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. Author manuscript; available in PMC: 2022 Jun 4.
Published in final edited form as: Sci Immunol. 2022 Mar 4;7(69):eabj1080. doi: 10.1126/sciimmunol.abj1080

Fig. 3. Biliary tuft cell abundance is modulated by bile acids.

Fig. 3.

A) Schematic of BA regulation via enterohepatic recirculation. B) Flare25 mice were fed control (ctrl) diet or 2% cholestyramine (2% chol) diet for two wks; IL-25+ tuft cell frequency in biliary epithelial prep was determined by flow cytometry. Representative of three experiments. C) Flare25 mice fed 2% cholestyramine diet for ten days were injected IP every other day with GW4064 (GW4064 + chol) or vehicle (veh + chol) starting on day 1 of diet. Representative of three experiments. D) Flare25 mice were fed 0.5% cholic acid (CA) diet for the indicated time periods; IL-25+ tuft cell frequency in biliary epithelial prep was determined by flow cytometry. E) Wholemount confocal imaging of Flare25 GB from mice fed chow or 0.5% CA diet for three days. F) Germfree (GF) were fed irradiated 0.5% CA diet in sterile isolators. Frequency of DCLK1+ tuft cells among epithelial cells in total GB/EHBD digest was examined by flow cytometry at indicated timepoints and compared to SPF mice fed irradiated 0.5% CA diet. G) Flare25 mice received 0.2% sodium deoxycholate in drinking water ad libitum. Tuft cell frequency among epithelial cells from total GB/EHBD digest was determined by flow cytometry. H,I) Frequency of DCLK1+ tuft cells among epithelial cells from total GB/EHBD digests was analyzed by flow cytometry in age- and sex-matched H) SPF mice from the UCSF vivarium compared to GF mice and I) Fxr −/− and WT controls. J) The frequency of IL-25+ tuft cells among biliary epithelial cells from total GB/EHBD digests was examined by flow cytometry in Flare25 reporter Cyp8b1−/−, Cyp8b1+/−, and Cyp8b1+/+ mice. K-M) Flare25 mice were subjected to bile duct ligation (BDL) or sham surgery. Seven days later total GB/EHBD digests were analyzed by flow cytometry for frequency of IL-25+ tuft cells among epithelial cells (K,L) and frequency of CD45+ immune cells (M). N-P) Total GB/EHBD digests from Pou2f3−/− and littermate controls were analyzed by flow cytometry seven days after BDL or sham surgery. Total immune cells (CD45+, N), macrophages (CD64+CD11b+Ly6G-, O), and neutrophils (Ly6G+CD11b+CD64-, P) were enumerated. Significance determined by unpaired two-tailed T test (H,I,M) or one-way ANOVA (N-P), *p<.05, **p<.01, ***p<.001, ****p<.0001.