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


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Tomita
Mikio
M
Department of Drug Absorption and Pharmacokinetics, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan. tomita@ps.toyaku.ac.jp
Takizawa
Yusuke
Y
Kishimoto
Hisanao
H
Hayashi
Masahiro
M
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D000818:Animals; D015870:Gene Expression; D007082:Ileum / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D008297:Male; D026901:Membrane Transport Proteins / Q000187:drug effects / Q000378:metabolism; D052248:Nitric Oxide Synthase Type I / Q000235:genetics / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D015427:Reperfusion Injury / Q000378:metabolism* / Q000503:physiopathology; D020112:Rhodamine 123 / Q000493:pharmacokinetics*; D014700:Verapamil / Q000494:pharmacology
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D026901:Membrane Transport Proteins; D020112:Rhodamine 123; D014700:Verapamil; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II