Cytokine-induced barrier dysfunction via gut-on-a-chip

A gut-on-chip model that generates human intestinal tubules has been used to demonstrate the detectable effect of cytokine exposure, an effect of inflammation in vivo, that causes complex changes in the intestinal epithelial barrier.  

This cytokine-induced barrier dysfunction was measured by TEER (trans-epithelial electrical resistance), combined compromised cell health (DRAQ7) and cytoskeleton changes (actin) and by lipid signaling profiles (LC-MS/MS).

The work was led by Amy Harms and Thomas Hankemeier at the Leiden Academic Centre for Drug Research, NL.

For the combined analysis of cell health and cytoskeleton alterations, following negative control or cytokine exposure of the gut-on-a-chip intestinal tubules, DRAQ7 was used to detect cell plasma membrane failure (i.e. catastrophic or programmed cell death) during a 30' incubation.  Thereafter, tubules were fixed (3.7% PFA) and then permeabilised with Triton X-100 (0.03%) and then stained with NucBlue Fixed Cell ReadyProbes Reagent (Invitrogen, R37606) (for ALL nuclei) and ActinGreen (Invitrogen, R37110) for cytoskeleton.

Using the stated protocol the data show a clear cytokine dose-related increase in cell death.

This method is additionally important in general use since this describes the use of DRAQ7 to stain nuclei of dead/dying cells and to detect this following fixation and permeabilization steps, opening up a new route for DRAQ7's use, perhaps reliant upon careful titration of fixative and surfactant to moderate the required effect to avoid the undesirable transfer of DRAQ7 from the true dead cells to otherwise DRAQ7-negative fixed and permeabilized cells.

The wider results underpin the physiological response of the in vitro tubules in the OrganoPlate organ-on-a-chip system (Mimetas, b.v.) to the inflammatory insult of cytokine exposure.

Reference:

Morelli, M., Savova, M. V., Queiroz, K., Harms, A. C., & Hankemeier, T. (2025). Cytokine‐Induced Barrier Dysfunction and Lipid Signaling in a Gut‐On‐Chip Model. The FASEB Journal, 39(19), e71059