Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: (a) Body mass at baseline (week 0; striped) and the conclusion (solid) of the 8-week intervention [n=15-19/group]. (b) Plasma ALT [n=9-12/group], (c) plasma AST [n=6-14/group], (d) liver mass [n=15-19/group], (e) liver TG [n=11-17/group], and (f) liver pathological NASH Activity and Brunt Fibrosis Scoring at the conclusion of the 8-week intervention [n=10-14/group]. (g) Representative images of gross morphology, H&E stained (magnification = 20X; scale bar = 50 μm), and Masson’s trichrome stained (magnification = 20X; scale bar = 50 μm) liver at the conclusion of the 8-week intervention. All data are shown as mean ± SEM and were analyzed by paired Student’s t-test within treatment group (a) or one-way ANOVA with Tukey post-hoc testing (b-f). We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. *p<0.05, **p< 0.01, ***p<0.005.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques: Clinical Proteomics, Activity Assay, Staining

Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: (a) Liver TGF-β1 at the conclusion of the 8-week intervention [n=13-17/group]. (b) Collagen synthesis rates during the 8-week intervention [n=14-18/group]. (c) Liver TGF-β1 hydroxyproline content at the conclusion of the 8-week intervention [n=14-19/group]. (d) Fractional percent [n=15-18/group], and (e) absolute content [n=15-18/group] of static and dynamic collagen in liver at the conclusion of the 8-week intervention. All data are shown as mean ± SEM and were analyzed by one-way ANOVA with Tukey post-hoc testing. We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. *p<0.05, **p< 0.01, ***p<0.005.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques:

Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: Liver (a) Loxl2 mRNA [n=9-12/group], and (b) LOXL2 protein [n=14/group] at the conclusion of the 8-week intervention. (c) Representative immunoblots of liver LOXL2 and GAPDH at the conclusion of the 8-week intervention. All data are shown as mean ± SEM and were analyzed by one-way ANOVA with Tukey post-hoc testing. We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. *p<0.05, ***p<0.005.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques: Western Blot

Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: Liver (a) Mmp2 mRNA [n=9-12/group], (b) Mmp9 mRNA [n=9-12/group], (c) Timp1 mRNA [n=8-11/group], (d) MMP2 protein [n=12-17/group], (e) MMP9 protein [n=12-17/group], and (f) TIMP1 protein [n=12-17/group] at the conclusion of the 8-week intervention. (g) Representative immunoblots of liver MMP2, MMP9, TIMP1, and GAPDH at the conclusion of the 8-week intervention. Liver (h) MMP2 [n=10-14/group], and (i) MMP9 [n=11-14/group] enzymatic activity at the conclusion of the 8-week intervention. (j) Representative images of liver gelatin zymography assessing MMP2 and MMP9 enzymatic activity. Signal intensity for each sample was normalized to the intensity of their corresponding positive control standard. All data are shown as mean ± SEM and were analyzed by one-way ANOVA with Tukey post-hoc testing. We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. *p<0.05, **p< 0.01, ***p<0.005.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques: Western Blot, Activity Assay, Zymography, Positive Control

Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: Liver (a) Mmp8 mRNA [n=8-12/group], (b) Mmp12 mRNA [n=9-12/group], (c) Mmp13 mRNA [n=9-12/group], (d) Mmp14 mRNA [n=9-12/group], (e) Mmp16 mRNA [n=9-12/group], and (f) Mmp19 mRNA [n=9-12/group] at the conclusion of the 8-week intervention. All data are shown as mean ± SEM and were analyzed by one-way ANOVA with Tukey post-hoc testing. We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. **p< 0.01, ***p<0.005.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques:

Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: Liver (a) Pparγ mRNA [n=9-12/group], (b) Scd1 mRNA [n=9-12/group], (c) PPARγ protein [n=12-17/group], and (d) SCD1 protein [n=8-14/group] at the conclusion of the 8-week intervention. (e) Representative immunoblots of liver PPARγ, SCD1, and GAPDH at the conclusion of the 8-week intervention. All data are shown as mean ± SEM and were analyzed by one-way ANOVA with Tukey post-hoc testing. We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. *p<0.05, **p< 0.01, ***p<0.005.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques: Western Blot

Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: (a) Liver fetuin-A protein at the conclusion of the 8-week intervention [n=12-17/group]. (b) Representative immunoblots of liver fetuin-A and GAPDH at the conclusion of the 8-week intervention. All data are shown as mean ± SEM and were analyzed by one-way ANOVA with Tukey post-hoc testing. We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. *p<0.05, **p< 0.01.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques: Western Blot

Journal: bioRxiv

Article Title: 17α-estradiol, a lifespan-extending compound, attenuates liver fibrosis by modulating collagen turnover rates in male mice

doi: 10.1101/2022.06.16.496423

Figure Lengend Snippet: (a) Representative immunofluorescence images of F4/80 (total macrophages), CD11c (M1: pro-inflammatory macrophages), and CD206 (M2: anti-inflammatory macrophages) in liver at the conclusion of the 8-week intervention (magnification = 320X; scale bar = 50 μm). Percentage of liver area for (b) F4/80 [n=5-7/group], (c) CD11c [n=5-7/group], and (d) CD206 [n=5-7/group] at the conclusion of the 8-week intervention. (e) Ratio of liver phospho to total p65 at the conclusion of the 8-week intervention [n=12-16/group]. (f) Representative immunoblots of liver phosphor p65, total p65, and GAPDH at the conclusion of the 8-week intervention. All data are shown as mean ± SEM were analyzed by one-way ANOVA with Tukey post-hoc testing. We did not indicate statistical differences between vehicle and 17α-E2 treatment groups (preventive and therapeutic), or between 17α-E2 treatment groups (preventive and therapeutic), for purposes of visual clarity. *p<0.05, ***p<0.005.

Article Snippet: At twelve weeks of age, mice were randomized by body mass into one of five groups , (1) Short-term carbon tetrachloride (CCl 4 ) (n=18): chow-fed (TestDiet 58YP; 66.6% CHO, 20.4% PRO, 13.0% FAT), CCl 4 -treated twice weekly for four weeks, (2) Vehicle (n=15): chow-fed, vehicle-treated (olive oil) controls, (3) CCl 4 (n=15): chow-fed, CCl 4 -treated twice weekly for eight weeks, (4) CCl 4 + 17α preventive (n=18): chow+17α (14.4 mg/kg; Steraloids, Newport, RI)-fed for eight weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks, or (5) CCl 4 + 17α therapeutic (n=15): chow+17α-fed for the final four weeks while simultaneously being CCl 4 -treated twice weekly for eight weeks.

Techniques: Immunofluorescence, Western Blot

Journal: Redox Biology

Article Title: Nlrp6 protects from corticosterone-induced NSPC ferroptosis by modulating RIG-1/MAVS-mediated mitophagy

doi: 10.1016/j.redox.2024.103196

Figure Lengend Snippet: Nlrp6 depletion promotes ferroptotic NSPC death through the system xc − /GSH/GPx4 axis. RNA-seq analysis of the primary NSPCs isolated from WT and Nlrp6 KO fetal mouse. (A) KEGG analysis of down-regulated genes of Nlrp6 KO NSPCs. (B) Enrichment plots in ferroptotic-related pathways by GSEA analysis. (C) Heat map of the core genes involved in ferroptosis. NE-4c cells transfected by control (CTL), siNlrp6 or simultaneously treated by Fer-1 and DFO. (D) Cell survival (n = 8). (E–G) MDA contents, ROS indicence and GSH contents (n = 4). (H–J) The protein expression of FTH1, SLC7A11 and GPX4 (n = 3). Data are expressed as mean ± SEM. Significance was calculated using a one-way ANOVA followed by the Dunnett's post-hoc test. *P < 0.05, **P < 0.01 and ***P < 0.001 compared to CTL group.

Article Snippet: In addition, NE-4c cells were induced by 1 μm corticosterone (HY–B1618, MCE) for 48 h, or simultaneously treated by SCFAs, a mixture of 30 μm sodium acetate (S2889, Procell), 2 μm sodium propionate (P1880, Sigma-Aldrich), 1 μm sodium butyrate (820236, Sigma-Aldrich).

Techniques: RNA Sequencing, Isolation, Transfection, Control, Expressing

Journal: Redox Biology

Article Title: Nlrp6 protects from corticosterone-induced NSPC ferroptosis by modulating RIG-1/MAVS-mediated mitophagy

doi: 10.1016/j.redox.2024.103196

Figure Lengend Snippet: Suppression of mitophagy contributes to NSPC ferroptosis induced by Nlrp6 depletion. The primary NSPCs were isolated from WT and Nlrp6 KO fetal mouse, and cultured for 7 days to neutrospheres, respectively. (A) Enrichment plots in mitophagy-related pathway by GSEA analysis. (B) Heat map of the core genes involved in mitophagy. (C) Flow cytometric analysis for the percentage of MDC positive cells (n = 7). (D–F) The protein expression of PINK1, BNIP3 and NIX (n = 3–4). FCCP was used to activate mitophagy. (G) Representative images of mitochondria labeled with red (Mito-Tracker Red CMXRos, MTR) and blue (DAPI). Scale bar, 100 μm. (H) The protein expression and the ratio of LC3II/I (n = 4). NE-4c cells transfected with CTL, siNlrp6 or simultaneously induced by FCCP. (I) Cell survival (n = 8). (J) The percentage of MDC positive cells (n = 4). (K) The protein expression and the ratio of LC3II/I (n = 4). (L–N) MDA contents, ROS indicence and GSH contents (n = 4). Data are expressed as mean ± SEM. Significance was calculated using a two-tailed unpaired t -test or a one-way ANOVA followed by the Dunnett's post-hoc test. *P < 0.05 and ***P < 0.001 compared to WT group or CTL group.

Article Snippet: In addition, NE-4c cells were induced by 1 μm corticosterone (HY–B1618, MCE) for 48 h, or simultaneously treated by SCFAs, a mixture of 30 μm sodium acetate (S2889, Procell), 2 μm sodium propionate (P1880, Sigma-Aldrich), 1 μm sodium butyrate (820236, Sigma-Aldrich).

Techniques: Isolation, Cell Culture, Expressing, Labeling, Transfection, Two Tailed Test

Journal: Redox Biology

Article Title: Nlrp6 protects from corticosterone-induced NSPC ferroptosis by modulating RIG-1/MAVS-mediated mitophagy

doi: 10.1016/j.redox.2024.103196

Figure Lengend Snippet: Nlrp6 regulates mitophagy through RIG-I/MAVS axis. (A–D) The mRNA and protein levels of DDX58 (RIG-1) and MAVS in primary NSPCs from WT and Nlrp6 KO mice (n = 3–4). (E, F) The protein levels of RIG-1 and LC3II/I of NE-4c cells transfected by CTL, siNlrp6 or simultaneously treated by Ploy (ic) (n = 4). (G, H) The protein expression of RIG-1 and LC3II/I in NE-4c cells induced by CTL, CORT or simultaneously treated by Ploy (ic) (n = 3). (I–L) The protein levels of Nlrp6, RIG-1, MAVS and LC3II/I in NE-4c cells induced by CTL, CORT or simultaneously transfected with Nlrp6 plasmid (n = 3–4). Statistical analysis of the protein levels was adjusted to loading control. Data are expressed as mean ± SEM. Significance was calculated using a two-tailed unpaired t -test or a one-way ANOVA followed by the Dunnett's post-hoc test. *P < 0.05, **P < 0.01 and ***P < 0.001 compared to WT group or CTL group.

Article Snippet: In addition, NE-4c cells were induced by 1 μm corticosterone (HY–B1618, MCE) for 48 h, or simultaneously treated by SCFAs, a mixture of 30 μm sodium acetate (S2889, Procell), 2 μm sodium propionate (P1880, Sigma-Aldrich), 1 μm sodium butyrate (820236, Sigma-Aldrich).

Techniques: Transfection, Expressing, Plasmid Preparation, Control, Two Tailed Test

Journal: Redox Biology

Article Title: Nlrp6 protects from corticosterone-induced NSPC ferroptosis by modulating RIG-1/MAVS-mediated mitophagy

doi: 10.1016/j.redox.2024.103196

Figure Lengend Snippet: SCFAs up-regulates Nlrp6 to protect NSPCs from corticosterone-induced ferroptosis. (A) Nlrp6 luciferase reporter system and the activity of luciferase in NE-4c cells (n = 3). NE-4c cells were induced by CTL, CORT or simultaneously treated with SCFAs. (B) The protein expression of Nlrp6 (n = 4). (C) Cell survival (n = 7). (D–F) MDA contents, ROS indicence and GSH contents (n = 4). (G) The percentage of MDC positive cells (n = 9). (H–J) The protein expression of RIG-1, MAVS and LC3II/I (n = 4). Statistical analysis of the protein levels was adjusted to loading control. Data are expressed as mean ± SEM. Significance was calculated using a two-tailed unpaired t -test or a one-way ANOVA followed by the Dunnett's post-hoc test. *P < 0.05, **P < 0.01 and ***P < 0.001 compared to CTL group.

Article Snippet: In addition, NE-4c cells were induced by 1 μm corticosterone (HY–B1618, MCE) for 48 h, or simultaneously treated by SCFAs, a mixture of 30 μm sodium acetate (S2889, Procell), 2 μm sodium propionate (P1880, Sigma-Aldrich), 1 μm sodium butyrate (820236, Sigma-Aldrich).

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

Journal: European journal of endocrinology

Article Title: ABIRATERONE ACETATE TREATMENT LOWERS 11-OXYGENATED ANDROGENS

doi: 10.1530/EJE-19-0905

Figure Lengend Snippet: List of steroid analytes, standards, internal standards, and mass spectrometry parameters

Article Snippet: Cortisol-d4 , Cerilliant , 367.2/121.1 , 97.0 , 7.3 , .

Techniques: Mass Spectrometry