Journal: Nature medicine

Article Title: Gut microbiota and intestinal FXR mediate the clinical benefits of metformin

doi: 10.1038/s41591-018-0222-4

Figure Lengend Snippet: a, TR-FRET FXR coactivator recruitment assay to assess whether GUDCA is a FXR antagonist in the presence of CDCA (20 μM). n = 4 replicates/treatment. b, Luciferase activity of control (DMSO), different concentrations of GUDCA or TUDCA without or with FXR agonist CDCA treatment. n = 3 replicates/treatment. P value was determined by one-way ANOVA with Tukey’s correction (F7,16 = 61.69). c, SHP and FGF19 mRNA expression in differentiated Caco-2 cells after treatment with different concentrations of GUDCA or TUDCA with CDCA. n = 3 replicates/treatment. P value was determined by one-way ANOVA with Tukey’s correction (SHP: F7,16 = 4.871, FGF19: F7,16 = 15.41). d, Relative expression of the target gene mRNAs of intestinal FXR in mice treated with vehicle, TCA or TCA and CDCA, GUDCA or TUDCA. n = 5 mice/group. P value was determined by one-way ANOVA with Tukey’s correction (Shp: F4,20 = 38.47, Fgf15: F4,20 = 25.02). e, Relative expression of intestinal Fxr mRNA and its target gene mRNAs in mice treated with metformin on a HFD for 1 week. n = 5 mice/group. P value was determined by two-tailed Student’s t-test (Shp: t8 = 2.779, Fgf15: t8 = 3.383). f, The relative expression of Cyp7a1, Cyp7b1, Cyp8b1 and Cyp27a1 mRNAs in the livers of metformin-treated mice after 1-week HFD feeding. n = 5 mice/group. P value was determined by two-tailed Student’s t-test (Cyp7a1: t8 = 2.805). g, Relative abundance of intestinal Fxr mRNA and its target gene mRNAs in metformin-treated Ampka1fl/fl and Ampka1ΔIE mice on a HFD for 1 week. n = 5 mice/group. P value was determined by one-way ANOVA with Tukey’s correction (Shp: F3,16 = 8.046, Fgf15: F3,16 = 11.83). h, Relative expression of intestinal Fxr mRNA and its target gene mRNAs in antibiotics-treated, microbiota-depleted mice treated with metformin on a HFD for 1 week. n = 5 mice/group. P value was determined by one-way ANOVA with Tukey’s correction (Shp: F3,16 = 6.859, Fgf15: F3,16 = 8.782). i, In microbiota-depleted mice, metformin had no effect in inhibiting the activation of intestinal FXR on top of TCA. The relative expression of Fxr and its target genes in the ileum. n = 6 mice/group. All the data are presented as the mean ± s.e.m.

Article Snippet: After 24 h post-transfection, the cells were exposed to GUDCA (50–200 μM, Sigma-Aldrich, Cat# 06863), or TUDCA (50–200 μM, Sigma-Aldrich, Cat# T0266) in the presence of CDCA (20 μM, Targetmol, Cat# T0847) for 16 h. Luciferase assays were performed using the dual-luciferase assay system (Promega).

Techniques: Luciferase, Activity Assay, Expressing, Two Tailed Test, Activation Assay

Journal: Journal of Lipid Research

Article Title: Parenteral lipids shape gut bile acid pools and microbiota profiles in the prevention of cholestasis in preterm pigs [S]

doi: 10.1194/jlr.RA120000652

Figure Lengend Snippet: Bile acid concentrations in gallbladder contents of preterm piglets after 22 days

Article Snippet: The free and conjugated bile acid concentrations in samples were determined by reverse isotope dilution methodology using internal standards of d9-chenodeoxycholic acid (CDCA) and d9-glyco-CDCA (Cambridge Isotopes, Andover, MA).

Techniques:

Journal: Journal of Lipid Research

Article Title: Parenteral lipids shape gut bile acid pools and microbiota profiles in the prevention of cholestasis in preterm pigs [S]

doi: 10.1194/jlr.RA120000652

Figure Lengend Snippet: Colonic contents bile acid profiles and relative ratios of conjugated to unconjugated bile acids. A: Heatmap showing profiles of bile acid species detected in samples from colon contents, with bile acid species arranged by hierarchical clustering. Values are relative standardized units (dimensionless). B: Relative ratios of total conjugated (both glyco- and tauro-conjugated) to unconjugated chenodeoxycholate (CDCA), hyocholate (HCA), and hyodeoxycholate (HDCA) found in colon contents samples. Values are ratios of relative standardized units (dimensionless). Bars represent standard error. *P < 0.05 versus ENT, #P < 0.05 versus IL (Holm-adjusted pairwise t-tests).

Article Snippet: The free and conjugated bile acid concentrations in samples were determined by reverse isotope dilution methodology using internal standards of d9-chenodeoxycholic acid (CDCA) and d9-glyco-CDCA (Cambridge Isotopes, Andover, MA).

Techniques:

Journal: JCI Insight

Article Title: Lipidomics unveils lipid dyshomeostasis and low circulating plasmalogens as biomarkers in a monogenic mitochondrial disorder

doi: 10.1172/jci.insight.123231

Figure Lengend Snippet: (A–D) LC-QTOF–based untargeted lipidomics from livers of fed H-Lrpprc–/– mice (n = 13) and their littermate controls (n = 8). (A) Volcano plot depicting the 1,386 features obtained following MS data processing. Using a P-corr threshold of 0.05 (corresponding to an uncorrected P value of 0.015; horizontal red dotted line) and an FC >2.5 or <0.4 (vertical red dotted lines), 92 features significantly discriminated H-Lrpprc–/– mice vs. controls, of which 60 were increased (red dots) and 32 decreased (green dots). See also Supplemental Table 3 for the list of lipids identified by MS/MS with FCs and P values (unpaired Student’s t test and with Benjamini-Hochberg correction). (B–D) Dot plots of selected lipids significantly discriminating H-Lrpprc–/– mice from controls and identified by MS/MS. Each dot represents a log2-transformed signal intensity ratio for the indicated lipid (sub)classes with their acyl side chain(s): (B) 15 glycerolipids (TGs); (C) 22 glycerophospholipids: 9 PC, 9 PE, 2 phosphatidylglycerol (PG) and 2 phosphatidylserine (PS); and (D) 4 ACs.

Article Snippet: Briefly, plasma samples or pulverized liver tissues previously freeze-clamped (20 mg) were homogenized in methanol/water (80:20, v/v) spiked with deuterated standards ([ 2 H 4 ]lithocholic acid, d 4 -LCA; [ 2 H 4 ]cholic acid, d 4 -CA; [ 2 H 4 ]chenodeoxycholic acid, d 4 -CDCA; [ 2 H 4 ]deoxycholic-2,2,4,4-d 4 acid, d 4 -DCA); [ 2 H 4 ]glycodeoxycholic acid, d 4 -GDCA; [ 2 H 4 ]glycochenodeoxycholic acid, d 4 -GCDCA; and [ 2 H 4 ]glycocholic acid, d 4 -GCA) (CDN Isotopes) and then acidified with HCl (1 mM) before homogenization and centrifugation.

Techniques: Tandem Mass Spectroscopy, Transformation Assay