New Endothelial Cell-Based Assay to Assess Variability of Nitric Oxide Production in Humans

PROJECT SUMMARY ABSTRACT Cardiovascular diseasesCVDare the leading cause of death in the US and other developed nationsInalonemore thanmillion Americans suffered from CVD with associated cost $billionThe development of CVD and other chronic circulatory disorders has been associated with exposure to drugs and environmental toxicantsThese chemicals can alter the bioavailability of a key regulator of vascular functionnitric oxideNOwhich is produced by endothelial cellsECsThe impairment of vascular EC NO production can contribute to the development of chronic circulatory disordersHoweverthis does not explain why some individuals develop the disease and others do notAccordinglythe National Institute of Environmental Health Sciences leads in efforts to understand the contribution of individual susceptibility to complex diseasessuch as CVDTo aid these effortsCreative ScientistIncCSIis developing advanced tools enabling the generation of data on individual vascular susceptibility to drugs and environmental toxicantsUsing proprietary protocols and cell growth media supplementsCSI isolates Endothelial Colony Forming CellsECFCfrom small volumes of peripheral blood and cryopreserves the cellsECFCs are unique primary cells involved in both embryonic and adult vasculogenesisThe donor specific nature and robust growth potential of ECFCs allow for the testing of tens of thousands of chemicals in cells derived from any number of individualsOur goal is to commercialize a NO testing service using a novel and highly specific NO reagent and a high content imaging platformIn Aimwe will determine the sensitivity and selectivity of our novel NO specific fluorescent probeCompletion of this Aim will demonstrate the utility of our reagent for measuring basal and induced NO levels in ECFCsIn Aimwe will determine the reproducibility of measuring NO levels in different ECFC lineages using different culture conditionse gwith inducerwith inducer and inhibitorCompletion of Aimwill establish the utility of ECFCs for identification of NO inhibitorsIn Phase IIwe will test the assayandapos s ability to detect human response variability totoxicants using a collection of ECFCs derived from up todonorsOur ultimate goal is to provide a commercial service to regulatory and research agencies and pharmaceutical companies aiming to evaluate the potential vascular toxicity of environmental toxicants and drug candidates in a population andin particulartrying to identify both sensitive and resistant individuals and or population sub groupsThe success of our assay development efforts will help subsequently commercialize a stand alone NO testing kit PROJECT NARRATIVE A new method to test nitric oxideNOproduction in donor specific vascular endothelial cells is proposed because current methods for testing the impairment of NO production in endothelial cells lack sensitivity and specificityNO is produced by endothelial cells and is a key regulator of vascular functionMany drugs and environmental toxicants alter the bioavailability of endothelial NO leading to cardiovascular and other circulatory diseaseswhich are the leading cause of death in the US and other developed countries