title: Activin receptor-like kinase 5 inhibition reverses impairment of endothelial cell viability by endogenous islet mesenchymal stromal cells.
abstract: Following islet transplantation, islet graft revascularization is compromised due to loss of endothelial cells (ECs) during islet culture. TGF-β signaling pathways are essential for vascular homeostasis but their importance for islet EC function is unclear. We have identified a population of multipotent mesenchymal stromal cells (MSCs) within islets and investigated how modulation of TGF-β signaling by these cells influences islet EC viability. Cultured islets exhibited reduced expression of EC markers (VEGFR2, VE-cadherin and CD31), which was associated with diminished but sustained expression of endoglin a marker of both ECs and MSCs. Double fluorescent labeling of islets in situ with the EC marker CD31 disclosed a population of CD31-negative cells which were positive for endoglin. In vitro coculture of microvascular ECs with endoglin-positive, CD31-negative islet MSCs reduced VEGFR2 protein expression, disrupted EC angiogenic behavior, and increased EC detachment. Medium conditioned by islet MSCs significantly decreased EC viability and increased EC caspase 3/7 activity. EC:MSC cocultures showed enhanced Smad2 phosphorylation consistent with altered ALK5 signaling. Pharmacological inhibition of ALK5 activity with SB431542 (SB) improved EC survival upon contact with MSCs, and SB-treated cultured islets retained EC marker expression and sensitivity to exogenous VEGF164 . Thus, endoglin-expressing islet MSCs influence EC ALK5 signaling in vitro, which decreases EC viability, and changes in ALK5 activity in whole cultured islets contribute to islet EC loss. Modifying TGF-β signaling may enable maintenance of islet ECs during islet isolation and thus improve islet graft revascularization post-transplantation.
authors.affiliation: Diabetes Research Group, Division of Diabetes and Nutritional Sciences, School of Medicine, Kings College London, London, UK. C.E.Clarkin@soton.ac.uk
authors.forename: Claire E
authors.initials: CE
authors.lastname: Clarkin
authors.forename: Aileen J
authors.initials: AJ
authors.lastname: King
authors.forename: Paramjeet
authors.initials: P
authors.lastname: Dhadda
authors.forename: Pedro
authors.initials: P
authors.lastname: Chagastelles
authors.forename: Nance
authors.initials: N
authors.lastname: Nardi
authors.forename: Caroline P
authors.initials: CP
authors.lastname: Wheeler-Jones
authors.forename: Peter M
authors.initials: PM
authors.lastname: Jones
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis; D001549:Benzamides / Q000494:pharmacology; D002470:Cell Survival / Q000187:drug effects; D018920:Coculture Techniques; D004149:Dioxoles / Q000494:pharmacology; D000071063:Endoglin; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000096:biosynthesis; D007515:Islets of Langerhans / Q000098:blood supply / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D016381:Islets of Langerhans Transplantation / Q000379:methods*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D051379:Mice; D008813:Mice, Inbred ICR; D018919:Neovascularization, Physiologic / Q000187:drug effects; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000077293:Receptor, Transforming Growth Factor-beta Type I; D011956:Receptors, Cell Surface / Q000096:biosynthesis; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015398:Signal Transduction; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology
chemical_list: C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; D015703:Antigens, CD; D001549:Benzamides; D004149:Dioxoles; C064206:ENG protein, human; D000071063:Endoglin; C502004:Eng protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D047428:Protein Kinase Inhibitors; D011956:Receptors, Cell Surface; D018125:Receptors, Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; C000625841:TGFBR1 protein, human; C000625842:Tgfbr1 protein, mouse