title: Thrombospondin-1 opens the paracellular pathway in pulmonary microvascular endothelia through EGFR/ErbB2 activation.
abstract: Thrombospondin-1 (TSP1) is a multidomain protein that contains epidermal growth factor (EGF)-like repeats that indirectly activate the EGF receptor (EGFR) and selected downstream signaling pathways. In these studies, we show that TSP1 opens the paracellular pathway in human lung microvascular endothelial cells (HMVEC-Ls) in a dose-, time-, and protein tyrosine kinase (PTK)-dependent manner. TSP1 increased tyrosine phosphorylation of proteins enriched to intercellular boundaries including the zonula adherens (ZA) proteins, vascular endothelial-cadherin, γ-catenin, and p120 catenin. In HMVEC-Ls, EGFR and ErbB2 are expressed at low levels, and both heterodimerize and tyrosine autophosphorylate in response to TSP1. Prior EGFR-selective PTK inhibition with AG1478 or ErbB2-selective PTK inhibition with AG825 protected against TSP1-induced tyrosine phosphorylation of ZA proteins and barrier disruption. Preincubation of HMVEC-Ls with an EGFR ectodomain-blocking antibody also prevented TSP1-induced opening of the paracellular pathway. Therefore, in HMVEC-Ls, TSP1 increases tyrosine phosphorylation of ZA proteins and opens the paracellular pathway, in part, through EGFR/ErbB2 activation. Surprisingly, recombinant TSP1 EGF-like repeats 1-3 and the high-affinity EGFR ligands, EGF, TGF-α, and amphiregulin, each failed to increase paracellular permeability. However, HMVEC-Ls in which EGFR was overexpressed became responsive to the EGF-like repeats of TSP1 as well as to EGF. These studies indicate that TSP1 disrupts the endothelial barrier through EGFR/ErbB2 activation although additional signals are necessary in cells with low receptor expression.
authors.affiliation: Mucosal Biology Research Center, and Departments of Medicine and Pathology, University of Maryland, School of Medicine, Baltimore, Maryland 21201, USA.
authors.forename: Pallavi
authors.initials: P
authors.lastname: Garg
authors.forename: Shiqi
authors.initials: S
authors.lastname: Yang
authors.forename: Anguo
authors.initials: A
authors.lastname: Liu
authors.forename: Manuel A
authors.initials: MA
authors.lastname: Pallero
authors.forename: Donald J
authors.initials: DJ
authors.lastname: Buchsbaum
authors.forename: Deane F
authors.initials: DF
authors.lastname: Mosher
authors.forename: Joanne E
authors.initials: JE
authors.lastname: Murphy-Ullrich
authors.forename: Simeon E
authors.initials: SE
authors.lastname: Goldblum
mesh_terms: D022005:Adherens Junctions / Q000187:drug effects / Q000378:metabolism; D002250:Carbon Radioisotopes; D042783:Endothelial Cells / Q000187:drug effects / Q000201:enzymology*; D004789:Enzyme Activation / Q000187:drug effects; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D008024:Ligands; D008168:Lung / Q000098:blood supply*; D055806:Microvessels / Q000166:cytology*; D010766:Phosphorylation / Q000187:drug effects; D019000:Phosphotyrosine / Q000378:metabolism; D055503:Protein Multimerization / Q000187:drug effects; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D020449:Repetitive Sequences, Amino Acid; D012710:Serum Albumin, Bovine / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013379:Substrate Specificity / Q000187:drug effects; D019700:Thrombospondin 1 / Q000737:chemistry / Q000494:pharmacology*; D013997:Time Factors
chemical_list: D002250:Carbon Radioisotopes; D066246:ErbB Receptors; D008024:Ligands; D019000:Phosphotyrosine; D018719:Erb-b2 Receptor Tyrosine Kinases; D012710:Serum Albumin, Bovine; D019700:Thrombospondin 1; C512478:EGFR protein, human; C508053:ERBB2 protein, human