Due to their high proline content, gliadins are resistant to complete gastrointestinal digestion and are not degraded into di- or tripeptides that can be transported into enterocytes via the peptide transporter PepT1. Although gliadin uptake by enterocytes has been shown to occur through endocytosis, differences in the uptake of distinct gliadin preparations between celiac disease and healthy control induced pluripotent stem cell-derived small intestinal epithelial cells (iPSC-SIECs) have not previously been investigated.
We examined the uptake and intracellular transport of ATTO-labeled digests of a commercial wheat gliadin, wheat gliadins from pure line of hexaploid bread wheat (BW208) and tetraploid durum wheat (Don Pedro), and their CRISPR-Cas9-edited wheat lines (V653 and V520, respectively) and maize zein in iPSC-SIECs derived from celiac disease patients and healthy controls. Uptake dynamics were monitored using time-lapse confocal microscopy. Raw image sequences were first restored via an automatic pipeline that estimates and removes signal-dependent noise. The restored images were then aligned using a multiscale registration procedure assuming affine temporal deformations. Thereafter the amount and location of endocytosed material were analyzed through image analysis.
All gliadin preparations and zein were readily endocytosed by both celiac and healthy control iPSC-SIECs during the one-hour imaging period. Among the tested proteins, Don Pedro-derived gliadin peptides exhibited a consistently higher rate of endocytosis. This increased uptake was observed in both celiac and healthy control cells.
Under the isolated epithelial conditions used in this study, peptide uptake and intracellular penetration were comparable between celiac and healthy iPSC-SIECs. The consistently elevated endocytic rate of Don Pedro-derived peptides in both groups suggests that specific structural characteristics of this gliadin preparation may influence cellular uptake and warrant further investigation.