T, Health-related Faculty Mannheim, Ruprecht Karls University, Heidelberg Mannheim, Germany Department of Chemistry, University of Cologne, Cologne, Germany d Department of Nephrology, Academic Healthcare Center, Groningen, The Netherlandsb cart ic l e i nf oArticle history: Received 7 May possibly 2014 Received in revised form 29 May possibly 2014 Accepted two June 2014 Readily available on line five June 2014 Key phrases: Endothelial cells Carbon monoxide Adhesion molecules Enzyme-triggered CORMsa b s t r a c tAcyloxydiene e(CO)three complexes can act as enzyme-triggered CO-releasing molecules (ET-CORMs). Their biological activity strongly will depend on the mother compound from which they’re derived, i.e. cyclohexenone or cyclohexanedione, and on the position in the ester functionality they harbour. The present study addresses if the latter characteristic affects CO release, if cytotoxicity of ET-CORMs is mediated by way of iron release or inhibition of cell respiration and to what extent cyclohexenone and cyclohexanedione derived ET-CORMs differ in their ability to counteract TNF- mediated inflammation. Irrespective of the formulation (DMSO or cyclodextrin), toxicity in HUVEC was significantly higher for ET-CORMs bearing the ester functionality at the outer (rac-4), as when compared with the inner (rac-1) position in the cyclohexenone moiety. This was paralleled by an increased CO release from the former ET-CORM. Toxicity was not mediated by means of iron as EC50 values for rac-4 had been drastically decrease than for FeCl2 or FeCl3 and were not influenced by iron chelation. ATP depletion preceded toxicity suggesting impaired cell respiration as putative cause for cell death. In long-term HUVEC cultures inhibition of VCAM-1 expression by rac-1 waned in time, while for the cyclohexanedione derived rac-8 inhibition seems to increase. NFB was inhibited by both rac-1 and rac-8 independent of IB degradation. Both ET-CORMs activated Nrf-2 and consequently induced the expression of HO-1. This study additional gives a rational framework for designing acyloxydiene e(CO)three complexes as ET-CORMs with differential CO release and biological activities. We also supply a much better understanding of how these complexes affect cell-biology in mechanistic terms. 2014 The Authors. Published by Elsevier B.V. This is an open access article beneath the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/3.0/).Introduction Carbon monoxide is endogenously produced in mammalian cells by means of the action of extremely conserved haem oxygenase enzymes [1,2], which catalyse the rate-limiting step in degradation of haem to biliverdin, iron and carbon monoxide (CO) [3?]. TheAbbreviations: CO, carbon monoxide; ET-CORM, enzyme-triggered carbon monoxide-releasing molecule; HUVEC, human umbilical vein endothelial cells; VCAM-1, vascular cell adhesion molecule 1; NF, nuclear factor kappa-lightchain enhancer of activated B-cells; HO-1, haem oxygenase 1; Nrf2, nuclear factor (erythroid-derived); TNF-, tumour necrosis aspect alpha n Correspondence to: Vth Medical Clinic, University Hospital Mannheim, Theodor-Kutzer-Ufer 1-3, Mannheim D-68167, Germany.Spiro[2.5]octane-1-carboxylic acid In stock Tel.1554086-90-6 web : 49-621-383 3771; fax: 49-621-383 3804.PMID:32261617 E-mail address: stamellou.eleni@googlemail (E. Stamellou). 1 SE and SD have contributed equally to this study.CO method has emerged in current years as a crucial essential element in cell physiology and pathophysiology. Based on the cytoprotective properties of this program, the therapeutic possible of CO has been extensively explored in a selection.