hydroxycholesterol d4  (LGC Standards)

Journal: The Journal of Endocrinology

Article Title: AKR1D1 knockout mice develop a sex-dependent metabolic phenotype

doi: 10.1530/JOE-21-0280

Figure Lengend Snippet: Hepatic and serum bile acids are lower in Akr1d1 –/– mice with sexually dimorphic changes to bile acid metabolism and composition. Mature (30 weeks) male and female Akr1d1 –/– mice have decreased total hepatic and serum bile acids (A) and altered bile acid composition with reduced 12α-hydroxylated/non-12α-hydroxylated bile ratio in the liver and serum (B and C) ( n = 12–16 mice). Principal component analysis shows greater divergence from WT in male Akr1d1 –/– mice (D). Akr1d1 deletion has a sexually dimorphic effect on mRNA expression of hepatic bile acid-metabolising genes and levels of bile acid intermediates. Cyp8b1 expression is increased in Akr1d1 –/– females but not in males (E) ( n = 10 mice) as are the AKR1D1 substrates 7α,12α-dihydroxy-4-chol-3-one (F) and 7α-hydroxy-4-chol-3-one (G) ( n = 10 mice). The oxysterol 27-hydroxycholesterol (27-OHC) is decreased in Akr1d1 –/– females (H) ( n = 9 mice). Female Akr1d1 –/– mice also have increased expression of the bile acid-detoxifying genes Cyp3a11 , Cyp2c55 , Cyp4a12a and Sult2a7 (I) ( n = 10 mice) and serum levels of LCA sulphate (J) ( n = 9 mice). Data are presented as mean ± s.d. , log2(FC), ratio or mean relative abundance. * P < 0.05, ** P < 0.01, *** P < 0.005, ∅ P < 0.001, ∅∅ P < 0.0005, ∅∅∅ P < 0.0001 compared to WT. P values for bile acid composition compare WT and Akr1d1 –/– within sex (male/female). (WT = WT C57BL/6; –/– = Akr1d1 –/– ). A full colour version of this figure is available at https://doi.org/10.1530/JOE-21-0280 .

Article Snippet: Principal component analysis was performed using the FactoMineR package ( Lê et al. 2008 ) and the factoextra package to visualise the results in R. Free and esterified oxysterols were measured as previously described ( ) with the following modifications: Liver (100 mg) was spiked with 30 μL of 1 μM internal standard mix 25 (R/S), 26-hydroxycholesterol-d4, 7α-hydroxy-4-cholesten-3-one-d7, 7α,12α-dihydroxycholest-4-en-3-one-d7) (Toronto Research Chemicals, Ontario, Canada) and homogenised in chloroform/methanol (4 mL: CHCl 3 /MeOH, 2:1, v/v) containing 50 μg/mL butylated hydroxytoluene.

Techniques: Expressing

Journal: The Journal of Endocrinology

Article Title: AKR1D1 knockout mice develop a sex-dependent metabolic phenotype

doi: 10.1530/JOE-21-0280

Figure Lengend Snippet: Akr1d1 deletion does not protect male C57BL/6 mice against diet-induced obesity, hypercholesterolemia or insulin resistance but improves hypertriglyceridemia. On a high fat diet (HFD), male Akr1d1 –/– mice (grey line) gain weight at the same rate as their WT littermates (black line) (A) and after 20 weeks body composition is not different between genotypes (lean mass lower bar; fat mass upper bar) (B) ( n = 14–15 mice). Adipose weight (C) ( n = 14–15), hepatic triacylglycerol (D) ( n = 12–15), HDL and total cholesterol (E) ( n = 14–15) were unchanged between HFD fed WT and Akr1d1 –/– males. Serum triacylglycerol was reduced in HFD fed Akr1d1 –/– males but not to levels seen in the control diet (F) ( n = 10 mice). Akr1d1 –/– males were not protected against diet-induced reduction in ipGTT (G) or ipITT (H) ( n = 14–15 mice). Data are presented as mean ± s.d .* P < 0.05, ** P < 0.01, *** P < 0.005 compared to wildtype (WT = WT C57BL/6; –/– = Akr1d1 –/– ).

Article Snippet: Principal component analysis was performed using the FactoMineR package ( Lê et al. 2008 ) and the factoextra package to visualise the results in R. Free and esterified oxysterols were measured as previously described ( ) with the following modifications: Liver (100 mg) was spiked with 30 μL of 1 μM internal standard mix 25 (R/S), 26-hydroxycholesterol-d4, 7α-hydroxy-4-cholesten-3-one-d7, 7α,12α-dihydroxycholest-4-en-3-one-d7) (Toronto Research Chemicals, Ontario, Canada) and homogenised in chloroform/methanol (4 mL: CHCl 3 /MeOH, 2:1, v/v) containing 50 μg/mL butylated hydroxytoluene.

Techniques: Control

Journal: The Journal of Endocrinology

Article Title: AKR1D1 knockout mice develop a sex-dependent metabolic phenotype

doi: 10.1530/JOE-21-0280

Figure Lengend Snippet: Bile acid profile in male Akr1d1 –/– mice on a high fat diet (HFD). After 20 weeks on a HFD, male Akr1d1 –/– mice have reduced total liver (A) and serum (B) bile acids and altered bile acid composition (liver: C; serum D) compared to high fat-fed WT littermates. Serum 12α-hydroxylated/non-12α-hydroxylated bile ratio in the liver (E) and serum (F) is reduced compared to control and HFD fed WT littermates. Data are presented as mean ± s.d. in n =10–15 mice. * P < 0.05, ** P < 0.01, *** P < 0.005, ∅ P < 0.001, ∅∅ P < 0.0005, ∅∅∅ P < 0.0001. WT control diet data are presented for comparison, P value for bile acid composition compares WT and Akr1d1 –/– on HFD (WT = WT C57BL/6; –/– = Akr1d1 –/– ). A full colour version of this figure is available at https://doi.org/10.1530/JOE-21-0280 .

Article Snippet: Principal component analysis was performed using the FactoMineR package ( Lê et al. 2008 ) and the factoextra package to visualise the results in R. Free and esterified oxysterols were measured as previously described ( ) with the following modifications: Liver (100 mg) was spiked with 30 μL of 1 μM internal standard mix 25 (R/S), 26-hydroxycholesterol-d4, 7α-hydroxy-4-cholesten-3-one-d7, 7α,12α-dihydroxycholest-4-en-3-one-d7) (Toronto Research Chemicals, Ontario, Canada) and homogenised in chloroform/methanol (4 mL: CHCl 3 /MeOH, 2:1, v/v) containing 50 μg/mL butylated hydroxytoluene.

Techniques: Control, Comparison

Journal: bioRxiv

Article Title: Akr1d1-/- mice have a sexually dimorphic metabolic phenotype with reduced fat mass, increased insulin sensitivity and hypertriglyceridemia in males

doi: 10.1101/2021.02.02.429227

Figure Lengend Snippet: Akr1d1 deletion (A: western blot, liver), does not alter serum corticosterone levels (B) (n = 10 mice), or hepatic mRNA expression of glucocorticoid responsive genes, Sgk1, Gilz, Dusp1 and 11 β hsd1 (C) (n = 10 mice). Male and female Akr1d1-/- mice have decreased total hepatic (D) and serum (E) bile acids and altered bile acid composition with reduced 12α-hydroxylated/non-12α-hydroxylated bile ratio in the liver (F & G) and serum (H & I) (n = 12-15 mice). Principle Component Analysis shows greater divergence from wildtype in male Akr1d1-/- mice (J). Akr1d1 deletion has a sexually dimorphic effect on mRNA expression of hepatic bile acid metabolizing genes and levels of bile acid intermediates. Cyp8b1 expression is increased in Akr1d1-/- females but not males (K) (n = 10 mice) as are the AKR1D1 substrates 7α,12α-dihydroxy-4-chol-3-one (L) and 7α-hydroxy-4-chol-3-one (M) (n = 10 mice). The oxysterol 27-hydroxycholesterol (27-OHC) is decreased in Akr1d1-/- females (N) (n = 10 mice). Female Akr1d1-/- mice also have increased expression of the bile acid detoxifying genes Cyp3a11, Cyp2c55, Cyp4a12a and Sult2a7 (O) (n = 10 mice) and serum levels of LCA sulfate (P) (n = 10 mice). Data are presented as mean±se, log2(FC), ratio, or mean relative abundance. *p<0.05, * *p<0.01, * * *p<0.005, Ø p<0.001, ØØ p< 0.0005, ØØØ p<0.0001 compared to wildtype. mRNA expression was measured by RNASeq. p-values for bile acid composition compare WT and Akr1d1-/- within sex (male/female). (WT = wildtype C57BL/6; -/-= Akr1d1-/- )

Article Snippet: Principal component analysis was performed using the FactoMineR package and the factoextra package to visualise the results in R. Free and esterified oxysterols were measured as previously described with the following modifications: Liver (100 mg) was spiked with 30 μl of 1 μM internal standard mix 25(R/S), 26-Hydroxycholesterol-d4, 7α-hydroxy-4-cholesten-3-one-d7, 7α,12α-dihydroxycholest-4-en-3-one-d7) (Toronto Research Chemicals, Ontario, Canada) and homogenized in chloroform/methanol (4 ml: CHCl3/MeOH, 2:1, v/v) containing 50 μg/mL butylated hydroxytoluene.

Techniques: Western Blot, Expressing

Journal: bioRxiv

Article Title: Akr1d1-/- mice have a sexually dimorphic metabolic phenotype with reduced fat mass, increased insulin sensitivity and hypertriglyceridemia in males

doi: 10.1101/2021.02.02.429227

Figure Lengend Snippet: Male Akr1d1-/- mice on HFD have reduced total liver (A) and serum (B) bile acids and altered bile acid composition (liver: C; serum D) compared to high fat fed WT littermates. Serum 12α-hydroxylated/non-12α-hydroxylated bile ratio in the liver (E) and serum (F) is reduced compared to control and HFD fed WT littermates. Data are presented as mean±se in n=10-15 mice. *p<0.05, * *p<0.01, * * *p<0.005, Ø p<0.001, ØØ p< 0.0005, ØØØ p<0.0001. WT control diet data is presented for comparison, p value for bile acid composition compares WT and Akr1d1-/- on HFD. (WT = wildtype C57BL/6; -/- = Akr1d1-/- )

Article Snippet: Principal component analysis was performed using the FactoMineR package and the factoextra package to visualise the results in R. Free and esterified oxysterols were measured as previously described with the following modifications: Liver (100 mg) was spiked with 30 μl of 1 μM internal standard mix 25(R/S), 26-Hydroxycholesterol-d4, 7α-hydroxy-4-cholesten-3-one-d7, 7α,12α-dihydroxycholest-4-en-3-one-d7) (Toronto Research Chemicals, Ontario, Canada) and homogenized in chloroform/methanol (4 ml: CHCl3/MeOH, 2:1, v/v) containing 50 μg/mL butylated hydroxytoluene.

Techniques: Control, Comparison