Electrical Impedance of Cultured Endothelium Under Fluid Flow NATACHA DEPAOLA, 1 JEFFREY E. PHELPS, 1 LUCIO FLOREZ, 1 CHARLES R. KEESE, 2 FRED L. MINNEAR, 4 IVAR GIAEVER, 3 and PETER VINCENT 4 1 Departments of Biomedical Engineering, 2 Biology, and 3 Physics, Rensselaer Polytechnic Institute, Troy, NY and 4 Center for Cardiovascular Sciences, Albany Medical College, Albany, NY (Received 4 June 1999; accepted 11 May 2001) Abstract—The morphological and functional status of organs, tissues, and cells can be assessed by evaluating their electrical impedance. Fluid shear stress regulates the morphology and function of endothelial cells in vitro. In this study, an electrical biosensor was used to investigate the dynamics of flow-induced alterations in endothelial cell morphology in vitro. Quantitative, real-time changes in the electrical impedance of endothelial monolayers were evaluated using a modified electric cell- substrate impedance sensing ECIS system. This ECIS/Flow system allows for a continuous evaluation of the cell mono- layer impedance upon exposure to physiological fluid shear stress forces. Bovine aortic endothelial cells grown to conflu- ence on thin film gold electrodes were exposed to fluid shear stress of 10 dynes/cm 2 for a single uninterrupted 5 h time period or for two consecutive 30 min time periods separated by a 2 h no-flow interval. At the onset of flow, the monolayer electrical resistance sharply increased reaching 1.2 to 1.3 times the baseline in about 15 min followed by a sustained decrease in resistance to 1.1 and 0.85 times the baseline value after 30 min and 5 h of flow, respectively. The capacitance decreased at the onset of flow, started to recover after 15 min and after slightly overshooting the baseline values, decreased again with a prolonged exposure to flow. Measured changes in capacitance were in the order of 5% of the baseline values. The observed changes in endothelial impedance were reversible upon flow removal with a recovery rate that varied with the duration of the preceding flow exposure. These results demonstrate that the impedance of endothelial monolayers changes dynamically with flow indicating morphological and/or functional changes in the cell layer. This in vitro model system ECIS/Flow may be a very useful tool in the quantitative evaluation of flow- induced dynamic changes in cultured cells when used in con- junction with biological or biochemical assays able to deter- mine the nature and mechanisms of the observed changes. © 2001 Biomedical Engineering Society. DOI: 10.1114/1.1385811 Keywords—Endothelial cells, Electrical impedance, Shear stress, Flow. INTRODUCTION The electrical properties of organs and tissues provide valuable information on their functional and structural status. Measurements of electrical resistance have been traditionally used to evaluate barrier function in epithe- lial tissues forming the interfaces between biological compartments. The endothelium is the functional barrier between the circulating blood and the arterial wall. It controls trans- port across the vascular wall, transduces blood-borne sig- nals, and is involved in the regulation of vascular tone, vascular growth, and hemostasis and thrombosis, among others. Several devices and chambers with various elec- trode configurations have been used to measure the elec- trical resistance of cultured endothelial layers grown on microporous membranes. 35 Most recently, Giaever and Keese 18–20 developed a biosensor that allows real-time measurements of electrical impedance in cell cultures grown on small gold electrodes. This method, referred to as electric cell-substrate impedance sensing ECIS, has been successfully used to monitor cell behavior in tissue culture including cell attachment and spreading, micro- motion, permeability, changes in cell shape, pore forma- tion and resealing in the plasma membrane of mamma- lian cells, and alteration in cellular function under various drug and biochemical treatments. 2,14,17–20,34 Hemodynamic forces modulate endothelial cell mor- phology and function in vivo and in vitro. The time frame of endothelial response to fluid forces varies from seconds to hours. 3,6 The endothelium constitutes a com- plex interface that plays a key role in mechanotransduc- tion and regulation of signaling events between the mov- ing blood and the vessel wall. Endothelial responses to shear stress range from electrophysiological changes in membrane potential, transcription factor activation, gene regulation, synthesis and protein expression, and signifi- cant changes of cell structure, among others. Numerous review articles have been published in which the effect of shear stress in endothelial cell structure and function, signaling events, and flow-mediated endothelial mechan- otransduction and its mechanisms are discussed. 3,4,6 Real time evaluation of cell morphology and function is of increasing value to determine the dynamics of the endo- thelial response to flow. 3,5,15,25,32,33 In their vast majority, Address all correspondence to Natacha DePaola, PhD, Department of Biomedical Engineering, Rensselaer Polytechnic Institute, 110 8th Street, Troy, NY 12180. Electronic mail: depaola@rpi.edu Annals of Biomedical Engineering, Vol. 29, pp. 648–656, 2001 0090-6964/2001/298/648/9/$15.00 Printed in the USA. All rights reserved. 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