Evaluating the potential of foodborne chemicals to modulate intestinal barrier function: a combined in vitro/in silico toolbox to study the effects on tight junction protein claudin-4

Intestinal barrier integrity is of paramount importance for our health. Several food constituents and contaminants have been described for positive or detrimental effects, however, a correlation between gut associated disorders and xenobiotics exposures remains challenging and modeling options limited. This study presents a NAMs-based hybrid in vitro/in silico workflow to investigate, and eventually prioritize, foodborne xenobiotics for their effect on gut barrier homeostasis. Model compounds comprised endocrine-active chemicals, such as plastic components (bisphenol A, bisphenol F), phytoestrogens (chrysin, daidzein, genistein), mycotoxins (alternariol, zearalenone), and natural/synthetic steroids (17-beta-estradiol, cholesterol, levonorgestrel). Plasticizer diisodecyl phthalate was included as foodborne, non-endocrine active substance. The Caco-2/HT29-MTX-E12 intestinal epithelial co-culture was used to design a workflow capable of grasping discrete alterations in tight junction organization. The protein claudin-4 was taken as reference to study the structural organization along cell-cell contacts. Complementarily, docking and molecular dynamics simulations were applied to explore ligandclaudin-4 interactions, providing a pipeline which is potentially suitable to screen larger compounds libraries. Finally, proof-of-principle experiments including repeated exposures to reference compound 17-beta-estradiol and plasticizer diisodecyl phthalate, supported the plausibility of a functional impairment of claudin-4. Both compounds significantly increased epithelial permeability without altering cell viability, indicating selective modulation of mechanical barrier properties. Together, this work outlines a mechanistic toolbox for the study of the potential of orally derived xenobiotics in modulating intestinal homeostasis.