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Soluble VE‐cadherin contributes to loss of VE‐cadherin‐mediated adhesion and endothelial barrier function
Author(s) -
Flemming Sven,
Burkard Natalie,
Renschler Melanie,
Vielmuth Franziska,
Meir Michael,
Schick Martin,
Wunder Christian,
Germer ChristopThomas,
Spindler Volker,
Waschke Jens,
Schegel Nicolas
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.923.1
Subject(s) - cadherin , ve cadherin , adam10 , microbiology and biotechnology , barrier function , extracellular , inflammation , adhesion , biology , chemistry , cell adhesion , metalloproteinase , disintegrin , immunology , biochemistry , matrix metalloproteinase , cell , organic chemistry
Endothelial barrier breakdown in systemic inflammation and sepsis is a hallmark preceding organ failure and death in patients. Vascular endothelial (VE)‐cadherin‐mediated adhesion represents a crucial determinant of endothelial barrier integrity. Previously we demonstrated that under inflammatory conditions the disintegrin and metalloproteinase 10 (ADAM10) caused shedding of VE‐cadherin which resulted in the formation of soluble VE‐cadherin (sVE‐cadherin) in close temporal correlation of inflammation‐induced endothelial barrier disruption. Here we further characterized ADAM10‐induced VE‐cadherin shedding and tested whether recombinant sVE‐cadherin inferes VE‐cadherin mediated adhesion. First we identified possible cleavages sites of VE‐cadherin by mass spectrometry (MSM). Therefore, digestion of recombinant VE‐cadherin (rVE‐cadherin) with recombinant ADAM10 was performed. This led to the formation of sVE‐cadherin at 90 and 110kDa, as shown by coomassie stained gel an western blot, which correlated with several cleavage sites of the extracellular domain of VE‐cadherin. MSM showed only peptides up to 452 amino acids (aa) and one peptide at 503–530 aa, which also corresponds to the extracellular domains 1–5 of human VE‐cadherin. Additionally two other cleavage sites at 80–127aa and 160–175aa were observed. In the next step we performed atomic force microscopy measurements in a cell free sytem to test whether sVe‐cadherin blocked VE‐cadherin‐mediated adhesion. In these experiments both, sVE‐cadherin and rVE‐cadherin blocked VE‐cadherin‐mediated adhesion. Accordingly, application of rVE‐cadherin on Human dermal microvascular endothelial cells (HDMECs) resulted in a dose‐dependent loss of endothelial barrier functions as revealed by measurements of transendothelial electrical resistance (TER). In immunostaining of endothelial monolayers incubation with rVE‐cadherin led to intercellular gap formation. In summary our data demonstrate specific cleavage sites of VE‐cadherin by ADAM10. The resulting sVE‐cadherin fragments interfere VE‐cadherin mediated adhesion and thereby contribute to loss of endothelial barrier disruption. This may play a novel role in inflammation‐induced endothelial barrier breakdown. Support or Funding Information The present study was supported by grants from the German Research Foundation (DFG) to N.S. (Grant numbers: SCHL1962/2‐1 and SCHL 1962/4‐1).