Transcellular relocation of tetrahydrobiopterin across Caco-2 cells: A model study of tetrahydrobiopterin absorption through epithelial cells of intestinal mucosa

Oral administration of tetrahydrobiopterin (BH₄) has been known to be effective in treating BH₄-deficient patients. It has long been established that BH₄ is absorbed by the intestinal mucosa. However, the mechanism for translocation of BH₄ across epithelial cells has not been elucidated. In order to study BH₄ transport mechanisms, Caco-2 cells were employed in this study as an epithelial cell model. Caco-2 cells were cultured (2 × 10⁴ cells/0.3 cm² well) for 21 days in a 24-well format using Transwell, a porous membrane-based culture dish, at which point they had established themselves as a tight sheet with a definite polarity. When BH₄ (100 μmol/L) was given to cells from the apical side, a considerable translocation toward their basolateral side was noted. The rate of BH₄ movement was around 150 pmol/h per well. This was comparable to the highest rate of BH₄ uptake or its release so far obtained using a monolayer culture of Caco-2 cells on an ordinary plastic plate. The transcellular movement of BH₄ across the polar culture on the porous membrane was effectively prevented by benzbromarone (10 μmol/L), a well known inhibitor of a group of transporters including urate transporter (URAT1), organic anion transporters (OATs), and multidrug-resistance-associated proteins (MRPs). It was thus concluded that in Caco-2 cells, BH₄ moved across the cell interior in a rapid ligand-specific manner that was driven by a transporter.