The MTX-induced apoptosis of IEC-6 cells was shown to be depressed by AGE. AGE may be useful for the cancer chemotherapy with MTX, since AGE reduces the MTX-induced intestinal damage.

Gastrointestinal toxicity is one of the most serious side effects of methotrexate (MTX) treatment. The side effects often disrupt the cancer chemotherapy. We previously reported that aged garlic extract (AGE) protects the small intestine of rats from MTX-induced damage. In this study, the protection of AGE against MTX-induced damage of IEC-6 cells originating from the rat jejunum crypt was investigated. MTX decreased the viability of IEC-6 cells, but this effect was prevented by AGE (0.5%). The MTX-induced apoptosis of IEC-6 cells was depressed by AGE. These results indicated that AGE protects IEC-6 cells from the MTX-induced damage. AGE may be useful in cancer chemotherapy with MTX because it reduces MTX-induced intestinal damage.

The transport characteristics of fluorescein methotrexate (F-MTX) were studied by using the rat intestinal crypt cell line IEC-6. Enhanced accumulation of F-MTX at 4 degrees C suggests the existence of an active efflux system. MK-571, an inhibitor of the multidrug resistance-associated protein/ATP binding cassette C (MRP/ABCC) family, also enhanced the accumulation of F-MTX. Transcellular transport of F-MTX from the apical to the basolateral compartment was 2.5 times higher than the opposite direction. This vectorial transport was also reduced by MK-571, indicating the presence of Mrp-type transporter(s) on the basolateral membrane. Mrp3 mRNA was readily detectable, and the protein was localized on the basolateral membrane. Uptake of FMTX into membrane vesicles from IEC-6 cells and Spodoptera frugiperda-9 cells expressing rat Mrp3 were both ATP dependent and saturable as a function of the F-MTX concentration. Similar Km values (11.0 +/- 1.8 and 4.5 +/- 1.1 microM) and inhibition profiles by MK-571, estradiol-17beta-d-glucuronide, and taurocholate for the ATP-dependent transport of F-MTX into these vesicles were obtained. These findings suggest that the efflux of F-MTX is mediated by Mrp3 on the basolateral membrane of IEC-6 cells.

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