Skip to main content

Advertisement

SpringerLink
Log in

Urinary trypsin inhibitor attenuates LPS-induced endothelial barrier dysfunction by upregulation of vascular endothelial-cadherin expression

  • Original Research Paper
  • Published:
Inflammation Research Aims and scope Submit manuscript

Abstract

Introduction

Urinary trypsin inhibitor (UTI) decreases inflammatory cytokine levels and mortality in experimental animal models of inflammation. Here, we observed the effect of UTI on lipopolysaccharide (LPS)-induced hyperpermeability in human umbilical vein endothelial cells (HUVECs) and explored the role of vascular endothelial-cadherin (VE-cadherin) in its effect.

Methods

The effect of UTI on endothelial barrier hyperpermeability was detected by an electrical cell–substrate impedance sensing (ECIS) system and a transwell chamber system. The expression of VE-cadherin in HUVECs was examined by real-time PCR and western blot.

Results

We demonstrated that the alleviation of LPS-induced barrier dysfunction could be achieved by pretreatment with 3000 U/mL of UTI. VE-cadherin monoclonal antibody (mAb) could inhibit the protective effects. UTI maintained VE-cadherin expression by increasing protein stability at both the transcriptional and post-transcriptional levels. Meanwhile, VE-cadherin expression on the cell surface increased when the cells were pretreated with UTI. Furthermore, pretreatment with UTI decreased the phosphorylation of VE-cadherin at Tyr658 but not Tyr731.

Conclusions

Our data show that prophylactic UTI maintains the endothelial barrier function, increases VE-cadherin expression, and inhibits the phosphorylation of VE-cadherin at Tyr658 under inflammatory conditions. It suggests a scientific and potential clinical therapeutic importance of UTI in treatment of inflammatory disorders.

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
Fig. 8

Similar content being viewed by others

References

  1. Fox ED, Heffernan DS, Cioffi WG, Reichner JS. Neutrophils from critically ill septic patients mediate profound loss of endothelial barrier integrity. Crit Care. 2013;17:R226.

    Article  PubMed Central  PubMed  Google Scholar 

  2. Lee WL, Slutsky AS. Sepsis and endothelial permeability. N Engl J Med. 2010;363:689–91.

    Article  CAS  PubMed  Google Scholar 

  3. Bogatcheva NV, Zemskova MA, Kovalenkov Y, Poirier C, Verin AD. Molecular mechanisms mediating protective effect of cAMP on lipopolysaccharide (LPS)-induced human lung microvascular endothelial cells (HLMVEC) hyperpermeability. J Cell Physiol. 2009;221:750–9.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  4. Xing J, Wang Q, Coughlan K, Viollet B, Moriasi C, Zou MH. Inhibition of AMP-activated protein kinase accentuates lipopolysaccharide-induced lung endothelial barrier dysfunction and lung injury in vivo. Am J Pathol. 2013;182:1021–30.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  5. Aghajanian A, Wittchen ES, Allingham MJ, Garrett TA, Burridge K. Endothelial cell junctions and the regulation of vascular permeability and leukocyte transmigration. J Thromb Haemost JTH. 2008;6:1453–60.

    Article  CAS  PubMed  Google Scholar 

  6. Gustot T. Multiple organ failure in sepsis: prognosis and role of systemic inflammatory response. Curr Opin Crit Care. 2011;17:153–9.

    Article  PubMed  Google Scholar 

  7. Dejana E. Endothelial cell-cell junctions: happy together. Nat Rev Mol Cell Biol. 2004;5:261–70.

    Article  CAS  PubMed  Google Scholar 

  8. Herwig MC, Tsokos M, Hermanns MI, Kirkpatrick CJ, Muller AM. Vascular endothelial cadherin expression in lung specimens of patients with sepsis-induced acute respiratory distress syndrome and endothelial cell cultures. Pathobiol J Immunopathol Mol Cell Biol. 2013;80:245–51.

    Article  CAS  Google Scholar 

  9. May C, Doody JF, Abdullah R, Balderes P, Xu X, Chen CP, et al. Identification of a transiently exposed VE-cadherin epitope that allows for specific targeting of an antibody to the tumor neovasculature. Blood. 2005;105:4337–44.

    Article  CAS  PubMed  Google Scholar 

  10. Taddei A, Giampietro C, Conti A, Orsenigo F, Breviario F, Pirazzoli V, et al. Endothelial adherens junctions control tight junctions by VE-cadherin-mediated upregulation of claudin-5. Nat Cell Biol. 2008;10:923–34.

    Article  CAS  PubMed  Google Scholar 

  11. Bannerman DD, Sathyamoorthy M, Goldblum SE. Bacterial lipopolysaccharide disrupts endothelial monolayer integrity and survival signaling events through caspase cleavage of adherens junction proteins. J Biol Chem. 1998;273:35371–80.

    Article  CAS  PubMed  Google Scholar 

  12. Angelini DJ, Hyun SW, Grigoryev DN, Garg P, Gong P, Singh IS, et al. TNF-alpha increases tyrosine phosphorylation of vascular endothelial cadherin and opens the paracellular pathway through fyn activation in human lung endothelia. Am J Physiol Lung Cell Mol Physiol. 2006;291:L1232–45.

    Article  CAS  PubMed  Google Scholar 

  13. Corada M, Liao F, Lindgren M, Lampugnani MG, Breviario F, Frank R, et al. Monoclonal antibodies directed to different regions of vascular endothelial cadherin extracellular domain affect adhesion and clustering of the protein and modulate endothelial permeability. Blood. 2001;97:1679–84.

    Article  CAS  PubMed  Google Scholar 

  14. Gao X, Kouklis P, Xu N, Minshall RD, Sandoval R, Vogel SM, et al. Reversibility of increased microvessel permeability in response to VE-cadherin disassembly. Am J Physiol Lung Cell Mol Physiol. 2000;279:L1218–25.

    CAS  PubMed  Google Scholar 

  15. Gavard J. Endothelial permeability and VE-cadherin: a wacky comradeship. Cell Adhes Migr. 2013;7:455–61.

    Article  Google Scholar 

  16. Sato H, Kajikawa S, Kuroda S, Horisawa Y, Nakamura N, Kaga N, et al. Impaired fertility in female mice lacking urinary trypsin inhibitor. Biochem Biophys Res Commun. 2001;281:1154–60.

    Article  CAS  PubMed  Google Scholar 

  17. Hirose J, Ozawa T, Miura T, Isaji M, Nagao Y, Yamashiro K, et al. Human neutrophil elastase degrades inter-alpha-trypsin inhibitor to liberate urinary trypsin inhibitor related proteins. Biol Pharm Bull. 1998;21:651–6.

    Article  CAS  PubMed  Google Scholar 

  18. Yano T, Anraku S, Nakayama R, Ushijima K. Neuroprotective effect of urinary trypsin inhibitor against focal cerebral ischemia-reperfusion injury in rats. Anesthesiology. 2003;98:465–73.

    Article  CAS  PubMed  Google Scholar 

  19. Inoue K, Takano H, Shimada A, Yanagisawa R, Sakurai M, Yoshino S, et al. Urinary trypsin inhibitor protects against systemic inflammation induced by lipopolysaccharide. Mol Pharmacol. 2005;67:673–80.

    Article  CAS  PubMed  Google Scholar 

  20. Bae HB, Jeong CW, Li M, Kim HS, Kwak SH. Effects of urinary trypsin inhibitor on lipopolysaccharide-induced acute lung injury in rabbits. Inflammation. 2012;35:176–82.

    Article  CAS  PubMed  Google Scholar 

  21. Molor-Erdene P, Okajima K, Isobe H, Uchiba M, Harada N, Okabe H. Urinary trypsin inhibitor reduces LPS-induced hypotension by suppressing tumor necrosis factor-alpha production through inhibition of Egr-1 expression. Am J Physiol Heart Circ Physiol. 2005;288:H1265–71.

    Article  CAS  PubMed  Google Scholar 

  22. Song Z, Chen G, Lin G, Jia C, Cao J, Ao G. The ultra-early protective effect of ulinastatin on rabbit acute lung injury induced by paraquat. BMC Emerg Med. 2013;13(Suppl 1):S7.

    PubMed Central  PubMed  Google Scholar 

  23. Takubo T. Human urinary trypsin inhibitor. Nihon rinsho Jpn J Clin Med. 2010;68(Suppl 7):810–3.

    Google Scholar 

  24. Linder A, Russell JA. An exciting candidate therapy for sepsis: ulinastatin, a urinary protease inhibitor. Intensive Care Med. 2014;40:1164–7.

    Article  PubMed  Google Scholar 

  25. Chen H, He MY, Li YM. Treatment of patients with severe sepsis using ulinastatin and thymosin alpha1: a prospective, randomized, controlled pilot study. Chin Med J. 2009;122:883–8.

    CAS  PubMed  Google Scholar 

  26. Moon SW, Lee SW, Hong YS, Park DW, Jang IJ, Yoon YH, Lim SI. The effects of urinary trypsin inhibitor on the outcomes of severe sepsis and septic shock patients. J Korean Soc Emerg Med. 2009;20:80–5.

    Google Scholar 

  27. Karnad DR, Bhadade R, Verma PK, Moulick ND, Daga MK, Chafekar ND, et al. Intravenous administration of ulinastatin (human urinary trypsin inhibitor) in severe sepsis: a multicenter randomized controlled study. Intensive Care Med. 2014;40:830–8.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  28. Winiarska M, Bil J, Wilczek E, Wilczynski GM, Lekka M, Engelberts PJ, et al. Statins impair antitumor effects of rituximab by inducing conformational changes of CD20. PLoS Med. 2008;5:e64.

    Article  PubMed Central  PubMed  Google Scholar 

  29. Cowan CE, Kohler EE, Dugan TA, Mirza MK, Malik AB, Wary KK. Kruppel-like factor-4 transcriptionally regulates VE-cadherin expression and endothelial barrier function. Circ Res. 2010;107:959–66.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  30. Wang L, Taneja R, Wang W, Yao LJ, Veldhuizen RA, Gill SE, et al. Human alveolar epithelial cells attenuate pulmonary microvascular endothelial cell permeability under septic conditions. PLoS One. 2013;8:e55311.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  31. Redmond EM, Cahill PA, Hirsch M, Wang YN, Sitzmann JV, Okada SS. Effect of pulse pressure on vascular smooth muscle cell migration: the role of urokinase and matrix metalloproteinase. Thromb Haemost. 1999;81:293–300.

    CAS  PubMed  Google Scholar 

  32. Prota LF, Cebotaru L, Cheng J, Wright J, Vij N, Morales MM, et al. Dexamethasone regulates CFTR expression in Calu-3 cells with the involvement of chaperones HSP70 and HSP90. PLoS One. 2012;7:e47405.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  33. Payvandi F, Wu L, Haley M, Schafer PH, Zhang LH, Chen RS, et al. Immunomodulatory drugs inhibit expression of cyclooxygenase-2 from TNF-alpha, IL-1beta, and LPS-stimulated human PBMC in a partially IL-10-dependent manner. Cell Immunol. 2004;230:81–8.

    Article  CAS  PubMed  Google Scholar 

  34. Lee SH, Jee JG, Bae JS, Liu KH, Lee YM. A group of novel HIF-1alpha inhibitors, glyceollins, blocks HIF-1alpha synthesis and decreases its stability via inhibition of the PI3 K/AKT/mTOR pathway and Hsp90 binding. J Cell Physiol. 2015;230:853–62.

    Article  CAS  PubMed  Google Scholar 

  35. Ye X, Ding J, Zhou X, Chen G, Liu SF. Divergent roles of endothelial NF-kappaB in multiple organ injury and bacterial clearance in mouse models of sepsis. J Exp Med. 2008;205:1303–15.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  36. Groger M, Pasteiner W, Ignatyev G, Matt U, Knapp S, Atrasheuskaya A, Bukin E, Friedl P, Zinkl D, Hofer-Warbinek R, Zacharowski K, Petzelbauer P, Reingruber S. Peptide Bbeta(15-42) preserves endothelial barrier function in shock. PLoS One. 2009;4:e5391.

    Article  PubMed Central  PubMed  Google Scholar 

  37. Ivanov AI, Parkos CA, Nusrat A. Cytoskeletal regulation of epithelial barrier function during inflammation. Am J Pathol. 2010;177:512–24.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  38. Wolfson RK, Chiang ET, Garcia JG. HMGB1 induces human lung endothelial cell cytoskeletal rearrangement and barrier disruption. Microvasc Res. 2011;81:189–97.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  39. Mehta D, Malik AB. Signaling mechanisms regulating endothelial permeability. Physiol Rev. 2006;86:279–367.

    Article  CAS  PubMed  Google Scholar 

  40. Li R, Ren M, Chen N, Luo M, Zhang Z, Wu J. Vitronectin increases vascular permeability by promoting VE-cadherin internalization at cell junctions. PLoS One. 2012;7:e37195.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  41. Potter MD, Barbero S, Cheresh DA. Tyrosine phosphorylation of VE-cadherin prevents binding of p120- and beta-catenin and maintains the cellular mesenchymal state. J Biol Chem. 2005;280:31906–12.

    Article  CAS  PubMed  Google Scholar 

  42. Donners MM, Wolfs IM, Olieslagers S, Mohammadi-Motahhari Z, Tchaikovski V, Heeneman S, van Buul JD, Caolo V, Molin DG, Post MJ, Waltenberger J. A disintegrin and metalloprotease 10 is a novel mediator of vascular endothelial growth factor-induced endothelial cell function in angiogenesis and is associated with atherosclerosis. Arterioscler Thromb Vasc Biol. 2010;30:2188–95.

    Article  CAS  PubMed  Google Scholar 

  43. Shen W, Li S, Chung SH, Zhu L, Stayt J, Su T, Couraud PO, Romero IA, Weksler B, Gillies MC. Tyrosine phosphorylation of VE-cadherin and claudin-5 is associated with TGF-beta1-induced permeability of centrally derived vascular endothelium. Eur J Cell Biol. 2011;90:323–32.

    Article  CAS  PubMed  Google Scholar 

  44. Wessel F, Winderlich M, Holm M, Frye M, Rivera-Galdos R, Vockel M, Linnepe R, Ipe U, Stadtmann A, Zarbock A, Nottebaum AF, Vestweber D. Leukocyte extravasation and vascular permeability are each controlled in vivo by different tyrosine residues of VE-cadherin. Nat Immunol. 2014;15:223–30.

    Article  CAS  PubMed  Google Scholar 

  45. Schildberger A, Rossmanith E, Weber V, Falkenhagen D. Monitoring of endothelial cell activation in experimental sepsis with a two-step cell culture model. Innate Immun. 2010;16:278–87.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We thank Prof. Hang Xiao for his invaluable technical assistance throughout the study period. And this work was supported by the Priority Academic Program Development of Jiangsu Higher Education Institutions and the Natural Science Foundation of Jiangsu Province (BK20131444).

Author information

Author notes

    Authors and Affiliations

    1. Department of Emergency, The First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing, 210029, People’s Republic of China

      Jie Chen, Hao Sun, Junjie Chen, Huazhong Zhang & Jinsong Zhang

    2. The Laboratory of Neurotoxicology, School of Public Health, Nanjing Medical University, Nanjing, 210029, People’s Republic of China

      Jun Wang & Wenyi Qian

    3. Department of Emergency, The Affiliated Hospital of Xuzhou Medical College, Xuzhou, 221000, People’s Republic of China

      Chenglei Su

    4. Department of Basic Medical College, Nanjing Medical University, Nanjing, 210029, People’s Republic of China

      Li Sun

    Authors
    1. Jie Chen
      View author publications

      You can also search for this author in PubMed Google Scholar

    2. Jun Wang
      View author publications

      You can also search for this author in PubMed Google Scholar

    3. Chenglei Su
      View author publications

      You can also search for this author in PubMed Google Scholar

    4. Wenyi Qian
      View author publications

      You can also search for this author in PubMed Google Scholar

    5. Li Sun
      View author publications

      You can also search for this author in PubMed Google Scholar

    6. Hao Sun
      View author publications

      You can also search for this author in PubMed Google Scholar

    7. Junjie Chen
      View author publications

      You can also search for this author in PubMed Google Scholar

    8. Huazhong Zhang
      View author publications

      You can also search for this author in PubMed Google Scholar

    9. Jinsong Zhang
      View author publications

      You can also search for this author in PubMed Google Scholar

    Corresponding author

    Correspondence to Jinsong Zhang.

    Ethics declarations

    Conflict of interest

    The authors declare that they have no conflict of interest.

    Additional information

    Responsible Editor: Artur Bauhofer.

    J. Chen and J. Wang contributed equally to this work.

    Rights and permissions

    Reprints and permissions

    About this article

    Check for updates. Verify currency and authenticity via CrossMark

    Cite this article

    Chen, J., Wang, J., Su, C. et al. Urinary trypsin inhibitor attenuates LPS-induced endothelial barrier dysfunction by upregulation of vascular endothelial-cadherin expression. Inflamm. Res. 65, 213–224 (2016). https://doi.org/10.1007/s00011-015-0907-9

    Download citation

    • Received:

    • Revised:

    • Accepted:

    • Published:

    • Issue Date:

    • DOI: https://doi.org/10.1007/s00011-015-0907-9

    Keywords

    Search

    Navigation