Skip to main content

Advertisement

SpringerLink
Log in

Dual Regulation of L-Selectin-Mediated Leukocyte Adhesion by Endothelial Surface Glycocalyx

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

Abstract

Endothelial surface glycocalyx (ESG) is a carbohydrate-rich, gel-like layer found on vascular endothelium, serving critical functions in mechanotransduction of blood flows, maintenance of the endothelial permeability, and the control of leukocyte adhesion and inflammation. This study aimed to clarify the role of ESG in the adhesion between leukocytes and Human Umbilical Vein Endothelial Cells (HUVECs) under resting or inflammatory conditions. Using an atomic force microscopy-based single-cell adhesion assay, we directly quantified the detachment force and work perpendicular to the cell membrane. Detachment force and work were measured for every separation event of a leukocyte from a HUVEC with ESG, or with the major ESG glycosaminoglycan components, heparan sulfate (HS) and hyaluronic acid (HA) removed. For the resting HUVECs, when HS and/or HA were removed, the detachment force and work increased dramatically. For the HUVECs activated by inflammatory cytokine tumor necrosis factor alpha, we observed increases in the detachment force and work compared to the resting HUVECs, and removal of HS and/or HA resulted in significant decreases in the detachment force and work. The results demonstrate that the ESG layer serves a dual function: (1) on resting endothelium, it prevents leukocyte adhesion, and (2) under inflammatory conditions, it participates in endothelial-leukocyte interactions with molecules such as selectins.

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

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  1. Atkins, E. D., and J. K. Sheehan. Structure for hyaluronic acid. Nature 235:253–254, 1972.

    Article  Google Scholar 

  2. Bell, G. I., M. Dembo, and P. Bongrand. Cell adhesion. Competition between nonspecific repulsion and specific bonding. Biophys. J. 45:1051–1064, 1984.

    Article  Google Scholar 

  3. Bet, M. R., G. Goissis, S. Vargas, and H. S. Selistre-de-Araujo. Cell adhesion and cytotoxicity studies over polyanionic collagen surfaces with variable negative charge and wettability. Biomaterials 24:131–137, 2003.

    Article  Google Scholar 

  4. Birdsall, H. H., C. Lane, M. N. Ramser, and D. C. Anderson. Induction of VCAM-1 and ICAM-1 on human neural cells and mechanisms of mononuclear leukocyte adherence. J. Immunol. 148:2717–2723, 1992.

    Google Scholar 

  5. Buscher, K., S. B. Riese, M. Shakibaei, C. Reich, J. Dernedde, R. Tauber, and K. Ley. The transmembrane domains of L-selectin and CD44 regulate receptor cell surface positioning and leukocyte adhesion under flow. J. Biol. Chem. 285:13490–13497, 2010.

    Article  Google Scholar 

  6. Chen, X., Y. Wang, J. Liu, P. Xu, X. M. Zhang, Y. Y. Tian, Y. M. Xue, X. Y. Gao, Y. Liu, and J. H. Wang. Synergistic effect of HMGB1 knockdown and cordycepin in the K562 human chronic myeloid leukemia cell line. Mol. Med. Rep. 12:4462–4468, 2015.

    Google Scholar 

  7. Constantinescu, A. A., H. Vink, and J. A. Spaan. Endothelial cell glycocalyx modulates immobilization of leukocytes at the endothelial surface. Arterioscler. Thromb. Vasc. Biol. 23:1541–1547, 2003.

    Article  Google Scholar 

  8. Couffinhal, T., C. Duplaa, L. Labat, J. M. Lamaziere, C. Moreau, O. Printseva, and J. Bonnet. Tumor necrosis factor-alpha stimulates ICAM-1 expression in human vascular smooth muscle cells. Arterioscler. Thromb. 13:407–414, 1993.

    Article  Google Scholar 

  9. Erlandsen, S. L., S. R. Hasslen, and R. D. Nelson. Detection and spatial distribution of the beta 2 integrin (Mac-1) and L-selectin (LECAM-1) adherence receptors on human neutrophils by high-resolution field emission SEM. J. Histochem. Cytochem. 41:327–333, 1993.

    Article  Google Scholar 

  10. Fransson, L. A., M. Belting, F. Cheng, M. Jonsson, K. Mani, and S. Sandgren. Novel aspects of glypican glycobiology. Cell. Mol. Life Sci. 61:1016–1024, 2004.

    Article  Google Scholar 

  11. Fu, B. M. M., and J. M. Tarbell. Mechano-sensing and transduction by endothelial surface glycocalyx: composition, structure, and function. Wiley Interdiscipl. Rev.-Syst. Biol. Med. 5:381–390, 2013.

    Article  Google Scholar 

  12. Giuffre, L., A. S. Cordey, N. Monai, Y. Tardy, M. Schapira, and O. Spertini. Monocyte adhesion to activated aortic endothelium: role of L-selectin and heparan sulfate proteoglycans. J. Cell Biol. 136:945–956, 1997.

    Article  Google Scholar 

  13. Hanson, A. M., J. Gambill, V. Phomakay, C. T. Staten, and M. D. Kelley. 9-cis-retinoic Acid and troglitazone impacts cellular adhesion, proliferation, and integrin expression in K562 cells. PLoS One 9:e93005, 2014.

    Article  Google Scholar 

  14. Hutter, J. L., and J. Bechhoefer. Calibration of Atomic-Force Microscope Tips (Vol 64, pp 1868, 1993). Rev. Sci. Instrum. 64:3342, 1993.

    Article  Google Scholar 

  15. Ihrcke, N. S., L. E. Wrenshall, B. J. Lindman, and J. L. Platt. Role of heparan sulfate in immune system-blood vessel interactions. Immunol. Today 14:500–505, 1993.

    Article  Google Scholar 

  16. Jackson, R. L., S. J. Busch, and A. D. Cardin. Glycosaminoglycans: molecular properties, protein interactions, and role in physiological processes. Physiol. Rev. 71:481–539, 1991.

    Google Scholar 

  17. Jung, T. M., W. M. Gallatin, I. L. Weissman, and M. O. Dailey. Down-regulation of homing receptors after T cell activation. J. Immunol. 141:4110–4117, 1988.

    Google Scholar 

  18. Kishimoto, T. K., M. A. Jutila, and E. C. Butcher. Identification of a human peripheral lymph node homing receptor: a rapidly down-regulated adhesion molecule. Proc. Natl. Acad. Sci. USA 87:2244–2248, 1990.

    Article  Google Scholar 

  19. Koenig, A., K. Norgard-Sumnicht, R. Linhardt, and A. Varki. Differential interactions of heparin and heparan sulfate glycosaminoglycans with the selectins—implications for the use of unfractionated and low molecular weight heparins as therapeutic agents. J. Clin. Invest. 101:877–889, 1998.

    Article  Google Scholar 

  20. Latkovic, S. Ultrastructural localization of lectin-binding sites on the surface of the guinea pig conjunctival epithelium. Graefe’s Arch. Clin. Exp. Ophthalmol 229:153–156, 1991.

    Article  Google Scholar 

  21. Laurent, V. M., A. Duperray, V. Sundar Rajan, and C. Verdier. Atomic force microscopy reveals a role for endothelial cell ICAM-1 expression in bladder cancer cell adherence. PLoS One 9:e98034, 2014.

    Article  Google Scholar 

  22. Lever, R., M. J. Rose, E. A. McKenzie, and C. P. Page. Heparanase induces inflammatory cell recruitment in vivo by promoting adhesion to vascular endothelium. Am. J. Physiol. Cell Physiol. 306:C1184–C1190, 2014.

    Article  Google Scholar 

  23. Ley, K., T. F. Tedder, and G. S. Kansas. L-Selectin can mediate leukocyte rolling in untreated mesenteric venules in vivo independent of E-selection or P-selectin. Blood 82:1632–1638, 1993.

    Google Scholar 

  24. Lozzio, B. B., C. B. Lozzio, E. G. Bamberger, and A. S. Feliu. A multipotential leukemia cell line (K-562) of human origin. Proc. Soc. Exp. Biol. Med. Soc. Exp. Biol. Med. 166:546–550, 1981.

    Article  Google Scholar 

  25. Ma, X. L., A. S. Weyrich, D. J. Lefer, M. Buerke, K. H. Albertine, T. K. Kishimoto, and A. M. Lefer. Monoclonal antibody to L-selectin attenuates neutrophil accumulation and protects ischemic reperfused cat myocardium. Circulation 88:649–658, 1993.

    Article  Google Scholar 

  26. Miyakawa, Y., T. Nishimura, Y. Ueyama, K. Miyake, M. Miyasaka, Y. Ikeda, and S. Habu. Cell adhesion via murine alpha4 human beta1 integrin chimera on transfected K562 cells to endothelial cells. Exp. Cell Res. 226:75–79, 1996.

    Article  Google Scholar 

  27. Moore, K. L., K. D. Patel, R. E. Bruehl, F. Li, D. A. Johnson, H. S. Lichenstein, R. D. Cummings, D. F. Bainton, and R. P. McEver. P-selectin glycoprotein ligand-1 mediates rolling of human neutrophils on P-selectin. J. Cell Biol. 128:661–671, 1995.

    Article  Google Scholar 

  28. Mulivor, A. W., and H. H. Lipowsky. Inflammation- and ischemia-induced shedding of venular glycocalyx. Am. J. Physiol. Heart Circ. Physiol. 286:H1672–H1680, 2004.

    Article  Google Scholar 

  29. Nandi, A., P. Estess, and M. H. Siegelman. Hyaluronan anchoring and regulation on the surface of vascular endothelial cells is mediated through the functionally active form of CD44. J. Biol. Chem. 275:14939–14948, 2000.

    Article  Google Scholar 

  30. Nelson, R. M., O. Cecconi, W. G. Roberts, A. Aruffo, R. J. Linhardt, and M. P. Bevilacqua. Heparin oligosaccharides bind L- and P-selectin and inhibit acute inflammation. Blood 82:3253–3258, 1993.

    Google Scholar 

  31. Norgardsumnicht, K., and A. Varki. Endothelial heparan-sulfate proteoglycans that bind to L-selectin have glucosamine residues with unsubstituted amino-groups. J. Biol. Chem. 270:12012–12024, 1995.

    Article  Google Scholar 

  32. Oberleithner, H., W. Peters, K. Kusche-Vihrog, S. Korte, H. Schillers, K. Kliche, and K. Oberleithner. Salt overload damages the glycocalyx sodium barrier of vascular endothelium. Pflugers Arch. 462:519–528, 2011.

    Article  Google Scholar 

  33. Ohlson, M., J. Sorensson, and B. Haraldsson. A gel-membrane model of glomerular charge and size selectivity in series. Am. J. Physiol. Renal Physiol. 280:F396–F405, 2001.

    Google Scholar 

  34. Pai, R., B. Bassa, M. A. Kirschenbaum, and V. S. Kamanna. TNF-alpha stimulates monocyte adhesion to glomerular mesangial cells. The role of intercellular adhesion molecule-1 gene expression and protein kinases. J. Immunol. 156:2571–2579, 1996.

    Google Scholar 

  35. Pries, A. R., T. W. Secomb, and P. Gaehtgens. The endothelial surface layer. Pflugers Arch. 440:653–666, 2000.

    Article  Google Scholar 

  36. Reitsma, S., D. W. Slaaf, H. Vink, M. A. van Zandvoort, and M. G. oude Egbrink. The endothelial glycocalyx: composition, functions, and visualization. Pflugers Arch. 454:345–359, 2007.

    Article  Google Scholar 

  37. Rops, A. L., J. van der Vlag, J. F. Lensen, T. J. Wijnhoven, L. P. van den Heuvel, T. H. van Kuppevelt, and J. H. Berden. Heparan sulfate proteoglycans in glomerular inflammation. Kidney Int. 65:768–785, 2004.

    Article  Google Scholar 

  38. Sabri, S., M. Soler, C. Foa, A. Pierres, A. M. Benoliel, and P. Bongrand. Glycocalyx modulation is a physiological means of regulating cell adhesion. J. Cell Sci. 113:1589–1600, 2000.

    Google Scholar 

  39. Sarrazin, S., W. C. Lamanna, and J. D. Esko. Heparan sulfate proteoglycans. Cold Spring Harb. Perspect. Biol. 3:583–590, 2011.

    Article  Google Scholar 

  40. Shriver, Z., I. Capila, G. Venkataraman, and R. Sasisekharan. Heparin and heparan sulfate: analyzing structure and microheterogeneity. Handb. Exp. Pharmacol. 207:159–176, 2012.

    Article  Google Scholar 

  41. Snapp, K. R., A. J. Wagers, R. Craig, L. M. Stoolman, and G. S. Kansas. P-selectin glycoprotein ligand-1 is essential for adhesion to P-selectin but not E-selectin in stably transfected hematopoietic cell lines. Blood 89:896–901, 1997.

    Google Scholar 

  42. Soler, M., S. Desplat-Jego, B. Vacher, L. Ponsonnet, M. Fraterno, P. Bongrand, J. M. Martin, and C. Foa. Adhesion-related glycocalyx study: quantitative approach with imaging-spectrum in the energy filtering transmission electron microscope (EFTEM). FEBS Lett. 429:89–94, 1998.

    Article  Google Scholar 

  43. Sorensson, J., M. Ohlson, and B. Haraldsson. A quantitative analysis of the glomerular charge barrier in the rat. Am. J. Physiol. Renal Physiol. 280:F646–F656, 2001.

    Google Scholar 

  44. Springer, T. A. Traffic signals for lymphocyte recirculation and leukocyte emigration: the multistep paradigm. Cell 76:301–314, 1994.

    Article  Google Scholar 

  45. Stein, J. V., G. Cheng, B. M. Stockton, B. P. Fors, E. C. Butcher, and U. H. von Andrian. L-selectin-mediated leukocyte adhesion in vivo: microvillous distribution determines tethering efficiency, but not rolling velocity. J. Exp. Med. 189:37–50, 1999.

    Article  Google Scholar 

  46. van den Berg, B. M., H. Vink, and J. A. E. Spaan. The endothelial glycocalyx protects against myocardial edema. Circ. Res. 92:592–594, 2003.

    Article  Google Scholar 

  47. Vink, H., and B. R. Duling. Identification of distinct luminal domains for macromolecules, erythrocytes, and leukocytes within mammalian capillaries. Circ. Res. 79:581–589, 1996.

    Article  Google Scholar 

  48. Walcheck, B., J. Kahn, J. M. Fisher, B. B. Wang, R. S. Fisk, D. G. Payan, C. Feehan, R. Betageri, K. Darlak, A. F. Spatola, and T. K. Kishimoto. Neutrophil rolling altered by inhibition of L-selectin shedding in vitro. Nature 380:720–723, 1996.

    Article  Google Scholar 

  49. Weinbaum, S., J. M. Tarbell, and E. R. Damiano. The structure and function of the endothelial glycocalyx layer. Annu. Rev. Biomed. Eng. 9:121–167, 2007.

    Article  Google Scholar 

  50. Zakrzewicz, A., M. Grafe, D. Terbeek, M. Bongrazio, W. Auch-Schwelk, B. Walzog, K. Graf, E. Fleck, K. Ley, and P. Gaehtgens. L-selectin-dependent leukocyte adhesion to microvascular but not to macrovascular endothelial cells of the human coronary system. Blood 89:3228–3235, 1997.

    Google Scholar 

  51. Zeng, Y., E. E. Ebong, B. M. Fu, and J. M. Tarbell. The structural stability of the endothelial glycocalyx after enzymatic removal of glycosaminoglycans. PLoS One 7:e43168, 2012.

    Article  Google Scholar 

  52. Zhang, X., A. Chen, D. De Leon, H. Li, E. Noiri, V. T. Moy, and M. S. Goligorsky. Atomic force microscopy measurement of leukocyte-endothelial interaction. Am. J. Physiol. Heart Circ. Physiol. 286:H359–H367, 2004.

    Article  Google Scholar 

  53. Zhao, J. H., S. Nagao, and Z. L. Zhang. Loading and unloading of a spherical contact: from elastic to elastic-perfectly plastic materials. Int. J. Mech. Sci. 56:70–76, 2012.

    Article  Google Scholar 

Download references

Acknowledgments

This work was supported in part by Lehigh University scholarship to M.A.D, Lehigh University Grants for Experiential Learning in Health to K.G., and an American Heart Association grant (11SDG5420008), and start-up funding from Lehigh University to X.F.Z. The authors also wish to thank Dr. Bingmei Fu and Dr. John Tarbell of CUNY for their insightful discussions of the experimental design.

Conflict of interest

Authors Matthew Dragovich, Kaylynn Genemaras, Hannah L. Dailey and Sabrina Jedlicka have no conflicts of interest to disclose. Author X. Frank Zhang has received funding from the American Heart Association (11SDG5420008) and Lehigh University (start-up funding) to conduct the research.

Ethical Statement

Professors Zhang, Dailey, Jedlicka and Matthew Dragovich have no personal, professional or financial conflicts of interest in this project and/or proposal. We have conducted no research on human subjects, live or dead vertebrate animal subjects and our laboratories have accommodations for safe practice: Gloves, hoods, autoclaves, sterile equipment, proper refrigeration, etc.…. All data acquisition for the applied force and work was gathered and analyzed in Igor; the figures seen in this paper of the AFM force-extension curves are renditions of the raw data gathered from Igor and are displayed in Origin. The Box and whiskers plots of the analyzed data are displayed in Excel. The equipment is owned by Professor Zhang and was purchased with his funding from Lehigh University. The equipment used in this project is managed by Mr. Dragovich with further oversight from Professor Zhang. Mr. Dragovich was responsible for gathering and analyzing the cell–cell adhesion data. The equipment in this project was also under use by the then undergraduate researcher Kaylynn Genemaras who was managed by Mr. Dragovich and assisted Mr. Dragovich in gathering and analyzing the cell–cell adhesion data. The AFM was previously used in collaboration with several other groups as well. Professor Dailey provided the statistical analysis for this project which is displayed in the figures and the appendix. Professor Jedlicka aided in providing the antibody staining images. The assertion that any publications from this project make will be handled appropriately so author integrity will be maintained. The data is presented with the expectation that it will be made public to accept challenges and/or corroborations. Since these findings may influence health care and be used by other scientific researchers they will particularly be handled with the upmost ethical responsibility. Intellectual property gained during this study from all participating in this project will be handled through the appropriate channels to be sure that appropriate credits and rights are obtained.

Informed consent

No human studies were carried out by the authors for this article.

Author information

Author notes

  1. Kaylynn Genemaras

    Present address: Bioinnovation Program, Tulane University, 6823 Saint Charles Avenue, New Orleans, LA, 70118, USA

Authors and Affiliations

  1. Department of Mechanical Engineering & Mechanics, Lehigh University, 19 Memorial Drive West, Bethlehem, PA, 18015, USA

    Matthew A. Dragovich, Hannah L. Dailey & X. Frank Zhang

  2. Bioengineering Program, Lehigh University, 19 Memorial Drive West, Bethlehem, PA, 18015, USA

    Kaylynn Genemaras, Sabrina Jedlicka & X. Frank Zhang

  3. Department of Materials Science and Engineering, Lehigh University, 19 Memorial Drive West, Bethlehem, PA, 18015, USA

    Sabrina Jedlicka

Authors
  1. Matthew A. Dragovich
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Kaylynn Genemaras
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hannah L. Dailey
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Sabrina Jedlicka
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. X. Frank Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Matthew A. Dragovich or X. Frank Zhang.

Additional information

Associate Editor Michael R. King oversaw the review of this article.

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 96 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Dragovich, M.A., Genemaras, K., Dailey, H.L. et al. Dual Regulation of L-Selectin-Mediated Leukocyte Adhesion by Endothelial Surface Glycocalyx. Cel. Mol. Bioeng. 10, 102–113 (2017). https://doi.org/10.1007/s12195-016-0463-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12195-016-0463-6

Keywords

Search

Navigation