Journal: The Journal of Clinical Investigation

Article Title: Hepatic FoxOs link insulin signaling with plasma lipoprotein metabolism through an apolipoprotein M/sphingosine-1-phosphate pathway

doi: 10.1172/JCI146219

Figure Lengend Snippet: ( A ) Liver Apom gene expression in 13-week-old db/db and db/+ control mice. ( B ) Liver Apom gene expression in 25-week-old mice. ( C ) Western blot of ApoM expression in total plasma from 13-week-old and 25-week-old mice. ( D ) Western blot of ApoM and ApoA1 expression in HDL fractionated by sequential density ultracentrifugation from plasma of 13-week-old and 25-week-old mice. ( E ) Representative Western blot of ApoM and ApoA1 expression in ultracentrifuge-fractionated lipoproteins from plasma of 13-week-old mice. ( F – H ) S1P levels in 13-week-old db/db and db/+ control mice. ( F ) Total plasma S1P levels. ( G ) Plasma S1P distribution on ultracentrifuge-fractionated lipoproteins. ( H ) Percentage of plasma S1P distribution. n = 5–6/group. Data are presented as the mean ± SEM. * P < 0.05, ** P < 0.01, and *** P < 0.001, by Student’s t test.

Article Snippet: For the adenovirus experiments, adult male mice were injected intravenously with murine ApoM adenovirus (Welgen) at 0.5 × 10 9 virus particles per gram of body weight, 8 days prior to euthanasia, and were fed a standard chow diet.

Techniques: Gene Expression, Control, Western Blot, Expressing, Clinical Proteomics

Journal: The Journal of Clinical Investigation

Article Title: Hepatic FoxOs link insulin signaling with plasma lipoprotein metabolism through an apolipoprotein M/sphingosine-1-phosphate pathway

doi: 10.1172/JCI146219

Figure Lengend Snippet: ( A ) Hepatic Apom gene expression in adult male mice ( n = 5–8/group). ( B ) Representative Western blots of ApoM expression in liver lysates from adult male mice. C, littermate control mice; F, L-FoxO1,3,4 mice. ( C ) Hepatic Apom gene expression in mice of both sexes that were sacrificed on P2 ( n = 10–20/group). ( D ) Hepatic Apom gene expression following acute knockdown via AAV8.Tbg.Cre in adult male Foxo1 fl/fl , Foxo3 fl/fl , and Foxo4 fl/Y mice ( n = 5/group). Values are shown relative to littermate controls. ( E ) Schematic representation of FoxO1 protein. ( F – H ) Foxo1 , G6pc , and Apom gene expression in primary hepatocytes from WT mice that were transduced with different FoxO1 mutants. ( F ) FoxO1-ADA mutant: the 3 Akt phosphorylation sites are mutated, causing FoxO1 to be constitutively nuclear. Data indicate the mean ± SEM of triplicates of 3 independent experiments. ( G ) FoxO1-ADA-DBD mutant: contains the ADA mutation and a mutation disrupting the DNA binding domain. Data indicate the mean ± SEM of triplicates of 3 independent experiments. ( H ) FoxO1-Δ256 mutant: a dominant-negative version of FoxO1 that lacks the transactivation domain. ( I ) Chip-qPCR of Apom , Igfbp1 , and G6pc from livers of mice with a knockin allele of FoxO1-Venus. The mice were fed either chow or a HFD for 4 weeks. Data are presented as the mean ± SEM. * P < 0.05 and *** P < 0.001, by Student’s t test.

Article Snippet: For the adenovirus experiments, adult male mice were injected intravenously with murine ApoM adenovirus (Welgen) at 0.5 × 10 9 virus particles per gram of body weight, 8 days prior to euthanasia, and were fed a standard chow diet.

Techniques: Gene Expression, Western Blot, Expressing, Control, Knockdown, Transduction, Mutagenesis, Phospho-proteomics, Binding Assay, Dominant Negative Mutation, ChIP-qPCR, Knock-In

Journal: The Journal of Clinical Investigation

Article Title: Hepatic FoxOs link insulin signaling with plasma lipoprotein metabolism through an apolipoprotein M/sphingosine-1-phosphate pathway

doi: 10.1172/JCI146219

Figure Lengend Snippet: Chow-fed mice were injected intraperitoneally with gold-thioglucose or saline and were continued on a chow diet for 13 weeks. ( A ) Total body weight. ( B ) Plasma glucose levels after 5 hours fasting. ( C ) Plasma insulin levels after 5 hours fasting. ( D ) Hepatic Foxo1 , Apom , Scarb1 , and G6pc gene expression. ( E ) Representative Western blot of ApoM and ApoA1 expression in total plasma from chow-fed mice. CS, littermate control mice treated with saline; FS, L-FoxO1,3,4 mice treated with saline; CG, littermate control mice treated with gold-thioglucose; FG, L-FoxO1,3,4 mice treated with gold-thioglucose. * P < 0.05, ** P < 0.01, *** P < 0.001 versus control saline. ## P < 0.01 and ### P < 0.001 versus L-FoxO134-saline. && P < 0.01 versus control-GTG, by 2-way ANOVA. Data are presented as the mean ± SEM. ( n = 5–9/group for all panels)

Article Snippet: For the adenovirus experiments, adult male mice were injected intravenously with murine ApoM adenovirus (Welgen) at 0.5 × 10 9 virus particles per gram of body weight, 8 days prior to euthanasia, and were fed a standard chow diet.

Techniques: Injection, Saline, Clinical Proteomics, Gene Expression, Western Blot, Expressing, Control

Journal: The Journal of Clinical Investigation

Article Title: Hepatic FoxOs link insulin signaling with plasma lipoprotein metabolism through an apolipoprotein M/sphingosine-1-phosphate pathway

doi: 10.1172/JCI146219

Figure Lengend Snippet: ( A ) Representative Western blot of ApoM expression in total plasma from chow-fed mice ( n = 4/group). ( B ) Total S1P levels in plasma from chow-fed mice. Differences were evaluated by Student’s t tests. ( C ) Distribution of S1P in plasma fractionated by size-exclusion chromatography. Cholesterol levels in the control mice are shown as a reference to demonstrate the fractions in which HDL particles were eluted (gray shaded area). VLDL and LDL peaks are not shown, but were eluted in fractions approximately 25 and 35, respectively. ( D ) Distribution of S1P in lipoproteins fractions from mice fed a WTD for 3 weeks. Cholesterol levels from the control mice are shown as a reference to demonstrate the fractions in which HDL particles were eluted (gray shaded area). VLDL and LDL peaks are not shown, but were eluted in fractions approximately 21 and 31, respectively. ( E ) Western blot of ApoA1 and ApoM in lipoprotein fractions from chow diet–fed L-FoxO1,3,4 mice. ( F ) Western blot of ApoA1 and ApoM in lipoprotein fractions from Western diet–fed L-FoxO1,3,4 mice. ( G ) Plasma S1P distribution on ultracentrifuge-fractionated lipoproteins from control, CETP, L-FoxO1,3,4, and L-FoxO1,3,4:CETP mice. ( H ) Cholesterol efflux capacity of HDL isolated from Western diet–fed L-FoxO1,3,4 mice and littermate controls. Each sample is pooled HDL from 2 mice. * P < 0.05 versus control mice, by Kruskal-Wallis 1-way ANOVA; # P < 0.05 versus CETP mice, by Mann-Whitney U post hoc test. Data are presented as the mean ± SEM.

Article Snippet: For the adenovirus experiments, adult male mice were injected intravenously with murine ApoM adenovirus (Welgen) at 0.5 × 10 9 virus particles per gram of body weight, 8 days prior to euthanasia, and were fed a standard chow diet.

Techniques: Western Blot, Expressing, Clinical Proteomics, Size-exclusion Chromatography, Control, Isolation, MANN-WHITNEY

Journal: The Journal of Clinical Investigation

Article Title: Hepatic FoxOs link insulin signaling with plasma lipoprotein metabolism through an apolipoprotein M/sphingosine-1-phosphate pathway

doi: 10.1172/JCI146219

Figure Lengend Snippet: Chow-fed mice were injected intravenously with murine ApoM adenovirus (0.5 × 10 9 virus particles/gram of body weight), 8 days prior to euthanasia. ( A ) Apom and Foxo1 gene expression in liver ( n = 4–8/group). ** P < 0.01 versus controls; # P < 0.05 versus L-FoxO1,3,4-GFP. ( B ) Western blot of expression of ApoM and ApoA1 from HDL fractionated by ultracentrifugation. ( C ) Total plasma S1P levels ( n = 5–6/group). ( D ) Plasma S1P distribution on ultracentrifuge-fractionated lipoproteins ( n = 3–8/group). * P < 0.05 versus control-GFP; # P < 0.05 versus L-FoxO1,3,4-GFP. ( E ) Intraperitoneal glucose tolerance test, 3 days prior to euthanasia ( n = 4–6/group). * P < 0.05, ** P < 0.01, and *** P < 0.001, control-GFP versus L-FoxO1,3,4. # P < 0.05, control-AdApoM versus L-FoxO1,3,4-AdApoM. Statistical significance was determined by Kruskal-Wallis 1-way ANOVA with the Mann-Whitney U post hoc test ( A and C – E ). Data are presented as the mean ± SEM.

Article Snippet: For the adenovirus experiments, adult male mice were injected intravenously with murine ApoM adenovirus (Welgen) at 0.5 × 10 9 virus particles per gram of body weight, 8 days prior to euthanasia, and were fed a standard chow diet.

Techniques: Injection, Virus, Gene Expression, Western Blot, Expressing, Clinical Proteomics, Control, MANN-WHITNEY