Arjun Jain1, Shali Chen2, Hannah Yong1, Subrata Chakrabarti2. 1. 1Department of Physiology, Development and Neuroscience, University of Cambridge, Cambridge, UK. 2. 2Department of Pathology, University of Western Ontario, London, ON Canada.
Abstract
BACKGROUND: Diabetes mellitus is a group of metabolic disorders in which there are high blood glucose levels over a prolonged period. Diabetes is one of many diseases associated with pathologically elevated levels of endothelin (ET)-1. We have recently proposed the development of ET-traps, which are an antibody - based fusion protein that potently bind and sequester pathologically elevated levels of endothelin-1. METHODS: We constructed ET-traps that were found to be very potent binders to ET-1, with a KD of 32.5ρM. We then treated human retinal microvascular endothelial cells (HRMECs), which are an in vitro model of glucose induced cellular damage, with 10 nM ET-1 or high glucose levels (25 mM). RESULTS: In this study, we investigated the effects of our ET-trap constructs on the expression levels of both collagen 4α1 and fibronectin, which are both important pathologic markers in diabetes. Treating HRMECs with 10 nM ET-1 or 25 mM glucose significantly induces the expression of the ECM proteins fibronectin and collagen 4α1, as is found in chronic diabetic complications; Incubation of the cells with the ET-traps significantly prevented the increased expression of fibronectin and collagen 4α1 back to basal levels. This was found with both mRNA and protein expression levels of the two ECM proteins. CONCLUSION: Our results provide the first evidence of the efficacy of ET-traps in reducing pathologic markers in an in vitro model (of diabetes). Further research is warranted to determine the efficacy of ET-traps as a therapeutic tool for diabetes, which is a major public health burden around the world.
BACKGROUND: Diabetes mellitus is a group of metabolic disorders in which there are high blood glucose levels over a prolonged period. Diabetes is one of many diseases associated with pathologically elevated levels of endothelin (ET)-1. We have recently proposed the development of ET-traps, which are an antibody - based fusion protein that potently bind and sequester pathologically elevated levels of endothelin-1. METHODS: We constructed ET-traps that were found to be very potent binders to ET-1, with a KD of 32.5ρM. We then treated human retinal microvascular endothelial cells (HRMECs), which are an in vitro model of glucose induced cellular damage, with 10 nM ET-1 or high glucose levels (25 mM). RESULTS: In this study, we investigated the effects of our ET-trap constructs on the expression levels of both collagen 4α1 and fibronectin, which are both important pathologic markers in diabetes. Treating HRMECs with 10 nM ET-1 or 25 mM glucose significantly induces the expression of the ECM proteins fibronectin and collagen 4α1, as is found in chronic diabetic complications; Incubation of the cells with the ET-traps significantly prevented the increased expression of fibronectin and collagen 4α1 back to basal levels. This was found with both mRNA and protein expression levels of the two ECM proteins. CONCLUSION: Our results provide the first evidence of the efficacy of ET-traps in reducing pathologic markers in an in vitro model (of diabetes). Further research is warranted to determine the efficacy of ET-traps as a therapeutic tool for diabetes, which is a major public health burden around the world.
Authors: John Deanfield; Ann Donald; Claudio Ferri; Cristina Giannattasio; Julian Halcox; Sean Halligan; Amir Lerman; Giuseppe Mancia; James J Oliver; Achille C Pessina; Damiano Rizzoni; Gian Paolo Rossi; Antonio Salvetti; Ernesto L Schiffrin; Stefano Taddei; David J Webb Journal: J Hypertens Date: 2005-01 Impact factor: 4.844