Figure 5.

(a) Tacrolimus metabolite formation by intestinal enterocytes. Enterocyte cultures were exposed to 50 μmol l⁻¹ dexamethasone for 48 h. Controls consisted of dishes not exposed to dexamethasone. Thereafter the enterocyte cultures were exposed to 3, 7 μmol l⁻¹ tacrolimus for 2 h. Metabolites were identified by LC/particle beam-MS: demethyl-tacrolimus (M1), bisdemethyl-tacrolimus (M2), demethyl-hydroxy-tacrolimus (M3) and hydroxy-tacrolimus (M4). Data is shown as mean±s.d. of four dishes (2×10⁶ cells). (b) Western blot analysis of enterocytes (10 μg microsomes) at 2 days in culture (i.e. 48 h induction period). Following isolation and cell seeding enterocytes were cultured in the presence of 50 μmol l⁻¹ dexamethasone for 48 h. Thereafter cells were harvested and frozen at −20°C until further preparation. Microsomes were produced and subjected to Western blot analysis as detailed in the Methods section. Cells not exposed to dexamethasone were used as controls (Co).