FIGURE 9.

Pharmacological inhibition of glucocerebrosidase protects colonoids from BFT and ibiglustat-induced bursting. Colonoids treated with concentrated bacterial culture supernatant from ETBFΔbft (control) (A) or control + 5 μmol/L ibiglustat (B) showed no obvious morphology changes over 48 h as visualized by confocal microscopy. Colonoids treated with BFT undergo dramatic swelling and bubbling and are still in the process of recovering and returning to normal morphology by 48 h (C). Colonoids treated with BFT and 5 μmol/L ibiglustat also display a normal BFT response and delayed recovery, but no colonoid bursting (D). Colonoids in all videos were tracked and bursting events were counted. Colonoids that appeared to initiate, but not complete, bursting were scored as a potential colonoid explosion. Potential bursting events from normal colonoids and colonoids cultured in CBE were counted and compared across all conditions (E). CBE-treated colonoids overall had less potential bursting events when compared to colonoids without CBE. Further, BFT + ibiglustat-treated colonoids had significantly less potential bursting events in the presence of CBE. Single comparisons were made using an unpaired t-test. Statistical significance for comparison between treatment groups is indicated by asterisks: *P < .05 or **P < .01. Error bars represent the standard deviation of the mean. Normal represents colonoids grown under normal conditions, while CBE represents colonoids grown in CBE. Control represents concentrated bacterial culture supernatant from ETBFΔbft and BFT represents concentrated bacterial culture supernatant from ETBF. Confocal images were captured using 10x magnification. Scale bar indicates a distance of 100 μm. Compiled 48-hour time-lapse videos can be found in Videos S5–S8