%0 Figure %A Jannaway, Melanie %A Yang, Xiaoyuan %A Meegan, Jamie E. %A Coleman, Danielle C. %A Y. Yuan, Sarah %D 2019 %T Thrombin generates syndecan ectodomain fragments. %U https://plos.figshare.com/articles/figure/Thrombin_generates_syndecan_ectodomain_fragments_/8132972 %R 10.1371/journal.pone.0214737.g006 %2 https://plos.figshare.com/ndownloader/files/15160259 %K thrombin receptor PAR 1. %K barrier integrity %K disease states %K barrier function %K syndecan -3 ectodomain fragments %K Syndecan proteins %K plasma protein leakage %K glycocalyx %K Rho kinase inhibitor %K ELISA %K VE-cadherin-based adherens junction disorganization %K albumin-bound Evans %K transmission electron microscopy %K syndecan subtypes %K F-actin stress fibers %K mouse lung %K intercellular gaps %K transmigrating cells %K barrier dysfunction Objective %K paracellular hyperpermeability %K thrombotic disease states %K cleavage products %K paracellular permeability %K blot analysis %K lung microvasculature %X

To aid our understanding of the decrease in S3ED/S4ED proteins, western blots were carried out with recombinant syndecan ectodomains treated with thrombin. (A) Neither rhS1ED or rhS2ED were found to be cleaved by thrombin (10 U/ml, 24 hours). (B) In contrast, rhS3ED and rhS4ED were both cleaved by thrombin (10 U/ml, 24 hours) to produce 4 and 1 fragments respectively (labelled a-e). (C) A shorter treatment time with thrombin of 1 hour was sufficient to cleave and generate the same rhS3ED (a-d) and rhS4ED (e) fragments. (D) Probing of the SDC fragments with anti-6-his tag showed three of the four rhS3ED fragments all contain the C-terminal 6-his tag, whilst the rhS4ED fragment generate through thrombin treatment did not contain the 6-his tag. (E) A schematic to illustrate the probable identities of some of the different SDC ectodomain fragments generated by thrombin based on evidence from western blots. Western blots shown are representative images of n = 3. Arrowhead indicates position of thrombin on blot.

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