gw7647 cat Search Results


503 gw7647 targetmol cat  (TargetMol)
94
TargetMol 503 gw7647 targetmol cat
503 Gw7647 Targetmol Cat, supplied by TargetMol, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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503 gw7647 targetmol cat - by Bioz Stars, 2026-03
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gw7647  (Tocris)
93
Tocris gw7647
Gw7647, supplied by Tocris, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 1 article reviews
gw7647 - by Bioz Stars, 2026-03
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gw7647  (Cayman Chemical)
90
Cayman Chemical gw7647
Autophagy inhibition impairs pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in mice. Fed or 24 h fasted Atg7 F/F , and Atg7 LKO mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or <t>GW7647,</t> a synthetic PPAR𝛼 agonist (10 mg/kg BW) twice a day. A total of 5 h after last treatment, all mice were killed to collect livers, of which total RNAs were isolated for qPCR analysis. ( b ) Hepatic expression levels of PPAR𝛼 target genes Acox1 , Acot3 , Fgf21 , Pdk4 , Acot1, and Cidec were determined in Atg7 F/F and Atg7 LKO mice shown in panel ( a ) by qPCR analysis. n = 4 per group, ** p < 0.01 vs. Atg7 F/F mice treated with vehicle. ## p < 0.01. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual mouse. Statistics by two-tailed t -test. ( c ) A schematic diagram of an experimental procedure in AML12 cells, a mouse hepatocyte-derived cell line. AML12 cells were treated with bafilomycin A1 (BafA1: 0.1, 0.5, or 1 μM), a known autophagy inhibitor in the absence or presence of synthetic PPAR𝛼 agonists (1 μM of GW764 or 100 μM of Wy-14,643) for 24 h. Vehicle is 0.1% DMSO. Total RNAs from these cells were prepared to perform qPCR analysis. ( d , e ) Expression levels of PPAR𝛼 target genes Pdk4 , Acot3 , and Ucp2, were determined in AML12 cells shown in panel ( c ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. ## p < 0.01 vs. AML12 cells treated with GW7647 or Wy-14,643. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual sample. Statistics by a two-tailed, unpaired Student t -test. BW, body weight.
Gw7647, supplied by Cayman Chemical, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/gw7647/product/Cayman Chemical
Average 90 stars, based on 1 article reviews
gw7647 - by Bioz Stars, 2026-03
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gw7647  (Santa Cruz Biotechnology)
90
Santa Cruz Biotechnology gw7647
ZFP961 interacts with PPAR α to block thermogenic activation. a) Co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961 and Flag‐PPAR α . b) HEK293T cells were co‐transfected with indicated plasmids, and the transcriptional activity of PPAR α was measured with luciferase reporter assay ( n = 4, per group). c) Left panel: co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961‐WT, HA‐ZFP961‐Mut, or Flag‐PPAR α in the presence or absence of <t>GW7647</t> (1 × 10 −6 m ). Right panel: ZFP961‐Mut and ZFP961 mRNA levels were analyzed by qRT‐PCR ( n = 4, per group). d) The transcriptional activity of PPAR α was measured in HEK293T cells that were co‐transfected with indicated plasmids and treated with or without GW7647 (1 × 10 −6 m ) ( n = 4, per group). e) Mature brown adipocytes were infected with adenovirus expressing ZFP961‐WT, ZFP961‐Mut, or GFP and then treated with or without GW7647 (1 × 10 −6 m ) for 24 h. Gene expression was analyzed ( n = 4, per group). f) Ucp1 protein in mature brown adipocytes generated as in (e). g) Brown preadipocytes infected with lentivirus containing Scr or PPAR α shRNAs were differentiated and then infected with adenovirus expressing ZFP961‐WT or GFP, and gene expression was analyzed ( n = 4, per group). h) Ucp1 protein in mature brown adipocytes generated as in (g). All error bars represent s.e.m. Two‐tailed unpaired Student's t ‐test was performed. * p < 0.05; n.s., not significant.
Gw7647, supplied by Santa Cruz Biotechnology, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/gw7647/product/Santa Cruz Biotechnology
Average 90 stars, based on 1 article reviews
gw7647 - by Bioz Stars, 2026-03
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lanthascreen tm competitive binding assay kits  (Thermo Fisher)
90
Thermo Fisher lanthascreen tm competitive binding assay kits
ZFP961 interacts with PPAR α to block thermogenic activation. a) Co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961 and Flag‐PPAR α . b) HEK293T cells were co‐transfected with indicated plasmids, and the transcriptional activity of PPAR α was measured with luciferase reporter assay ( n = 4, per group). c) Left panel: co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961‐WT, HA‐ZFP961‐Mut, or Flag‐PPAR α in the presence or absence of <t>GW7647</t> (1 × 10 −6 m ). Right panel: ZFP961‐Mut and ZFP961 mRNA levels were analyzed by qRT‐PCR ( n = 4, per group). d) The transcriptional activity of PPAR α was measured in HEK293T cells that were co‐transfected with indicated plasmids and treated with or without GW7647 (1 × 10 −6 m ) ( n = 4, per group). e) Mature brown adipocytes were infected with adenovirus expressing ZFP961‐WT, ZFP961‐Mut, or GFP and then treated with or without GW7647 (1 × 10 −6 m ) for 24 h. Gene expression was analyzed ( n = 4, per group). f) Ucp1 protein in mature brown adipocytes generated as in (e). g) Brown preadipocytes infected with lentivirus containing Scr or PPAR α shRNAs were differentiated and then infected with adenovirus expressing ZFP961‐WT or GFP, and gene expression was analyzed ( n = 4, per group). h) Ucp1 protein in mature brown adipocytes generated as in (g). All error bars represent s.e.m. Two‐tailed unpaired Student's t ‐test was performed. * p < 0.05; n.s., not significant.
Lanthascreen Tm Competitive Binding Assay Kits, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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rosiglitazone cat. no. 5325/10  (Tocris)
90
Tocris rosiglitazone cat. no. 5325/10
ZFP961 interacts with PPAR α to block thermogenic activation. a) Co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961 and Flag‐PPAR α . b) HEK293T cells were co‐transfected with indicated plasmids, and the transcriptional activity of PPAR α was measured with luciferase reporter assay ( n = 4, per group). c) Left panel: co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961‐WT, HA‐ZFP961‐Mut, or Flag‐PPAR α in the presence or absence of <t>GW7647</t> (1 × 10 −6 m ). Right panel: ZFP961‐Mut and ZFP961 mRNA levels were analyzed by qRT‐PCR ( n = 4, per group). d) The transcriptional activity of PPAR α was measured in HEK293T cells that were co‐transfected with indicated plasmids and treated with or without GW7647 (1 × 10 −6 m ) ( n = 4, per group). e) Mature brown adipocytes were infected with adenovirus expressing ZFP961‐WT, ZFP961‐Mut, or GFP and then treated with or without GW7647 (1 × 10 −6 m ) for 24 h. Gene expression was analyzed ( n = 4, per group). f) Ucp1 protein in mature brown adipocytes generated as in (e). g) Brown preadipocytes infected with lentivirus containing Scr or PPAR α shRNAs were differentiated and then infected with adenovirus expressing ZFP961‐WT or GFP, and gene expression was analyzed ( n = 4, per group). h) Ucp1 protein in mature brown adipocytes generated as in (g). All error bars represent s.e.m. Two‐tailed unpaired Student's t ‐test was performed. * p < 0.05; n.s., not significant.
Rosiglitazone Cat. No. 5325/10, supplied by Tocris, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/rosiglitazone cat. no. 5325/10/product/Tocris
Average 90 stars, based on 1 article reviews
rosiglitazone cat. no. 5325/10 - by Bioz Stars, 2026-03
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ppar-α antagonist gw6471  (Tocris)
90
Tocris ppar-α antagonist gw6471
ZFP961 interacts with PPAR α to block thermogenic activation. a) Co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961 and Flag‐PPAR α . b) HEK293T cells were co‐transfected with indicated plasmids, and the transcriptional activity of PPAR α was measured with luciferase reporter assay ( n = 4, per group). c) Left panel: co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961‐WT, HA‐ZFP961‐Mut, or Flag‐PPAR α in the presence or absence of <t>GW7647</t> (1 × 10 −6 m ). Right panel: ZFP961‐Mut and ZFP961 mRNA levels were analyzed by qRT‐PCR ( n = 4, per group). d) The transcriptional activity of PPAR α was measured in HEK293T cells that were co‐transfected with indicated plasmids and treated with or without GW7647 (1 × 10 −6 m ) ( n = 4, per group). e) Mature brown adipocytes were infected with adenovirus expressing ZFP961‐WT, ZFP961‐Mut, or GFP and then treated with or without GW7647 (1 × 10 −6 m ) for 24 h. Gene expression was analyzed ( n = 4, per group). f) Ucp1 protein in mature brown adipocytes generated as in (e). g) Brown preadipocytes infected with lentivirus containing Scr or PPAR α shRNAs were differentiated and then infected with adenovirus expressing ZFP961‐WT or GFP, and gene expression was analyzed ( n = 4, per group). h) Ucp1 protein in mature brown adipocytes generated as in (g). All error bars represent s.e.m. Two‐tailed unpaired Student's t ‐test was performed. * p < 0.05; n.s., not significant.
Ppar α Antagonist Gw6471, supplied by Tocris, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/ppar-α antagonist gw6471/product/Tocris
Average 90 stars, based on 1 article reviews
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Image Search Results


Autophagy inhibition impairs pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in mice. Fed or 24 h fasted Atg7 F/F , and Atg7 LKO mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647, a synthetic PPAR𝛼 agonist (10 mg/kg BW) twice a day. A total of 5 h after last treatment, all mice were killed to collect livers, of which total RNAs were isolated for qPCR analysis. ( b ) Hepatic expression levels of PPAR𝛼 target genes Acox1 , Acot3 , Fgf21 , Pdk4 , Acot1, and Cidec were determined in Atg7 F/F and Atg7 LKO mice shown in panel ( a ) by qPCR analysis. n = 4 per group, ** p < 0.01 vs. Atg7 F/F mice treated with vehicle. ## p < 0.01. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual mouse. Statistics by two-tailed t -test. ( c ) A schematic diagram of an experimental procedure in AML12 cells, a mouse hepatocyte-derived cell line. AML12 cells were treated with bafilomycin A1 (BafA1: 0.1, 0.5, or 1 μM), a known autophagy inhibitor in the absence or presence of synthetic PPAR𝛼 agonists (1 μM of GW764 or 100 μM of Wy-14,643) for 24 h. Vehicle is 0.1% DMSO. Total RNAs from these cells were prepared to perform qPCR analysis. ( d , e ) Expression levels of PPAR𝛼 target genes Pdk4 , Acot3 , and Ucp2, were determined in AML12 cells shown in panel ( c ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. ## p < 0.01 vs. AML12 cells treated with GW7647 or Wy-14,643. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual sample. Statistics by a two-tailed, unpaired Student t -test. BW, body weight.

Journal: Cells

Article Title: Basal Autophagy Is Necessary for A Pharmacologic PPARα Transactivation

doi: 10.3390/cells11040754

Figure Lengend Snippet: Autophagy inhibition impairs pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in mice. Fed or 24 h fasted Atg7 F/F , and Atg7 LKO mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647, a synthetic PPAR𝛼 agonist (10 mg/kg BW) twice a day. A total of 5 h after last treatment, all mice were killed to collect livers, of which total RNAs were isolated for qPCR analysis. ( b ) Hepatic expression levels of PPAR𝛼 target genes Acox1 , Acot3 , Fgf21 , Pdk4 , Acot1, and Cidec were determined in Atg7 F/F and Atg7 LKO mice shown in panel ( a ) by qPCR analysis. n = 4 per group, ** p < 0.01 vs. Atg7 F/F mice treated with vehicle. ## p < 0.01. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual mouse. Statistics by two-tailed t -test. ( c ) A schematic diagram of an experimental procedure in AML12 cells, a mouse hepatocyte-derived cell line. AML12 cells were treated with bafilomycin A1 (BafA1: 0.1, 0.5, or 1 μM), a known autophagy inhibitor in the absence or presence of synthetic PPAR𝛼 agonists (1 μM of GW764 or 100 μM of Wy-14,643) for 24 h. Vehicle is 0.1% DMSO. Total RNAs from these cells were prepared to perform qPCR analysis. ( d , e ) Expression levels of PPAR𝛼 target genes Pdk4 , Acot3 , and Ucp2, were determined in AML12 cells shown in panel ( c ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. ## p < 0.01 vs. AML12 cells treated with GW7647 or Wy-14,643. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual sample. Statistics by a two-tailed, unpaired Student t -test. BW, body weight.

Article Snippet: AML12 cells were purchased from ATCC (Manassas, VA, USA) (CRL-2254); GW7647 (Cat. #10008613), Wy-14,643 (Cat. #70703), and L-sulforaphane (SFN, Cat. #14797) from Cayman; dimethylfumarate (DMF, Cat. #242926), butylated hydroxyanisole (BHA, Cat. #B1253-500G), dexamethasone (Cat. #D4902), polyethylene glycol 400 (PEG 400, Cat. #P3265-1KG), 3-methyladenine (Cat. #M9281) and Tween 80 (Cat. #P1754-500ML) from Sigma-Aldrich (St. Louis, MO, USA); Bafilomycin A1 (BafA1, Cat. #BML-CM110) from Enzo Life Science (Farmingdale, NY, USA); HyClone DMEM/F12 (Cat. #SH30023.01) and fetal bovine serum (FBS, Cat. #SH20084.03) from HyClone (Logan, UT, USA); Trizol Reagent (Cat. #15596018) from Invitrogen (Waltham, MA, USA); RbTaq TM qPCR 2X PreMIX (SYBR Green with high ROX, Cat. #RT531M) from Enzynomics (Daejeon, Korea); PrimeScript TM 1st strand cDNA Synthesis kit (Cat. #6110A) from TaKaRa Bio Inc. (Kusatsu, Shiga, Japan); cOmplete TM , Mini, EDTA-free Protease Inhibitor Cocktail (Cat. #11836170001), PhosSTOP TM (Cat. #04906837001), and Insulin-Transferrin-Sodium Selenite Supplement (ITS, Cat. #11074547001) from Roche (Basel, Switzerland); penicillin-streptomycin (Cat. #15140122), Pierce TM BCA protein assay kit (Cat. #23225) and Pierce ECL Western Blotting Substrate (Cat. #32106) from Thermo Scientific (Waltham, MA, USA); BSA (Cat. #0332-100G) from VWR (Radnor, PA, USA); Immun-Blot PVDF Membrane (Cat. #1620177) from Bio-Rad Laboratories (Hercules, CA, USA); Rabbit monoclonal anti-β-actin (13E5) antibody (Cat. #5125, 1:3000 dilution), Rabbit monoclonal anti-Atg7 (D12B11) antibody (Cat. #8558S, 1:1000 dilution), Rabbit SQSTM1/p62 antibody (Cat. #5114S, 1:1000 dilution) and anti-rabbit IgG HRP-linked antibody (Cat. #7074S, 1:3000 dilution) from Cell Signaling Technology (Danvers, MA, USA); Dimethyl sulfoxide (DMSO, Cat. #sc-358801) from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA); Atg5 −/− (RCB 2711) and Atg7 −/− (RCB 3707) MEFs from Riken BRC.

Techniques: Inhibition, Injection, Isolation, Expressing, Two Tailed Test, Derivative Assay

SFN-mediated NRF2 activation in a dose-dependent manner impairs pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in AML12 cells. AML12 cells were treated with sulforaphane (SFN, 12.5, 25, or 50 μM), a known NRF2 activator in a dose-dependent manner in the absence or presence of synthetic PPAR𝛼 agonists (GW7647, 0.01, 0.1 or 1 μM; Wy-14,643, 1, 10 or 100 μM) for 24 h. Vehicle is 0.1% DMSO. Total RNAs from these cells were prepared to perform qPCR analysis. ( b , c ) Expression levels of NRF2 target genes Nqo1 and Hmox1 were determined in AML12 cells shown in panel ( a ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual well. ( d , e ) Expression levels of PPAR𝛼 target genes Pdk4 , Acot3 , and Ucp2, were determined in AML12 cells shown in panel ( a ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. ## p < 0.01 vs. AML12 cells treated with 1 μM GW7647 or 100 μM Wy-14,643. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual sample. Statistics by a two-tailed, unpaired Student t -test.

Journal: Cells

Article Title: Basal Autophagy Is Necessary for A Pharmacologic PPARα Transactivation

doi: 10.3390/cells11040754

Figure Lengend Snippet: SFN-mediated NRF2 activation in a dose-dependent manner impairs pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in AML12 cells. AML12 cells were treated with sulforaphane (SFN, 12.5, 25, or 50 μM), a known NRF2 activator in a dose-dependent manner in the absence or presence of synthetic PPAR𝛼 agonists (GW7647, 0.01, 0.1 or 1 μM; Wy-14,643, 1, 10 or 100 μM) for 24 h. Vehicle is 0.1% DMSO. Total RNAs from these cells were prepared to perform qPCR analysis. ( b , c ) Expression levels of NRF2 target genes Nqo1 and Hmox1 were determined in AML12 cells shown in panel ( a ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual well. ( d , e ) Expression levels of PPAR𝛼 target genes Pdk4 , Acot3 , and Ucp2, were determined in AML12 cells shown in panel ( a ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. ## p < 0.01 vs. AML12 cells treated with 1 μM GW7647 or 100 μM Wy-14,643. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual sample. Statistics by a two-tailed, unpaired Student t -test.

Article Snippet: AML12 cells were purchased from ATCC (Manassas, VA, USA) (CRL-2254); GW7647 (Cat. #10008613), Wy-14,643 (Cat. #70703), and L-sulforaphane (SFN, Cat. #14797) from Cayman; dimethylfumarate (DMF, Cat. #242926), butylated hydroxyanisole (BHA, Cat. #B1253-500G), dexamethasone (Cat. #D4902), polyethylene glycol 400 (PEG 400, Cat. #P3265-1KG), 3-methyladenine (Cat. #M9281) and Tween 80 (Cat. #P1754-500ML) from Sigma-Aldrich (St. Louis, MO, USA); Bafilomycin A1 (BafA1, Cat. #BML-CM110) from Enzo Life Science (Farmingdale, NY, USA); HyClone DMEM/F12 (Cat. #SH30023.01) and fetal bovine serum (FBS, Cat. #SH20084.03) from HyClone (Logan, UT, USA); Trizol Reagent (Cat. #15596018) from Invitrogen (Waltham, MA, USA); RbTaq TM qPCR 2X PreMIX (SYBR Green with high ROX, Cat. #RT531M) from Enzynomics (Daejeon, Korea); PrimeScript TM 1st strand cDNA Synthesis kit (Cat. #6110A) from TaKaRa Bio Inc. (Kusatsu, Shiga, Japan); cOmplete TM , Mini, EDTA-free Protease Inhibitor Cocktail (Cat. #11836170001), PhosSTOP TM (Cat. #04906837001), and Insulin-Transferrin-Sodium Selenite Supplement (ITS, Cat. #11074547001) from Roche (Basel, Switzerland); penicillin-streptomycin (Cat. #15140122), Pierce TM BCA protein assay kit (Cat. #23225) and Pierce ECL Western Blotting Substrate (Cat. #32106) from Thermo Scientific (Waltham, MA, USA); BSA (Cat. #0332-100G) from VWR (Radnor, PA, USA); Immun-Blot PVDF Membrane (Cat. #1620177) from Bio-Rad Laboratories (Hercules, CA, USA); Rabbit monoclonal anti-β-actin (13E5) antibody (Cat. #5125, 1:3000 dilution), Rabbit monoclonal anti-Atg7 (D12B11) antibody (Cat. #8558S, 1:1000 dilution), Rabbit SQSTM1/p62 antibody (Cat. #5114S, 1:1000 dilution) and anti-rabbit IgG HRP-linked antibody (Cat. #7074S, 1:3000 dilution) from Cell Signaling Technology (Danvers, MA, USA); Dimethyl sulfoxide (DMSO, Cat. #sc-358801) from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA); Atg5 −/− (RCB 2711) and Atg7 −/− (RCB 3707) MEFs from Riken BRC.

Techniques: Activation Assay, Expressing, Two Tailed Test

SFN-mediated NRF2 activation in a time-dependent manner impairs pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in AML12 cells. AML12 cells were treated with 25 μM of SFN, a known NRF2 activator in a time-dependent manner (6, 12, or 18 h) in the absence or presence of synthetic PPAR𝛼 agonists (GW7647, 0.01, 0.1 or 1 μM; Wy-14,643, 1, 10 or 100 μM) for 24 h. Vehicle is 0.1% DMSO. Total RNAs from these cells were prepared to perform qPCR analysis. ( b , c ) Expression levels of NRF2 target genes Nqo1 and Hmox1 were determined in AML12 cells shown in panel ( a ) by qPCR analysis. ( d ) Expression levels of PPAR𝛼 target genes Pdk4 , Acot3 , and Ucp2, were determined in AML12 cells shown in panel ( a ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. ## p < 0.01 vs. AML12 cells treated with GW7647 or Wy-14,643. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual sample. Statistics by a two-tailed, unpaired Student t -test.

Journal: Cells

Article Title: Basal Autophagy Is Necessary for A Pharmacologic PPARα Transactivation

doi: 10.3390/cells11040754

Figure Lengend Snippet: SFN-mediated NRF2 activation in a time-dependent manner impairs pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in AML12 cells. AML12 cells were treated with 25 μM of SFN, a known NRF2 activator in a time-dependent manner (6, 12, or 18 h) in the absence or presence of synthetic PPAR𝛼 agonists (GW7647, 0.01, 0.1 or 1 μM; Wy-14,643, 1, 10 or 100 μM) for 24 h. Vehicle is 0.1% DMSO. Total RNAs from these cells were prepared to perform qPCR analysis. ( b , c ) Expression levels of NRF2 target genes Nqo1 and Hmox1 were determined in AML12 cells shown in panel ( a ) by qPCR analysis. ( d ) Expression levels of PPAR𝛼 target genes Pdk4 , Acot3 , and Ucp2, were determined in AML12 cells shown in panel ( a ) by qPCR analysis. n = 3 per group, ** p < 0.01 vs. AML12 cells treated with vehicle. ## p < 0.01 vs. AML12 cells treated with GW7647 or Wy-14,643. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual sample. Statistics by a two-tailed, unpaired Student t -test.

Article Snippet: AML12 cells were purchased from ATCC (Manassas, VA, USA) (CRL-2254); GW7647 (Cat. #10008613), Wy-14,643 (Cat. #70703), and L-sulforaphane (SFN, Cat. #14797) from Cayman; dimethylfumarate (DMF, Cat. #242926), butylated hydroxyanisole (BHA, Cat. #B1253-500G), dexamethasone (Cat. #D4902), polyethylene glycol 400 (PEG 400, Cat. #P3265-1KG), 3-methyladenine (Cat. #M9281) and Tween 80 (Cat. #P1754-500ML) from Sigma-Aldrich (St. Louis, MO, USA); Bafilomycin A1 (BafA1, Cat. #BML-CM110) from Enzo Life Science (Farmingdale, NY, USA); HyClone DMEM/F12 (Cat. #SH30023.01) and fetal bovine serum (FBS, Cat. #SH20084.03) from HyClone (Logan, UT, USA); Trizol Reagent (Cat. #15596018) from Invitrogen (Waltham, MA, USA); RbTaq TM qPCR 2X PreMIX (SYBR Green with high ROX, Cat. #RT531M) from Enzynomics (Daejeon, Korea); PrimeScript TM 1st strand cDNA Synthesis kit (Cat. #6110A) from TaKaRa Bio Inc. (Kusatsu, Shiga, Japan); cOmplete TM , Mini, EDTA-free Protease Inhibitor Cocktail (Cat. #11836170001), PhosSTOP TM (Cat. #04906837001), and Insulin-Transferrin-Sodium Selenite Supplement (ITS, Cat. #11074547001) from Roche (Basel, Switzerland); penicillin-streptomycin (Cat. #15140122), Pierce TM BCA protein assay kit (Cat. #23225) and Pierce ECL Western Blotting Substrate (Cat. #32106) from Thermo Scientific (Waltham, MA, USA); BSA (Cat. #0332-100G) from VWR (Radnor, PA, USA); Immun-Blot PVDF Membrane (Cat. #1620177) from Bio-Rad Laboratories (Hercules, CA, USA); Rabbit monoclonal anti-β-actin (13E5) antibody (Cat. #5125, 1:3000 dilution), Rabbit monoclonal anti-Atg7 (D12B11) antibody (Cat. #8558S, 1:1000 dilution), Rabbit SQSTM1/p62 antibody (Cat. #5114S, 1:1000 dilution) and anti-rabbit IgG HRP-linked antibody (Cat. #7074S, 1:3000 dilution) from Cell Signaling Technology (Danvers, MA, USA); Dimethyl sulfoxide (DMSO, Cat. #sc-358801) from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA); Atg5 −/− (RCB 2711) and Atg7 −/− (RCB 3707) MEFs from Riken BRC.

Techniques: Activation Assay, Expressing, Two Tailed Test

NRF2 activation impairs pharmacologic PPAR𝛼 transactivation in vivo. ( a ) A schematic diagram of an experimental procedure in mice. The 8- to 9-week-old male C57BL/6J wild-type mice were orally gavaged with vehicle or butylated hydroxyanisole (BHA, 200 mg/kg BW) once a day for 3 days. During the last 24 h, all mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647, a synthetic PPAR𝛼 agonist (10 mg/kg BW) twice a day (first injection: 00:00 am, second injection: 12:00 pm). A total of 5 h after last treatment, all mice were killed to collect livers of which total RNAs were prepared for qPCR analysis. ( b ) Hepatic expression levels of PPAR𝛼 target genes Pdk4 , Upc2 , Acot3 , Acox1 , and Fgf21 were determined as shown in panel ( a ) by qPCR analysis. n = 8 per group, * p < 0.05, ** p < 0.01 vs. WT treated with vehicle. ( c ) A schematic diagram of an experimental procedure in mice. The 8- to 9-week-old male Alb-Cre/+ and Keap1 LKO mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647 (10 mg/kg BW) twice a day. A total of 5 h after last treatment, all mice were killed to collect livers, of which total RNAs were isolated for qPCR analysis. ( d ) Hepatic expression levels of Keap1 , NRF2 target genes ( Nqo1 and Gsta1 ), and PPAR𝛼 target genes Pdk4 , Upc2 , Cidec , Acot1 to 3 , and Acox1 were determined shown in panel ( c ) by qPCR analysis. n = 4 per group, ** p < 0.01 vs. Alb-Cre/+ treated with vehicle. # p < 0.05, ## p < 0.01. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual mouse. Statistics by a two-tailed, unpaired Student t -test. WT, wild-type.

Journal: Cells

Article Title: Basal Autophagy Is Necessary for A Pharmacologic PPARα Transactivation

doi: 10.3390/cells11040754

Figure Lengend Snippet: NRF2 activation impairs pharmacologic PPAR𝛼 transactivation in vivo. ( a ) A schematic diagram of an experimental procedure in mice. The 8- to 9-week-old male C57BL/6J wild-type mice were orally gavaged with vehicle or butylated hydroxyanisole (BHA, 200 mg/kg BW) once a day for 3 days. During the last 24 h, all mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647, a synthetic PPAR𝛼 agonist (10 mg/kg BW) twice a day (first injection: 00:00 am, second injection: 12:00 pm). A total of 5 h after last treatment, all mice were killed to collect livers of which total RNAs were prepared for qPCR analysis. ( b ) Hepatic expression levels of PPAR𝛼 target genes Pdk4 , Upc2 , Acot3 , Acox1 , and Fgf21 were determined as shown in panel ( a ) by qPCR analysis. n = 8 per group, * p < 0.05, ** p < 0.01 vs. WT treated with vehicle. ( c ) A schematic diagram of an experimental procedure in mice. The 8- to 9-week-old male Alb-Cre/+ and Keap1 LKO mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647 (10 mg/kg BW) twice a day. A total of 5 h after last treatment, all mice were killed to collect livers, of which total RNAs were isolated for qPCR analysis. ( d ) Hepatic expression levels of Keap1 , NRF2 target genes ( Nqo1 and Gsta1 ), and PPAR𝛼 target genes Pdk4 , Upc2 , Cidec , Acot1 to 3 , and Acox1 were determined shown in panel ( c ) by qPCR analysis. n = 4 per group, ** p < 0.01 vs. Alb-Cre/+ treated with vehicle. # p < 0.05, ## p < 0.01. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual mouse. Statistics by a two-tailed, unpaired Student t -test. WT, wild-type.

Article Snippet: AML12 cells were purchased from ATCC (Manassas, VA, USA) (CRL-2254); GW7647 (Cat. #10008613), Wy-14,643 (Cat. #70703), and L-sulforaphane (SFN, Cat. #14797) from Cayman; dimethylfumarate (DMF, Cat. #242926), butylated hydroxyanisole (BHA, Cat. #B1253-500G), dexamethasone (Cat. #D4902), polyethylene glycol 400 (PEG 400, Cat. #P3265-1KG), 3-methyladenine (Cat. #M9281) and Tween 80 (Cat. #P1754-500ML) from Sigma-Aldrich (St. Louis, MO, USA); Bafilomycin A1 (BafA1, Cat. #BML-CM110) from Enzo Life Science (Farmingdale, NY, USA); HyClone DMEM/F12 (Cat. #SH30023.01) and fetal bovine serum (FBS, Cat. #SH20084.03) from HyClone (Logan, UT, USA); Trizol Reagent (Cat. #15596018) from Invitrogen (Waltham, MA, USA); RbTaq TM qPCR 2X PreMIX (SYBR Green with high ROX, Cat. #RT531M) from Enzynomics (Daejeon, Korea); PrimeScript TM 1st strand cDNA Synthesis kit (Cat. #6110A) from TaKaRa Bio Inc. (Kusatsu, Shiga, Japan); cOmplete TM , Mini, EDTA-free Protease Inhibitor Cocktail (Cat. #11836170001), PhosSTOP TM (Cat. #04906837001), and Insulin-Transferrin-Sodium Selenite Supplement (ITS, Cat. #11074547001) from Roche (Basel, Switzerland); penicillin-streptomycin (Cat. #15140122), Pierce TM BCA protein assay kit (Cat. #23225) and Pierce ECL Western Blotting Substrate (Cat. #32106) from Thermo Scientific (Waltham, MA, USA); BSA (Cat. #0332-100G) from VWR (Radnor, PA, USA); Immun-Blot PVDF Membrane (Cat. #1620177) from Bio-Rad Laboratories (Hercules, CA, USA); Rabbit monoclonal anti-β-actin (13E5) antibody (Cat. #5125, 1:3000 dilution), Rabbit monoclonal anti-Atg7 (D12B11) antibody (Cat. #8558S, 1:1000 dilution), Rabbit SQSTM1/p62 antibody (Cat. #5114S, 1:1000 dilution) and anti-rabbit IgG HRP-linked antibody (Cat. #7074S, 1:3000 dilution) from Cell Signaling Technology (Danvers, MA, USA); Dimethyl sulfoxide (DMSO, Cat. #sc-358801) from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA); Atg5 −/− (RCB 2711) and Atg7 −/− (RCB 3707) MEFs from Riken BRC.

Techniques: Activation Assay, In Vivo, Injection, Expressing, Isolation, Two Tailed Test

NRF2 activation is necessary and sufficient for suppressing pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in mice. The 8- to 9-week-old male Nrf2 F/F and Nrf2 LKO mice were orally gavaged with vehicle or butylated hydroxyanisole (BHA, 200 mg/kg BW) once a day for 3 days. During the last 24 h, all mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647, a synthetic PPAR𝛼 agonist (10 mg/kg BW) twice a day (first injection: 00:00 am, second injection: 12:00 pm). A total of 5 h after last treatment, all mice were killed to collect livers of which total RNAs were prepared for qPCR analysis. ( b ) Hepatic expression levels of Nrf2 and its target genes Nqo1 , Gsta1 , and Hmox1 were determined in AML12 cells shown in panel ( a ) by qPCR analysis, * p < 0.05. ( c ) Hepatic expression levels of PPAR𝛼 target genes Pdk4 , Upc2 , Cidec , and Acot1 to 3 were determined as shown in panel ( a ) by qPCR analysis. n = 3–4 per group, ** p < 0.01 vs. Nrf2 F/F treated with vehicle. # p < 0.05, ## p < 0.01. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual mouse. Statistics by a two-tailed, unpaired Student t -test.

Journal: Cells

Article Title: Basal Autophagy Is Necessary for A Pharmacologic PPARα Transactivation

doi: 10.3390/cells11040754

Figure Lengend Snippet: NRF2 activation is necessary and sufficient for suppressing pharmacologic PPAR𝛼 transactivation. ( a ) A schematic diagram of an experimental procedure in mice. The 8- to 9-week-old male Nrf2 F/F and Nrf2 LKO mice were orally gavaged with vehicle or butylated hydroxyanisole (BHA, 200 mg/kg BW) once a day for 3 days. During the last 24 h, all mice were intraperitoneally injected with vehicle (0.1% DMSO in 4:1 ratio of PEG 400 and Tween 80) or GW7647, a synthetic PPAR𝛼 agonist (10 mg/kg BW) twice a day (first injection: 00:00 am, second injection: 12:00 pm). A total of 5 h after last treatment, all mice were killed to collect livers of which total RNAs were prepared for qPCR analysis. ( b ) Hepatic expression levels of Nrf2 and its target genes Nqo1 , Gsta1 , and Hmox1 were determined in AML12 cells shown in panel ( a ) by qPCR analysis, * p < 0.05. ( c ) Hepatic expression levels of PPAR𝛼 target genes Pdk4 , Upc2 , Cidec , and Acot1 to 3 were determined as shown in panel ( a ) by qPCR analysis. n = 3–4 per group, ** p < 0.01 vs. Nrf2 F/F treated with vehicle. # p < 0.05, ## p < 0.01. Data represent mean ± s.e.m. and are plotted as fold change. Each dot indicates an individual mouse. Statistics by a two-tailed, unpaired Student t -test.

Article Snippet: AML12 cells were purchased from ATCC (Manassas, VA, USA) (CRL-2254); GW7647 (Cat. #10008613), Wy-14,643 (Cat. #70703), and L-sulforaphane (SFN, Cat. #14797) from Cayman; dimethylfumarate (DMF, Cat. #242926), butylated hydroxyanisole (BHA, Cat. #B1253-500G), dexamethasone (Cat. #D4902), polyethylene glycol 400 (PEG 400, Cat. #P3265-1KG), 3-methyladenine (Cat. #M9281) and Tween 80 (Cat. #P1754-500ML) from Sigma-Aldrich (St. Louis, MO, USA); Bafilomycin A1 (BafA1, Cat. #BML-CM110) from Enzo Life Science (Farmingdale, NY, USA); HyClone DMEM/F12 (Cat. #SH30023.01) and fetal bovine serum (FBS, Cat. #SH20084.03) from HyClone (Logan, UT, USA); Trizol Reagent (Cat. #15596018) from Invitrogen (Waltham, MA, USA); RbTaq TM qPCR 2X PreMIX (SYBR Green with high ROX, Cat. #RT531M) from Enzynomics (Daejeon, Korea); PrimeScript TM 1st strand cDNA Synthesis kit (Cat. #6110A) from TaKaRa Bio Inc. (Kusatsu, Shiga, Japan); cOmplete TM , Mini, EDTA-free Protease Inhibitor Cocktail (Cat. #11836170001), PhosSTOP TM (Cat. #04906837001), and Insulin-Transferrin-Sodium Selenite Supplement (ITS, Cat. #11074547001) from Roche (Basel, Switzerland); penicillin-streptomycin (Cat. #15140122), Pierce TM BCA protein assay kit (Cat. #23225) and Pierce ECL Western Blotting Substrate (Cat. #32106) from Thermo Scientific (Waltham, MA, USA); BSA (Cat. #0332-100G) from VWR (Radnor, PA, USA); Immun-Blot PVDF Membrane (Cat. #1620177) from Bio-Rad Laboratories (Hercules, CA, USA); Rabbit monoclonal anti-β-actin (13E5) antibody (Cat. #5125, 1:3000 dilution), Rabbit monoclonal anti-Atg7 (D12B11) antibody (Cat. #8558S, 1:1000 dilution), Rabbit SQSTM1/p62 antibody (Cat. #5114S, 1:1000 dilution) and anti-rabbit IgG HRP-linked antibody (Cat. #7074S, 1:3000 dilution) from Cell Signaling Technology (Danvers, MA, USA); Dimethyl sulfoxide (DMSO, Cat. #sc-358801) from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA); Atg5 −/− (RCB 2711) and Atg7 −/− (RCB 3707) MEFs from Riken BRC.

Techniques: Activation Assay, Injection, Expressing, Two Tailed Test

ZFP961 interacts with PPAR α to block thermogenic activation. a) Co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961 and Flag‐PPAR α . b) HEK293T cells were co‐transfected with indicated plasmids, and the transcriptional activity of PPAR α was measured with luciferase reporter assay ( n = 4, per group). c) Left panel: co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961‐WT, HA‐ZFP961‐Mut, or Flag‐PPAR α in the presence or absence of GW7647 (1 × 10 −6 m ). Right panel: ZFP961‐Mut and ZFP961 mRNA levels were analyzed by qRT‐PCR ( n = 4, per group). d) The transcriptional activity of PPAR α was measured in HEK293T cells that were co‐transfected with indicated plasmids and treated with or without GW7647 (1 × 10 −6 m ) ( n = 4, per group). e) Mature brown adipocytes were infected with adenovirus expressing ZFP961‐WT, ZFP961‐Mut, or GFP and then treated with or without GW7647 (1 × 10 −6 m ) for 24 h. Gene expression was analyzed ( n = 4, per group). f) Ucp1 protein in mature brown adipocytes generated as in (e). g) Brown preadipocytes infected with lentivirus containing Scr or PPAR α shRNAs were differentiated and then infected with adenovirus expressing ZFP961‐WT or GFP, and gene expression was analyzed ( n = 4, per group). h) Ucp1 protein in mature brown adipocytes generated as in (g). All error bars represent s.e.m. Two‐tailed unpaired Student's t ‐test was performed. * p < 0.05; n.s., not significant.

Journal: Advanced Science

Article Title: The KRAB Domain‐Containing Protein ZFP961 Represses Adipose Thermogenesis and Energy Expenditure through Interaction with PPAR α

doi: 10.1002/advs.202102949

Figure Lengend Snippet: ZFP961 interacts with PPAR α to block thermogenic activation. a) Co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961 and Flag‐PPAR α . b) HEK293T cells were co‐transfected with indicated plasmids, and the transcriptional activity of PPAR α was measured with luciferase reporter assay ( n = 4, per group). c) Left panel: co‐immunoprecipitation analysis using cell extracts from HEK293T cells that were co‐transfected with HA‐ZFP961‐WT, HA‐ZFP961‐Mut, or Flag‐PPAR α in the presence or absence of GW7647 (1 × 10 −6 m ). Right panel: ZFP961‐Mut and ZFP961 mRNA levels were analyzed by qRT‐PCR ( n = 4, per group). d) The transcriptional activity of PPAR α was measured in HEK293T cells that were co‐transfected with indicated plasmids and treated with or without GW7647 (1 × 10 −6 m ) ( n = 4, per group). e) Mature brown adipocytes were infected with adenovirus expressing ZFP961‐WT, ZFP961‐Mut, or GFP and then treated with or without GW7647 (1 × 10 −6 m ) for 24 h. Gene expression was analyzed ( n = 4, per group). f) Ucp1 protein in mature brown adipocytes generated as in (e). g) Brown preadipocytes infected with lentivirus containing Scr or PPAR α shRNAs were differentiated and then infected with adenovirus expressing ZFP961‐WT or GFP, and gene expression was analyzed ( n = 4, per group). h) Ucp1 protein in mature brown adipocytes generated as in (g). All error bars represent s.e.m. Two‐tailed unpaired Student's t ‐test was performed. * p < 0.05; n.s., not significant.

Article Snippet: In some experiments, cells were treated with GW7647 (1 × 10 −6 m ) (Santa Cruz Biotechnology, cat. no. CAS 265129‐71‐3) as indicated.

Techniques: Blocking Assay, Activation Assay, Immunoprecipitation, Transfection, Activity Assay, Luciferase, Reporter Assay, Quantitative RT-PCR, Infection, Expressing, Gene Expression, Generated, Two Tailed Test