Fig. 5. Tissue health was maintained over 72 h in the device and monitored after media treatment.

a Control tissue after 72 h experiment. (i) Tol blue staining showing maintenance of colon morphology, MUC = mucus layer, m = mucosa, cr = crypt, sm = submucosa, me = muscularis externa. (ii) Claudin-1 immunoreactivity in vivo. (iii) Claudin-1 immunoreactivity in 72 h ex vivo shows maintenance of tight junctions between epithelial cells and crypts. (iv) UEA-1⁺ material in vivo. (v) UEA-1⁺ material ex vivo confirming maintenance of epithelial cells and mucus layer. (vi) Quantification of goblet cell number per apical crypt in in vivo vs. ex vivo tissue, n = 3, error bars show the standard error. b Collagenase treated, and acidic luminal media resulted in alterations in goblet cell morphology and tight junction expression indicative of increased barrier permeability. (i) Goblet cells labeled with UEA-1 become circular after collagenase treatment. (ii) Acidic media resulted in loss of goblet cell shape and sloughing off of cells near the lumen. (iii) Alterations in tight junction protein expression (claudin-1) following collagenase treatment. (iv) Claudin-1 expression decreased considerably with exposure to acidic media indicative of substantial barrier disruption. (v) The bar graph shows a distinct reduction in transepithelial electrical resistance (TEER) after exposure to different media composition. The difference in TEER was measured from 24 to 48 h mark after the tissue was enclosed in the device, with the media change occurring at 24 h. The three media compositions consist of a control media, collagenase treated media, and low pH media (more details about media composition in “Animals, Tissue Collection, and Media Preparation”). L = lumen, TEER values are normalized to the membrane surface area of the chamber, 0.0314 cm². Control: n = 4, Collagenase: n = 10, Low pH: n = 3, error bars show standard error, **p < 0.005; ***p < 0.0001.