Effect of intestinal ischaemia/reperfusion on P-glycoprotein-mediated ileal excretion of rhodamine 123 in the rat

Abstract

We have shown that ischaemia/reperfusion in the small intestine at an early phase, such as 1 h after reperfusion, induced not only functional changes in the membrane, such as P-glycoprotein (P-gp) dysfunction, but also decreased expression of P-gp protein and mdr1a mRNA. In the present study we examined whether intestinal ischaemia/reperfusion modifies the P-gp-mediated ileal excretion transport system in rats beyond 1 h after reperfusion. To evaluate the contribution of P-gp-mediated transport to the ileal excretion of rhodamine 123, we used Western blotting to measure the expression of P-gp protein levels isolated from the ileum at different reperfusion times after 60 min of ischaemia. We also measured the expression of inducible nitric oxide synthase (iNOS) mRNA using real-time RT-PCR. Ileal excretion of rhodamine 123 decreased at 3 h after reperfusion and had recovered at 24 h. Changes in villi structure at 3 h and its recovery at 24 h were also observed. Verapamil, a competitive inhibitor of P-gp, significantly inhibited ileal clearance of rhodamine 123 to the lumen at 24 h after reperfusion, suggesting that P-gp was working at this time. These results suggest that intestinal ischaemia/reperfusion-induced decrease in P-gp-mediated ileal excretion of rhodamine 123 was probably due to impaired P-gp-mediated transport. Levels of P-gp protein and iNOS mRNA in the ileum decreased 3 h after ischaemia/reperfusion and returned to control levels after 24 h. These findings suggest that intestinal ischaemia/reperfusion markedly decreases P-gp-mediated ileal excretion of rhodamine 123, probably by decreasing the expression of P-gp protein, which is likely to be due to increased lipid peroxidation via iNOS.