Skip to main content
SpringerLink
Log in

The glycan profile of endothelial cells in the present of tumor-conditioned medium and potential roles of β-1,6-GlcNAc branching on HUVEC conformation

  • Published:
Molecular and Cellular Biochemistry Aims and scope Submit manuscript

Abstract

Endothelium plays a vital role in the logistics of the immune system, as well as the maintenance of the homeostasis. The major objective of this study is to unravel the relationship between expression changes of carbohydrate structures and the dysfunction of human umbilical vein endothelial cells (HUVEC) stimulated with tumor-conditioned medium (TCM), which is involved in tumor cell extravasation. Using flow cytometry (FCM) assay, the expression profiles of a selected group of 9 carbohydrate structures have been determined in HUVEC under control conditions and TCM-treated conditions, six of which increased significantly in expression after induction. Particularly, the expression level of β-1,6-GlcNAc branching glycan was extremely higher after the stimulation. In parallel, the conformation change of HUVEC monolayer has been detected with inverted phase contrast microscopy and confocal microscopy. Under TCM stimulation, the actin cytoskeleton underwent rearrangement and formed abundant stress fiber within cells; therefore cell contraction was induced, which resulted in paracellular gap formation and barrier dysfunction. We furthered our study to investigate the mechanism underlying the conformation change of HUVEC. The results demonstrated that TCM induced the increase in β-1,6-GlcNAc branching expression of PECAM-1, accompanied by the tyrosine phosphorylation of PECAM-1. The downstream effector RhoA was activated in consequence of the activation of PECAM-1. In conclusion, our results strongly suggested that the carbohydrate composition of endothelial cell surface is very important for the cells to exert their physiological effects correlated with cancer extravasation.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

Abbreviations

HUVEC:

Human umbilical vein endothelial cells

TCM:

Tumor-conditioned medium

FBS:

Fetal bovine serum

FITC:

Fluorescein isothyocianate

SDS-PAGE:

Sodium dodecyl sulfate polyacrylamide gel electrophoresis

mAb:

Monoclonal antibody

pAb:

Polyclonal antibody

HRP:

Horseradish peroxidase

ECGS:

Endothelial cell growth supplement

PBS:

Phosphate-buffered saline

BSA:

Bovine serum albumin

TBS:

Tris-buffered saline

Fn:

Fibronectin

GnT-V:

N-acetylglucosaminyltransferase V

MTT:

3-[4,5-Dimethylthiazol-2-yl]-2,5-diphenyltetrazoliun bromide

References

  1. Dube DH, Bertozzi CR (2005) Glycans in cancer and inflammation—potential for therapeutics and diagnostics. Nat Rev Drug Discov 4:477–488

    Article  CAS  PubMed  Google Scholar 

  2. Zhao Y, Li J, Wang J, Xing Y, Geng M (2007) Role of cell surface oligosaccharides of mouse mammary tumor cell lines in cancer metastasis. Indian J Biochem Biophys 44:145–151

    CAS  PubMed  Google Scholar 

  3. Guo HB, Lee I, Kamar M, Pierce M (2003) N-acetylglucosaminyltransferase V expression levels regulate cadherin-associated homotypic cell-cell adhesion and intracellular signaling pathways. J Biol Chem 278:52412–52424

    Article  CAS  PubMed  Google Scholar 

  4. Liwosz A, Lei T, Kukuruzinska MA (2006) N-glycosylation affects the molecular organization and stability of E-cadherin junctions. J Biol Chem 281:23138–23149

    Article  CAS  PubMed  Google Scholar 

  5. Yang J, Mani SA, Donaher JL, Ramaswamy S, Itzykson RA, Come C, Savagner P, Gitelman I, Richardson A, Weinberg RA (2004) Twist, a master regulator of morphogenesis, plays an essential role in tumor metastasis. Cell 117:927–939

    Article  CAS  PubMed  Google Scholar 

  6. Garcia-Vallejo JJ, Van Dijk W, Van Het Hof B et al (2006) Activation of human endothelial cells by tumor necrosis factor-α results in profound changes in the expression of glycosylation-related genes. J Cell Physiol 206:203–210

    Article  CAS  PubMed  Google Scholar 

  7. Luster AD, Alon R, Andrian UH (2005) Immune cell migration in inflammation: present and future therapeutic targets. Nature Immunol 6:1182–1190

    Article  CAS  Google Scholar 

  8. Dull RO, Dinavahi R, Schwartz L et al (2003) Lung endothelial heparan sulfates mediate cationic peptideinduced barrier dysfunction: a new role for the glycocalyx. Am J Physiol Lung Cell Mol Physiol 285:986–995

    Google Scholar 

  9. Miao BC, Li J, Fu XY et al (2005) Sulfated polymannuroguluronate, a novel anti-AIDS drug candidate, inhibits T cell apoptosis by combating oxidative damage of mitochondria. Mol Pharmacol 68:1716–1727

    CAS  PubMed  Google Scholar 

  10. Bruewer M, Hopkins AM, Hobert ME et al (2004) RhoA, Rac1, and Cdc42 exert distinct effects on epithelial barrier via selective structural and biochemical modulation of junctional proteins and F-actin. Am J Physiol Cell Physiol 287:327–335

    Article  Google Scholar 

  11. Damiani D, Michieli M, Michelutti A et al (2004) Antibody binding capacity for evaluation of MDR-related proteins in acute promyelocytic leukemia: onset versus relapse expression. Clin Cytom 59:40–45

    Article  Google Scholar 

  12. Dejana E, Corada M, Lampugnani MG (1995) Endothelial cell-to-cell junctions. FASEB J 9:910–918

    CAS  PubMed  Google Scholar 

  13. Rival Y, Del Maschio A, Rabiet MJ et al (1996) Inhibition of platelet endothelial cell adhesion molecule-1 synthesis and leukocyte transmigration in endothelial cells by the combined action of TNF-α and IFN-γ. J Immunol 157:1233–1241

    CAS  PubMed  Google Scholar 

  14. Waschke J, Curry FE, Adamson RH, Drenckhahn D (2005) Regulation of actin dynamics is critical for endothelial barrier functions. Am J Physiol Heart Circ Physiol 288:1296–1305

    Article  Google Scholar 

  15. Su WH, Chen HN, Jen CH (2002) Differential movements of VE-cadherin and PECAM-1 during transmigration of polymorphonuclear leukocytes through human umbilical vein endothelium. Blood 100:3597–3603

    Article  CAS  PubMed  Google Scholar 

  16. Lu TT, Yan LG, Madri JA (1996) Integrin engagement mediates tyrosine dephosphorylation on platelet-endothelial cell adhesion molecule 1. Proc Natl Acad Sci USA 93:11808–11813

    Article  CAS  PubMed  Google Scholar 

  17. Newton JP, Hunter AP, Simmons DL et al (1999) CD31 (PECAM-1) exists as a dimer and is heavily N-glycosylated. Biochem Biophys Res Commun 261:283–291

    Article  CAS  PubMed  Google Scholar 

  18. Tapon N, Hall A (1997) Rho, Rac, and Cdc42 GTPases regulate the organization of the actin cytoskeleton. Curr Opin Cell Biol 9:86–92

    Article  CAS  PubMed  Google Scholar 

  19. Gratzinger D, Canosa S, Engelhardt B, Madri JA (2003) Platelet endothelial cell adhesion molecule-1 modulates endothelial cell motility through the small G-protein Rho. FASEB J 17:1458–1469

    Article  CAS  PubMed  Google Scholar 

  20. Folkman J, Haudenschild C, Zetter BR (1979) Long-term culture of capillary endothelial cells. Proc Natl Acad Sci USA 76:5217–5221

    Article  CAS  PubMed  Google Scholar 

  21. Papetti M, Herman IM (2002) Mechanisms of normal and tumor-derived angiogenesis. Am J Physiol Cell Physiol 282:947–970

    Google Scholar 

  22. Utoguchi N, Mizuguchi H, Saeki K et al (1995) Tumor-conditioned medium increases macromolecular permeability of endothelial cell monolayer. Cancer Lett 89:7–14

    CAS  PubMed  Google Scholar 

  23. Castilla MA, Neria F, Renedo G et al (2004) Tumor-induced endothelial cell activation: role of vascular endothelial growth factor. Am J Physiol Cell Physiol 286:1170–1176

    Article  Google Scholar 

  24. Cook-Mills JM, Deem TL (2005) Active participation of endothelial cells in inflammation. J Leukoc Biol 77:487–495

    Article  CAS  PubMed  Google Scholar 

  25. O’Brien CD, Lim P, Sun J, Albelda SM (2003) PECAM-1-dependent neutrophil transmigration is independent of monolayer PECAM-1 signaling or localization. Blood 101:2816–2825

    Article  PubMed  Google Scholar 

Download references

Acknowledgment

This research was supported by the National Basic Research Program Grant (973) (2003CB716400).

Author information

Authors and Affiliations

  1. Laboratory of Stress Medicine, Department of Nautical Medicine, Second Military Medical University, Shanghai, China

    Yunli Peng

  2. Laboratory of Molecular Pharmacology, School of Medicine and Pharmacy, Ocean University of China, Qingdao, China

    Yunli Peng & Jing Li

  3. Division of Anti-tumor Pharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China

    Meiyu Geng

Authors
  1. Yunli Peng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jing Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Meiyu Geng
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Yunli Peng or Meiyu Geng.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Peng, Y., Li, J. & Geng, M. The glycan profile of endothelial cells in the present of tumor-conditioned medium and potential roles of β-1,6-GlcNAc branching on HUVEC conformation. Mol Cell Biochem 340, 143–152 (2010). https://doi.org/10.1007/s11010-010-0411-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11010-010-0411-z

Keywords

Search

Navigation