawji N, Deng JT, Leung D, Yan J, Sanghera J, Walsh MP, Dedhar S. Regulation of protein kinase B/Akt-serine 473 phosphorylation by integrin-linked kinase: critical roles for kinase activity and amino acids arginine 211 and serine 343. J Biol Chem 2001; 276:27462–27469.

    Article  PubMed  CAS  Google Scholar 

  20. Alessi DR, James SR, Downes CP, Holmes AB, Gaffney PR, Reese CB, Cohen P. Characterization of a 3-phosphoinositide-dependent protein kinase which phosphorylates and activates protein kinase B alpha. Curr Biol 1997; 7:261–269.

    Article  PubMed  CAS  Google Scholar 

  21. Friedrich EB, Liu E, Sinha S, Cook S, Milstone DS, MacRae CA, Mariotti M, Kuhlencordt PJ, Force T, Rosenzweig A, St-Arnaud R, Dedhar S, Gerszten RE. Integrin-linked kinase regulates endothelial cell survival and vascular development. Mol Cell Biol 2004; 24:8134–8144.

    Article  PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Advanced Research Laboratory, Taiho Pharmaceutical Co., Ltd., 357-8527, Saitama, Japan

    Motomu Watanabe & Mamoru Kiniwa

  2. Drug Discovery Laboratory, Taiho Pharmaceutical Co., Ltd., 357-8527, Saitama, Japan

    Yayoi Fujioka-Kaneko

  3. Biochemical Laboratory, Nitta Gelatin Inc., 581-0024, Osaka, Japan

    Hisayuki Kobayashi

  4. Research Center for Innovative Cancer Therapy, Kurume University, 830-0011, Fukuoka, Japan

    Michihiko Kuwano

  5. Station-II for Collaborative Research, Kyushu University, 812-8582, Fukuoka, Japan

    Michihiko Kuwano

  6. Advanced Research Laboratory, Taiho Pharmaceutical Co., Ltd., 357-8527, Saitama, Japan

    Yuji Basaki Ph.D.

  7. Station-II for Collaborative Research, Jyushu University, 812-8582, Fukuoka, Japan

    Yuji Basaki Ph.D.

  8. Department of Medical Biochemistry, Graduate School of Medical Science, Kyushu University, 3-1-1 Maidashi Higashi-ku, 812-8582, Fukuoka, Japan

    Yuji Basaki Ph.D.

Authors
  1. Motomu Watanabe
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  2. Yayoi Fujioka-Kaneko
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  3. Hisayuki Kobayashi
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  4. Mamoru Kiniwa
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  5. Michihiko Kuwano
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  6. Yuji Basaki Ph.D.
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Corresponding author

Correspondence to Yuji Basaki Ph.D..

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Watanabe and Basaki contributed equally to this manuscript.

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Open Access This article is published under license to BioMed Central Ltd. This is an Open Access article is distributed under the terms of the Creative Commons Attribution License ( https://creativecommons.org/licenses/by/2.0 ), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Watanabe, M., Fujioka-Kaneko, Y., Kobayashi, H. et al. Involvement of integrin-linked kinase in capillary/tube-like network formation of human vascular endothelial cells. Biol. Proced. Online 7, 41–47 (2005). https://doi.org/10.1251/bpo104

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  • DOI: https://doi.org/10.1251/bpo104

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Biological Procedures Online

ISSN: 1480-9222

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