title: Comparison of LDH, (51)Cr, and BCECF efflux as indices of non-steroidal anti-inflammatory drug-induced toxicity in human gastro-intestinal (HGT-1, HCT-8 and T84) cell lines: BCECF efflux is not an index of plasma membrane integrity.
abstract: Toxicity induced by non-steroidal anti-inflammatory drugs (NSAIDs) was assessed in vitro in three human gastro-intestinal epithelial cell lines (gastric HGT-1, ileo-caecal HCT-8 and colonic T84) using three proposed indices of cell plasma membrane integrity. Lactate dehydrogenase (LDH) and (51)CR efflux from cells over 5 hr under control conditions was small (about 3 and 9%, respectively). In contrast, the efflux of 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein (BCECF) was much greater, ranging from 45% in HGT-1 cells to 76% in HCT-8 and T84 cells over 5 hr. Indomethacin increased LDH and (51)Cr efflux at concentrations >/= 5 mm. The rank order for the toxicity of four NSAIDs, assessed by (51)Cr efflux in HCT-8 cells, was indomethacin > sulindac > ketoprofen = aspirin. In contrast, the effects of indomethacin on BCECF efflux were bimodal. At concentrations ranging from 10 mum to 5 mm, depending on the cell line, indomethacin reduced BCECF efflux to a minimum of approximately 20% of the control efflux rate. With higher concentrations of indomethacin, BCECF efflux increased to levels similar to, but not exceeding, control rates. Sulindac (0.01-10 mm) and ketoprofen (1-10 mm), but not aspirin (0.01-10 mm) also reduced BCECF efflux. The reduction in BCECF efflux observed with low concentrations of NSAIDs, and the very high rate of BCECF efflux under control conditions, negates the use of this tracer as an index of plasma membrane integrity. However, this inhibition could itself be a sensitive index of toxicity. The mechanism of inhibition of BCECF efflux is suggested to be due to inhibition of transport-mediated flux, and this is a possible site for NSAID action.
authors: Gastrointestinal Drug Delivery Research Centre, University of Newcastle upon Tyne, Medical School, Newcastle upon Tyne NE2 4HH, UK; Department of Physiological Sciences, University of Newcastle upon Tyne, Medical School, Newcastle upon Tyne NE2 4HH, UK.
authors: C N
authors: CN
authors: Allen
authors: T J
authors: TJ
authors: Gray
authors: E S
authors: ES
authors: Harpur
authors: B H
authors: BH
authors: Hirst