Identification of a commercial Xylan as a microfold cell-targeting polysaccharide in vivo and in vitro

Commercial Xylan undergoes efficient Microfold cell-mediated transport in vitro and in vivo . • Xylan possesses a β -(1 → 4)-linked backbone, branched with α -(1 → 2)-linked 4- O -methyl-glucuronic acid residues. • Uromodulin is predicted and validated as a potential receptor mediating Xylan transport. Microfold cells (M cells) in the gut serve as a key gateway to enhance the potency of oral vaccines. Although natural polysaccharides show promise as carriers, their development is limited by inherent drawbacks such as scarce availability and lack of intrinsic targeting capacity. Therefore, it is essential to identify suitable materials from well-defined, commercially available polysaccharide standards. In this study, a library of well-defined, high-purity commercial polysaccharides was screened, and identified Xylan as a lead candidate with superior M cell transport efficiency in an in vitro human M-like cell model. This transport was confirmed in vivo , where mass spectrometry quantified a significant delivery (2.45–9.97 μg) of Xylan into murine Peyer's patches. Structural confirmation via monosaccharide composition, enzymatic oligosaccharide profiling, methylation, and NMR analyses identified this active polymer as a β -1, 4-Xylan branched with α -1, 2-glucuronic acid bearing methyl substitution. Mechanistically, molecular docking predicted the M cell surface protein Uromodulin (Umod) as a potential receptor, a hypothesis subsequently validated by isothermal titration calorimetry and surface plasmon resonance. In conclusion, our findings reveal that a specific, commercial Xylan is selectively transported by M cells, a process potentially mediated by Umod, highlighting its great promise as a novel and well-defined carrier for oral vaccine development. Download: Download high-res image (263KB) Download: Download full-size image