Effects Of JBP485 On The Expression And Function Of PEPT1 In Indomethacin-induced Intestinal Injury In Rats And Damage In Caco-2 Cells

Objective: A major limitation of long term use of NSAIDs such as indomethacin and aspirin is the damage caused to the small intestine. PEPT1 is the important transporter in the absorption and excretion of drugs, primarily expressed in the brush-border membrane of the small intestine. JBP485 (cyclo-trans- 4-L-hydroxyprolyl-L-serine) is a dipeptide with obvious inflammation protective effect after oral administration. To investigate the effect of JBP485 on indomethacin-induced intestinal injury in rats and damage in Caco-2 cells, the activity and expression of PEPT1 were also examined.Methods: Male Wistar rats were administered JBP485 at a dose of 25 mg/kg, 0.5 h before and 2, 4.5 or 7 h after treatment with indomethacin (10mg/kg). 24 h after indomethacin treatment, diarrhea and ulcer were measured with and without JBP485. To illuminate the protective effect of JBP485, we determined histologic damage in intestine slices, maleic dialdehyde (MDA) and myeloperoxidase (MPO) in intestine homogenate after administration of JBP485 in indomethacin-induced intestine injury in rat. The effects of treatment with indomethacin and co-treatment with JBP485 were examined in terms of morphology, LDH-release and oxidative stress in Caco-2 cells. 24 h after indomethacin treatment, uptake of glycylsarcosine (Gly-Sar) by PEPT1 was determined by in vivo, in vitro and in situ. RT-PCR and Western blot were used to assess the expression of PEPT1 in rat intestine and Caco-2 cells.Results: Compared to the untreated control group, a single dose of indomethacin at10 mg/kg mostly provoked diarrhea and ulcer lesions in the small intestine, loss of surface epithelium, necrosis of the upper villi and transmural necrosis associated with massive mucosal infiltration by lymphocytes cells in intestine slices, while these effects were significantly reduced by co-treatment with JBP485. JBP485 relieves the indomethacin-induced mucosal inflammation and oxidative tissue damage in rats, caused a significant decrease in MDA and MPO levels. JBP485 promotes the repair of indomethacin-induced damage in Caco-2 cells, attenuating morphology of Caco-2 cells and reversing the activity of LDH and levels of MDA. Uptake of Gly-Sar by PEPT1 was decreased by indomethacin treatment, whereas the Gly-Sar plasma concentration was markedly increased in JBP485 co-treated rats in vivo, in vitro and in situ studies. Indomethacin down-regulated the expression of PEPT1 mRNA and protein in rat intestine and Caco-2 cells, the effects were reversed after administration of JBP485. No differences were found in experimental results in the rats and Caco-2 cells treated with JBP485 alone compared with control group.Conclusion: JBP485 was able to protect against indomethacin-induced intestinal injury in rats and cellular damage in a human intestinal cell line by decreasing oxidative stress. JBP485 partially blocked the down-regulation of PEPT1 expression and function in rat small intestine and Caco-2 cells induced by indomethacin.