UTILIZATION OF A HUMAN INTESTINAL EPITHELIAL-CELL CULTURE SYSTEM (CACO-2) FOR EVALUATING CYTOPROTECTIVE AGENTS

Human intestinal epithelial cells (Caco-2) were cultured as confluent monolayers on polycarbonate membranes in Transwells for investigating their applicability in evaluating the cytoprotective activity of sucra lfate. The control experiments established a reproducible chemical met hod (using 0.5 mM indomethacin in Hanks' balanced salt solution) for i nducing damage to the Caco-2 cell monolayers. Damage was determined by measuring changes in transepithelial electrical resistance (TEER). Tw enty-day-old Caco-2 cell monolayers were significantly and reproducibl y damaged (compared to buffer alone) (P < 0.001) by application of 0.5 mM indomethacin to the apical side for 1 hr. While sucralfate, at a 0 .5, 2, or 5 mg/mL concentration in the buffer, was shown not to revers e (treat) the damage caused by indomethacin in this cellular model, it was able to protect (prevent) the cells from indomethacin-induced dam age (P < 0.001). We observed that indomethacin-induced damage to the C aco-2 cell monolayers greatly affected the paracellular pathway since the percentage transport of [H-3]methoxyinulin was significantly eleva ted. In contrast, protection of the Caco-2 cells with 5 mg/mL sucralfa te in the presence of the damaging agent resulted in transport of the paracellular marker similar to that in the control (HBSS-treated) cell monolayers. This direct cytoprotective effect was thus independent of vascular factors at neutral pH and was observed to be dose dependent (0.5 to 5 mg/mL) when sucralfate was applied to the cells in the prese nce of the damaging agent. These findings, which are consistent with t hose observed for sucralfate in vivo (Okabe et al., Digest. Dis. Sci. 28:1034-1042, 1983), demonstrate the feasibility of using Caco-2 cell monolayers as an in vitro cell culture system which may serve to ident ify and rapidly screen the cytoprotective activity of potential drugs and their pharmaceutical formulations.