Extended Data Fig. 7. Interaction between p-AMPKα and SMPD3 in intestinal ceramide production.

a-c, HFHCD-fed SPF mice were treated with nicotine water or nicotine water plus 10 mg/kg GW4869 (by daily gavage) for 2 weeks, and ileal organoids were then isolated and cultured for 7 days and treated with GW4869 (10 μM) and nicotine (1 μg/mL) for the last 3 days before the detection of ceramide production and secretion (n = 5 mice/group). a, nSMase activity. b, Ceramide profiles in isolated organoids. c, Ceramide profiles in the supernatant of isolated organoids. d-f, HFHCD-fed Prkaa1^fl/fl and Prkaa1^ΔIE mice (SPF) were treated with nicotine water for 2 weeks, and organoids were then isolated and infected with LV (lentivirus)-Ctrl or LV-Smpd3, the infected organoids were plated and cultured for 7 days and treated with nicotine (1 μg/mL) for the last 3 days before the detection of ceramide production and secretion. Western blot analysis for verifying SMPD3 overexpression. (n = 3 mice/group) (d). Ceramide profiles in isolated organoids. (n = 8 mice/group) (e). Ceramide profiles in the supernatant of isolated organoids. (n = 8 mice/group) (f). g, HFHCD-fed WT mice were transplanted with PBS, B. xylanisolvens colonization, nicotine water, nicotine water plus B. xylanisolvens colonization, and nicotine water plus nicX knock-out B. xylanisolvens colonization for 20 weeks (SPF, n = 6 mice/group), and ileal tissues were collected for the ceramide profiles detection. Data are the means ± s.e.m. a, b, Two-tailed Student’s t-test. c, Two-tailed Mann-Whitney U-test. f, One-way ANOVA with Tukey’s post hoc test. e, g, Kruskal-Wallis test with Dunn’s test.