Journal: The Journal of experimental biology

Article Title: Different fuel regulation in two types of myofiber results in different antioxidant strategies in Daurian ground squirrels ( Spermophilus dauricus ) during hibernation.

doi: 10.1242/jeb.231639

Figure Lengend Snippet: Fig. 1. Expression of REDD1 in SOL and EDL muscles of Daurian ground squirrels during hibernation. Protein expression levels were visualized at four sampling points: pre-hibernation (PRE), hibernation (HIB), interbout arousal (IBA) and post-hibernation (POST). See Materials and Methods for more extensive definitions. Representative immunoblots and Stain-Free total protein loading controls are shown below for selected pairs of sampling points and muscles that are labeled on the top of the blot. Data were analyzed using an ANOVA with a post hoc Tukey’s test (P<0.05); values that are not statistically different from each other share the same letter notation.

Article Snippet: Membranes were then blocked with 4% Mowiol® PVA-203 (Aladdin, China) in Tris-buffered saline (TBS) for 10 min (Gholap et al., 2005; Rodda and Yamazaki, 1994), and incubated with REDD1 (Proteintech, 10638-1-AP), atrogin-1 (Proteintech, 12866-1-AP), PGC-1α (Novus, NBP1-04676SS), FOXO1 (C29H4) rabbit mAb (CST#2880), phospho-FOXO1 (Ser256) antibody (CST#9461), FOXO4 (CST#9472) and FOXO4 (phospho-Thr451) (Signalway antibody, 12053) in 0.1% TBST (TBS with 0.1% Tween 20) containing 2% polyvinylpyrrolidone (PVP-40; Amresco, USA, 0507-500G) at 4°C overnight.

Techniques: Expressing, Muscles, Sampling, Western Blot, Staining, Labeling

Journal: Nature Communications

Article Title: A REDD1/TXNIP pro-oxidant complex regulates ATG4B activity to control stress-induced autophagy and sustain exercise capacity

doi: 10.1038/ncomms8014

Figure Lengend Snippet: ( a ) Physical association of REDD1 with both transfected GFP-TXNIP (left) and endogenous TXNIP (right) following transfection of HA-REDD1 and IP with α-TXNIP in 293T cells. ( b ) The endogenous REDD1/TXNIP complex is induced by hypoxia (1% O 2 , 16 h) and energy stress (2-deoxyglucose (2-DG), 20 mM, 16 h) in 293T cells. Lysates were subjected to IP with α-REDD1 antibody. ( c ) Increased thioredoxin (TRX) antioxidant activity in Redd1 −/− and Txnip −/− cells, measured by reduction of insulin disulfides. Shown is the mean of three independent experiments performed in triplicate. ( d ) Reduced H 2 O 2 in Txnip −/− MEFs stained with CM-H2DCFDA (3 μM) followed by flow cytometry. ( e ) TXNIP fails to induce H 2 O 2 in Redd1 −/− cells unless co-transfected with REDD1. ( f ) Co-transfection of REDD1 and TXNIP potently inhibits TRX activity in Redd1 −/− cells. ( g ) Co-transfection of REDD1 and TXNIP is sufficient to suppress ATG4B activity in Redd1 −/− cells, assessed as in . ( h ) Impaired autophagy under basal and hypoxic (1% O 2 , 4 h) conditions in Txnip −/− MEFs, shown in the absence or presence of CQ (30 μM, 4 h). ( i ) Reduced MMP in Txnip −/− cells, assessed by staining with TMRE (100 nM) followed by flow cytometry. ( j ) Expression of REDD1 or TXNIP is sufficient to induce autophagy in transfected 293T cells. *** P <0.001, ** P <0.01, by paired t -test ( c – f , i ) or repeated measures ANOVA ( g ).

Article Snippet: Blots were incubated with antibodies recognizing the following proteins: LC3B (Cell Signaling, no. 2775), β-tubulin (Millipore MAB 3408), p62 (Cell Signaling no. 5114), TXNIP (MBL Laboratories K0204-3 and Santa Cruz Biotechnologies sc-33099), REDD1 (Bethyl Laboratories A302-169A and Santa Cruz Biotechnologies sc-46034), phospho-S6 S235/S236 (Cell Signaling no. 2211), total S6 (Cell Signaling no. 2217), phospho-S6 Kinase T389 (Cell Signaling no. 9205), Hypoxia-inducible factor 1-α (HIF-1α) (Novus Biologicals NB100-449), ATG4B (Santa Cruz Biotechnologies sc-130968), haemagglutinin (HA) epitope (Covance MMS-101R) and TOM20 (Santa Cruz Biotechnologies sc-11415).

Techniques: Transfection, Antioxidant Activity Assay, Staining, Flow Cytometry, Cotransfection, Activity Assay, Expressing

Journal: Nature Communications

Article Title: A REDD1/TXNIP pro-oxidant complex regulates ATG4B activity to control stress-induced autophagy and sustain exercise capacity

doi: 10.1038/ncomms8014

Figure Lengend Snippet: ( a ) Exercise (treadmill run, 2 h) induces autophagy in the muscle of wild-type mice, as assessed by LC3B processing. RPS6 (S6) protein, loading control. Duplicate panels correspond to individual mice. ( b ) REDD1 and TXNIP mRNA induction by exercise (treadmill run, 2 h) in the skeletal muscle of wild-type mice, measured using QRT–PCR relative to β-actin. Numbers refer to individual mice. ( c ) Exercise suppresses ATG4B activity in the heart of wild-type mice, assessed as in . Shown are the mean values from three mice per condition. ( d ) Hyperactivity of ATG4B in the heart and skeletal muscle of treadmill-exercised Redd1 −/− mice, assessed and quantitated as in c . ( N =3 mice per genotype.) ( e ) Increased TRX activity in the heart of treadmill-exercised Redd1 −/− mice, measured as in . ( N =3 mice per genotype.) ( f ) Impaired autophagy induction by treadmill exercise in the muscle of Redd1 −/− . Duplicate panels correspond to individual mice. ( g ) Decreased exercise capacity in Redd1 −/− mice (voluntary running wheel, 24 h). Each point represents an individual run; eight mice per genotype were subjected to at least three runs each; error bars, s.e.m. ( h ) Increased mitochondrial (mt) number in the muscle of treadmill-exercised Redd1 −/− compared with wild-type mice. ( N =3 mice per genotype.) ( i ) Reduced MMP in tissues as in h , assessed in cell-free mitochondrial fractions. ( j ) ATP concentration measured in lysates of tissues described in h . Except as noted, all error bars indicate s.d. *** P <0.001, ** P <0.01, by paired t -test ( b , e , h , j ), or repeated measures ANOVA ( c , d ), or Mann–Whitney U -test ( g ).

Article Snippet: Blots were incubated with antibodies recognizing the following proteins: LC3B (Cell Signaling, no. 2775), β-tubulin (Millipore MAB 3408), p62 (Cell Signaling no. 5114), TXNIP (MBL Laboratories K0204-3 and Santa Cruz Biotechnologies sc-33099), REDD1 (Bethyl Laboratories A302-169A and Santa Cruz Biotechnologies sc-46034), phospho-S6 S235/S236 (Cell Signaling no. 2211), total S6 (Cell Signaling no. 2217), phospho-S6 Kinase T389 (Cell Signaling no. 9205), Hypoxia-inducible factor 1-α (HIF-1α) (Novus Biologicals NB100-449), ATG4B (Santa Cruz Biotechnologies sc-130968), haemagglutinin (HA) epitope (Covance MMS-101R) and TOM20 (Santa Cruz Biotechnologies sc-11415).

Techniques: Quantitative RT-PCR, Activity Assay, Concentration Assay, MANN-WHITNEY

Journal: Nature Communications

Article Title: A REDD1/TXNIP pro-oxidant complex regulates ATG4B activity to control stress-induced autophagy and sustain exercise capacity

doi: 10.1038/ncomms8014

Figure Lengend Snippet: REDD1 and TXNIP are upregulated by physiologic stress, forming a protein complex required for induction of ROS that inhibit ATG4B-mediated LC3 delipidation and thereby promote autophagosome maturation. In the absence of REDD1, decreased ROS results in hyperactivity of ATG4B and disabled autophagy, leading to accumulation of aged, dysfunctional mitochondria and defective oxidative metabolism that compromises ATP generation and exercise capacity.

Article Snippet: Blots were incubated with antibodies recognizing the following proteins: LC3B (Cell Signaling, no. 2775), β-tubulin (Millipore MAB 3408), p62 (Cell Signaling no. 5114), TXNIP (MBL Laboratories K0204-3 and Santa Cruz Biotechnologies sc-33099), REDD1 (Bethyl Laboratories A302-169A and Santa Cruz Biotechnologies sc-46034), phospho-S6 S235/S236 (Cell Signaling no. 2211), total S6 (Cell Signaling no. 2217), phospho-S6 Kinase T389 (Cell Signaling no. 9205), Hypoxia-inducible factor 1-α (HIF-1α) (Novus Biologicals NB100-449), ATG4B (Santa Cruz Biotechnologies sc-130968), haemagglutinin (HA) epitope (Covance MMS-101R) and TOM20 (Santa Cruz Biotechnologies sc-11415).

Techniques:

Journal: F1000Research

Article Title: Validation of a commercially available anti-REDD1 antibody using RNA interference and REDD1-/- mouse embryonic fibroblasts

doi: 10.12688/f1000research.7691.1

Figure Lengend Snippet: Antibodies used and their manufacturer details.

Article Snippet: REDD1 levels were then immediately visualized by Western blot experiment using the Proteintech anti-REDD1 antibody (10638-1-AP).

Techniques:

Journal: F1000Research

Article Title: Validation of a commercially available anti-REDD1 antibody using RNA interference and REDD1-/- mouse embryonic fibroblasts

doi: 10.12688/f1000research.7691.1

Figure Lengend Snippet: Sense strand sequences of the shRNA constructs.

Article Snippet: REDD1 levels were then immediately visualized by Western blot experiment using the Proteintech anti-REDD1 antibody (10638-1-AP).

Techniques: shRNA, Construct, Sequencing, Control

Journal: F1000Research

Article Title: Validation of a commercially available anti-REDD1 antibody using RNA interference and REDD1-/- mouse embryonic fibroblasts

doi: 10.12688/f1000research.7691.1

Figure Lengend Snippet: REDD1 was detected using Proteintech’s 10638-1-AP anti-REDD1 antibody ( A ). Signal intensity was measured by densitometry analysis and blotted relative to α-tubulin control signal, n=2 ( B ).

Article Snippet: REDD1 levels were then immediately visualized by Western blot experiment using the Proteintech anti-REDD1 antibody (10638-1-AP).

Techniques: Control

Journal: F1000Research

Article Title: Validation of a commercially available anti-REDD1 antibody using RNA interference and REDD1-/- mouse embryonic fibroblasts

doi: 10.12688/f1000research.7691.1

Figure Lengend Snippet: REDD1 bands were indirectly detected using Proteintech’s 10638-1-AP anti-REDD1 antibody (n=3). Membrane exposure = 20 minutes. Numbers and block arrows indicate positions of 37 kDa and 25 kDa bands of the MW marker.

Article Snippet: REDD1 levels were then immediately visualized by Western blot experiment using the Proteintech anti-REDD1 antibody (10638-1-AP).

Techniques: Membrane, Blocking Assay, Marker

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a Weight gain in Redd1 −/− mice and their WT littermates fed NC or HFD for 16 weeks ( n = 6 per group). b Mass of eWAT and iWAT in NC- or HFD-fed Redd1 −/− mice and their WT littermates ( n = 8 per group). c Representative images of perilipin (green) and F4/80 (purple) staining in the eWAT of NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 8 per group). Scale bar, 100 μm. d Average adipocyte size in the eWAT of NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 8 per group). e Relative area of F4/80-positive cells in the eWAT of NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 8 per group). f Relative number of crown-like structures (CLSs) in the eWAT of NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 8 per group). g NF-κB activity in the eWAT from NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 6 per group). h Plasma levels of inflammatory cytokines in NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 6 per group). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using two-way ANOVA followed by the Holm–Sidak post hoc test. Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Staining, Activity Assay, Clinical Proteomics

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a Fasting plasma levels of glucose and insulin in NC- or HFD-fed Redd1 −/− mice and their WT littermates ( n = 6 per group). b Representative images of insulin (green)-stained pancreatic islets from NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 6 per group). Scale bar, 100 μm. c Quantification of average islet size ( n = 6 per group). d Calculation of the HOMA-IR scores ( n = 6 per group). e Assessment of GTT and ITT in mice fasting for 12 and 6 h, respectively, in NC- or HFD-fed Redd1 −/− mice and WT littermates ( n = 8 per group). f Representative western blots of Akt phosphorylation and plasma membrane-associated GLUT4 (PM-GLUT4) in eWAT and skeletal muscle from mice injected i.p. with saline or insulin ( n = 3). g Representative western blots of phosphorylated Akt and FOXO1 in the liver of mice injected with saline or insulin ( n = 6). h Quantification of the phosphorylated FOXO1 to total FOXO1 ratio ( n = 6 per group). i Quantification of G6pc , Pck1 , and Fbp1 mRNA levels in the liver ( n = 6 per group). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using two-way ANOVA followed by the Holm–Sidak post hoc test. Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Clinical Proteomics, Staining, Western Blot, Phospho-proteomics, Membrane, Injection, Saline

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a Weight gain over time in Redd1 fl/fl ( R fl/fl ) and Redd1 Δ Adipoq ( R Δ Adipoq ) mice fed HFD for 16 weeks ( n = 6 per group). b Mass measurements for the eWAT and iWAT in HFD-fed Redd1 fl/fl and Redd1 Δ Adipoq mice ( n = 6 per group). c Representative images showing perilipin (green) and F4/80 (purple) staining in the eWAT of Redd1 fl/fl and Redd1 Δ Adipoq mice fed HFD ( n = 6 per group). Scale bar, 100 μm. d NF-κB activity in the eWAT from HFD-fed Redd1 fl/fl and Redd1 Δ Adipoq mice ( n = 6 per group). e , f Plasma levels of inflammatory cytokines in HFD-fed Redd1 fl/fl and Redd1 Δ Adipoq mice ( n = 6 per group). g Fasting plasma levels of glucose and insulin in HFD-fed Redd1 fl/fl and Redd1 Δ Adipoq mice ( n = 6 per group). h Assessment of GTT and ITT in HFD-fed Redd1 fl/fl and Redd1 Δ Adipoq mice after fasting for 12 and 6 h, respectively ( n = 5 per group). i , j Representative western blots of phosphorylated IRS-1 and Akt in the eWAT and skeletal muscle ( i ) and phosphorylated Akt and FOXO1 in the liver ( j ) of NC- or HFD-fed Redd1 fl/fl and Redd1 Δ Adipoq mice after i.p. injection of saline or insulin ( n = 3). k Relative expression levels of G6pc , Pck1 , and Fbp1 in the liver of HFD-fed Redd1 fl/fl and Redd1 Δ Adipoq mice compared with NC-fed mouse groups ( n = 6 per group). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using an unpaired two-tailed t-test. Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Staining, Activity Assay, Clinical Proteomics, Western Blot, Injection, Saline, Expressing, Two Tailed Test

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a Weight gain in Redd1 fl/fl ( R fl/fl ) and Redd1 Δ LysM ( R Δ LysM ) mice fed HFD for 16 weeks ( n = 5 per group). b Measurement of fat (eWAT + iWAT) mass in HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice ( n = 5 per group). c Representative images showing perilipin (green) and F4/80 (purple) staining in the eWAT of HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice ( n = 5 per group). Scale bar, 100 μm. d NF-κB activity in the eWAT from HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice ( n = 5 per group). e Plasma levels of inflammatory cytokines in HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice ( n = 5 per group). f Fasting plasma levels of glucose and insulin in HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice ( n = 5 per group). g Calculation of HOMA-IR scores in HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice ( n = 5 per group). h Assessment of GTT and ITT in HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice fasting for 12 and 6 h, respectively ( n = 5 per group). i , j Representative western blots of the insulin-responsive phosphorylation of IRS-1 and Akt in the eWAT and skeletal muscle ( i ) and phosphorylation of Akt and FOXO1 in the liver ( j ) of NC- or HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice ( n = 3). k Relative expression levels of G6pc , Pck1 , and Fbp1 in the liver of HFD-fed Redd1 fl/fl and Redd1 Δ LysM mice compared with NC-fed mouse groups ( n = 5 per group). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using an unpaired two-tailed t -test. Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Staining, Activity Assay, Clinical Proteomics, Western Blot, Phospho-proteomics, Expressing, Two Tailed Test

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a – c Representative oil red-O (ORO)-stained images of WT and Redd1 −/− SVF cells ( a ), shControl (shC)- or sh-Redd1-transfected 3T3-L1 cells ( b ), and WT ( Redd1 fl/f l , R fl/fl ) and Redd1 Δ Adipoq ( R Δ Adipoq ) SVF cells ( c ) when cultured in differentiation medium (MDI) and quantification of relative ORO intensity ( n = 4). d – f , Expression levels of adipogenic genes ( d ), REDD1 ( e ), and lipogenic genes ( f ) in R fl/fl and R Δ Adipoq SVF cells cultured in MDI medium and quantification of relative ORO intensity ( n = 4). g Assessment of NF-κB–Luc activity in 3T3-L1 cells transfected either with siRNA for control, Ikka , Ikkb , or NF-κB p65 ( p65 ) or with pcDNA3.1/His- Ikba ( n = 5). h , i Representative images and realative quantification of ORO-stained images ( h ) and expression levels of Pparg and Cebpa ( i ) in 3T3-L1 cells infected with control adenovirus (Ad-C) or adenoviral Redd1 (Ad- R ) after transfection with vector alone or pcDNA3.1/His- Ikba ( n = 4). j NF-κB–Luc activity in mouse peritoneal macrophages infected with Ad-C or Ad- Redd1 ( n = 4). k Cytokine production in mouse peritoneal macrophages infected with Ad-C or Ad- Redd1 ( n = 5). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using one-way ANOVA ( g , h ) and two-way ANOVA ( a , i ) followed by the Holm–Sidak post hoc test and an unpaired two-tailed t -test ( b – f , j , k ). Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Staining, Transfection, Cell Culture, Expressing, Activity Assay, Control, Infection, Plasmid Preparation, Two Tailed Test

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a Predictive binding conformation between REDD1 and IκBα using computational protein-protein molecular docking methods. b Co-immunoprecipitation analysis of the interaction between REDD1 and IκBα in HEK293 cells transfected with pcDNA3.1/His- Ikba (His- Ikba ) and either pFlag-CMV-1- Redd1 ( Redd1 ) or Redd1 mutants ( R KKAA and R KKRAAA ) ( n = 3). c Representative confocal images of NF-κB p65 nuclear translocalization in HEK293 cells infected with Ad-C, Ad- Redd1 , or its mutants ( n = 4). Scale bar, 50 μm. d Assessment of NF-κB–Luc activity in 3T3-L1 cells infected with Ad-control, Ad- Redd1 , or its mutants ( n = 4). e Representative ORO-stained images of 3T3-L1 cells infected with Ad-control, Ad- Redd1 , or Ad- Redd1 KKAA and quantification of relative ORO intensity ( n = 4). f Expression levels of Pparg and Cebpa in 3T3-L1 cells infected with Ad-control, Ad- Redd1 , or Ad- Redd1 KKAA ( n = 4). g Production of MCP-1 and TNF-α in macrophages infected with Ad-control, Ad- Redd1 , or Ad- Redd1 KKAA ( n = 4). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using one-way ANOVA ( d , e ) and two-way ANOVA ( f , g ) followed by the Holm–Sidak post hoc test. Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Binding Assay, Immunoprecipitation, Transfection, Infection, Activity Assay, Control, Staining, Expressing

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a Weight gain in WT and Redd1 KKAA mice after being fed HFD for 16 weeks ( n = 10 per group). b eWAT and iWAT mass measurements in HFD-fed Redd1 KKAA mice and their WT littermates ( n = 10 per group). c Expression levels of Pparg and Cebpa in the eWAT of Redd1 KKAA mice and WT littermates fed HFD for 10 weeks ( n = 8 per group). d Representative images showing perilipin (green) and F4/80 (purple) staining in the eWAT of HFD-fed Redd1 KKAA mice and WT littermates ( n = 5 per group). Scale bar, 100 μm. e NF-κB activity in the eWAT from HFD-fed Redd1 KKAA mice and WT littermates ( n = 6 per group). f , g Plasma levels of inflammatory cytokines in HFD-fed Redd1 KKAA mice and WT littermates ( n = 8 per group). h Fasting plasma levels of glucose and insulin in HFD-fed Redd1 KKAA mice and WT littermates ( n = 8 per group). i Assessment of GTT and ITT in HFD-fed Redd1 KKAA mice and WT littermates after fasting for 12 and 6 h, respectively ( n = 6 per group). j , k Representative western blots of insulin-responsive Akt phosphorylation and plasma membrane-associated GLUT4 (PM-GLUT4) levels in the eWAT and skeletal muscle ( j ) and Akt and FOXO1 phosphorylation in the liver ( k ) of HFD-fed Redd1 KKAA mice and WT littermates ( n = 3). l Relative expression levels of G6pc , Pck1 , and Fbp1 in the liver of HFD-fed Redd1 KKAA mice and WT littermates compared with NC-fed mice ( n = 6 per group). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using an unpaired two-tailed t -test. Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Expressing, Staining, Activity Assay, Clinical Proteomics, Western Blot, Phospho-proteomics, Membrane, Two Tailed Test

Journal: Nature Communications

Article Title: REDD1 promotes obesity-induced metabolic dysfunction via atypical NF-κB activation

doi: 10.1038/s41467-022-34110-1

Figure Lengend Snippet: a Representative images of H&E-stained liver tissues from HFD-fed Redd1 −/− , Redd1 ΔAdipoq , Redd1 Δ LysM , Redd1 KKAA , and control mice, and quantification of hepatic steatosis from H&E-stained liver tissues ( n = 6 per group). Scale bars, 100 μm. b Expression levels of Acc , Fasn , and Scd-1 in the liver of HFD-fed Redd1 −/− , Redd1 ΔAdipoq , Redd1 Δ LysM , Redd1 KKAA , and their control mice ( n = 6 per group). Bar graphs represent mean ± s.e.m. Statistical significance was calculated using an unpaired two-tailed t -test. Source data are provided as a Source Data file.

Article Snippet: In addition, mouse adipose SVF cells and 3T3-L1 preadipocytes (5.0 × 10 5 cells/well, American Type Culture Collection) were plated and transfected with sh Redd1 (#sc-45807-SH, Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Staining, Control, Expressing, Two Tailed Test