Journal: Cancer letters

Article Title: Acetyl-CoA carboxylase rewires cancer metabolism to allow cancer cells to survive inhibition of the Warburg effect by cetuximab

doi: 10.1016/j.canlet.2016.09.020

Figure Lengend Snippet: ACC plays a critical role in maintaining survival of cells exhibiting an increased dependence on glucose in low glucose culture. A–C, HEK293 cells infected with a recombinant lentivirus as indicated were subjected to Western blotting using the antibodies indicated (A). Conditioned media from culture of the cells in normal glucose medium (5 mM) for the indicated time periods were used for comparing glucose consumption (B). Cell survival proportions after culture in low glucose medium (1 mM) were determined as described in Materials and Methods (C). D–F, HEK293 cells co-infected with lentivirus containing HIF-1α_P402A/P564A or not and lentivirus containing ACC1_S79A or ACC2_S212A as indicated were subjected to Western blotting using the antibodies indicated (D) and to analysis of cell survival proportions (E, F). G–I, HEK293 cells infected with lentivirus containing HIF-1α_P402A/P564A or not and transfected with a control siRNA, ACC1 siRNA, or ACC2 siRNA as described in Materials and Methods were subjected to Western blotting using the antibodies indicated (G) and to analysis of cell survival proportions (H, I). All differences between control and experimental conditions are statistically significant except those marked “ns” (not significant).

Article Snippet: The primary antibodies used for Western blotting and their sources were as follows: HIF-1α and SREBP-1c, BD Biosciences; T172-phosphorylated AMPK, AMPK, S79-phosphorylated ACC1, ACC1, ACC2, and FASN, Cell Signaling Technology; and β-actin, Sigma-Aldrich.

Techniques: Infection, Recombinant, Western Blot, Transfection

Journal: Cancer letters

Article Title: Acetyl-CoA carboxylase rewires cancer metabolism to allow cancer cells to survive inhibition of the Warburg effect by cetuximab

doi: 10.1016/j.canlet.2016.09.020

Figure Lengend Snippet: HNSCC cells with low sensitivity to cetuximab contain a high basal level of ACC associated with increased stability of ACC. A and B, Cetuximab-sensitive (HN5) and cetuximab-resistant (HN5-R, UMSCC1, and MDA1986) cells were treated with cetuximab (20 nM) or not for 24 h. Cell protein lysates were subjected to Western blotting using the antibodies indicated (left panels). RNA samples were used to determine the levels of ACC1 and ACC2 mRNA in the cells with and without cetuximab treatment by real-time PCR as described in Materials and Methods (right panels). C, HN5 and HN5-R cells were cultured in the presence of the protein biosynthesis inhibitor cycloheximide (CHX, 10 μM) for the indicated times. Cell protein lysates were collected at each indicated time point and subjected to Western blotting using the antibodies indicated. Densitometric values of the ACC protein band at each time point were plotted as shown.

Article Snippet: The primary antibodies used for Western blotting and their sources were as follows: HIF-1α and SREBP-1c, BD Biosciences; T172-phosphorylated AMPK, AMPK, S79-phosphorylated ACC1, ACC1, ACC2, and FASN, Cell Signaling Technology; and β-actin, Sigma-Aldrich.

Techniques: Western Blot, Real-time Polymerase Chain Reaction, Cell Culture

Journal: Cancer letters

Article Title: Acetyl-CoA carboxylase rewires cancer metabolism to allow cancer cells to survive inhibition of the Warburg effect by cetuximab

doi: 10.1016/j.canlet.2016.09.020

Figure Lengend Snippet: Knockdown of ACC sensitized cetuximab-resistant cells to cetuximab treatment. A and B, HN5 (A) and HN5-R (B) cells infected with lentivirus containing ACC1_S79A or ACC2_S212A as indicated were subjected to MTT assays following treatment with cetuximab (20 nM) or not for 72 h and to Western blotting using the antibodies indicated following treatment with cetuximab (20 nM) or not for 24 h. C and D, HN5 cells (C) and HN5-R cells (D) were transfected with a control siRNA, one of two different ACC1 siRNAs, or one of two different ACC2 siRNAs as indicated for 48 h and then treated with cetuximab (20 nM) or not for an additional 24 h. Cell lysates were then subjected to Western blotting using the antibodies indicated and to analysis of apoptosis measured by fold increases in DNA fragmentation as described in Materials and Methods. * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.0005. ns: not significant.

Article Snippet: The primary antibodies used for Western blotting and their sources were as follows: HIF-1α and SREBP-1c, BD Biosciences; T172-phosphorylated AMPK, AMPK, S79-phosphorylated ACC1, ACC1, ACC2, and FASN, Cell Signaling Technology; and β-actin, Sigma-Aldrich.

Techniques: Infection, Western Blot, Transfection

Journal: Diabetes & metabolism journal

Article Title: Effects Of Exercise Training And Chlorogenic Acid Supplementation On Hepatic Lipid Metabolism In Prediabetes Mice.

doi: 10.4093/dmj.2022.0265

Figure Lengend Snippet: Fig. 3. Treatments attenuated high-fat diet (HFD)-induced increases in triglyceride production in the liver. At the first stage of the study, mice were treated with two different diets for the first 12 weeks: a normal diet (ND, n=7) and HFD (n=42). Then, in the second stage, HFD mice were further divided into six groups (n=7/group): no treatment (pre-D), treated with green coffee (GC), chlorogenic acid (CGA), exercise training (EX), GC+EX, and CGA+EX. Treatments were applied for 10 weeks under the same diet. At the end of the second stage (22nd week), (A) sterol regulatory element-binding protein-1c (Srebp-1c), (B) fatty acid syn- thase (Fasn), (C) acetyl-CoA carboxylase 1 (Acc1), and (E) glycerol-3-phosphate acyltransferase 1 (Gpat1) gene expression levels in the liver were measured using quantitative reverse transcription polymerase chain reaction. (D) ACC1 and (F) GPAT1 protein expression levels in the liver were assessed by immunoblotting. 18s and β-actin were used to normalize gene and protein expres- sion, respectively. aP<0.05 vs. ND, bP<0.01 vs. ND, cP<0.05 vs. pre-D, dP<0.01 vs. pre-D.

Article Snippet: The primary antibodies used were as follows: acyl-CoA synthetase 3 (ACSL3; 1:1,000, sc-166374), acetyl-CoA carboxylase 1 (ACC1; 1:1,000, sc-137104), acetyl-CoA carboxylase 2 (ACC2; 1:1,000, sc-390344), glycerol-3-phosphate acyltransferase 1 (GPAT1; 1:1,000, sc-398135), carnitine palmitoyl transferase 1 (CPT1; 1:1,000, sc-514555), and β-actin (1:1,000, sc-47778) (Santa Cruz Biotechnology, Dallas, TX, USA).

Techniques: Binding Assay, Gene Expression, Reverse Transcription, Polymerase Chain Reaction, Expressing, Western Blot

Journal: Diabetes & metabolism journal

Article Title: Effects Of Exercise Training And Chlorogenic Acid Supplementation On Hepatic Lipid Metabolism In Prediabetes Mice.

doi: 10.4093/dmj.2022.0265

Figure Lengend Snippet: Fig. 4. Treatments mitigated high-fat diet (HFD)-induced decreases in β-oxidation in the liver. At the first stage of the study, mice were treated with two different diets for the first 12 weeks: a normal diet (ND, n=7) and HFD (n=42). Then, in the second stage, HFD mice were further divided into six groups (n=7/group): no treatment (pre-D), treated with green coffee (GC), chlorogenic acid (CGA), exercise training (EX), GC+EX, and CGA+EX. Treatments were applied for 10 weeks under the same diet. At the end of the second stage (22nd week), (A) peroxisome proliferator-activated receptor alpha (Ppar-α), (B) acetyl-CoA carboxylase 2 (Acc2), and (D) carnitine palmitoyl transferase I (Cpt1) gene expression levels in the liver were measured using quantitative re- verse transcription polymerase chain reaction. (C) ACC2 and (E) CPT1 protein expression levels in the liver were assessed by im- munoblotting. 18s and β-actin were used to normalize gene and protein expression, respectively. aP<0.05 vs. ND, bP<0.01 vs. ND, cP<0.05 vs. pre-D, dP<0.01 vs. pre-D.

Article Snippet: The primary antibodies used were as follows: acyl-CoA synthetase 3 (ACSL3; 1:1,000, sc-166374), acetyl-CoA carboxylase 1 (ACC1; 1:1,000, sc-137104), acetyl-CoA carboxylase 2 (ACC2; 1:1,000, sc-390344), glycerol-3-phosphate acyltransferase 1 (GPAT1; 1:1,000, sc-398135), carnitine palmitoyl transferase 1 (CPT1; 1:1,000, sc-514555), and β-actin (1:1,000, sc-47778) (Santa Cruz Biotechnology, Dallas, TX, USA).

Techniques: Gene Expression, Polymerase Chain Reaction, Expressing

Journal: Diabetes & metabolism journal

Article Title: Effects Of Exercise Training And Chlorogenic Acid Supplementation On Hepatic Lipid Metabolism In Prediabetes Mice.

doi: 10.4093/dmj.2022.0265

Figure Lengend Snippet: Fig. 5. Graphical abstract. A prediabetic hepatic cell representing de novo lipogenesis (DNL) (left) associated with a high-fat diet (HFD) and the preventive effects of treatments (right). HFD elevates plasma free fatty acids (FFAs) which enter hepatic cells and contribute to the activation of acyl-CoA synthetase (ACSL) known to convert fatty acids (FAs) into fatty acyl-CoAs. Acyl-CoAs then proceed with de novo lipogenesis (acetyl-CoA carboxylase 1 [ACC1]) and/or β-oxidation inhibition (ACC2). Consequently, with the involvement of fatty acid synthase (Fasn) and glycerol-3-phosphate acyltransferase 1 (GPAT1), triglyceride (TG) can be formed. In this study, lipogenic molecules were found to increase with HFD (red color) which can lead to an increase in lipid drop- lets inside the cell owing to increased TG production (left). However, exercise training (EX), green coffee (GC), and chlorogenic acid (CGA) treatments mitigated HFD-induced changes in the lipogenic molecules (green color). Thus, β-oxidation via carnitine palmitoyl transferase I (CPT1) can be increased, and FA levels and the lipid droplet size inside the hepatic cell can be decreased.

Article Snippet: The primary antibodies used were as follows: acyl-CoA synthetase 3 (ACSL3; 1:1,000, sc-166374), acetyl-CoA carboxylase 1 (ACC1; 1:1,000, sc-137104), acetyl-CoA carboxylase 2 (ACC2; 1:1,000, sc-390344), glycerol-3-phosphate acyltransferase 1 (GPAT1; 1:1,000, sc-398135), carnitine palmitoyl transferase 1 (CPT1; 1:1,000, sc-514555), and β-actin (1:1,000, sc-47778) (Santa Cruz Biotechnology, Dallas, TX, USA).

Techniques: Clinical Proteomics, Activation Assay, Inhibition

Journal: Heliyon

Article Title: miR-122/PPARβ axis is involved in hypoxic exercise and modulates fatty acid metabolism in skeletal muscle of obese rats

doi: 10.1016/j.heliyon.2024.e26572

Figure Lengend Snippet: Primers used for RT-qPCR analysis.

Article Snippet: The protein samples were then transferred to a PVDF membrane (0.45-μm pore size) at a constant pressure of 100 V for 10 min, followed by blocked overnight in Tris-buffered saline with 0.1% Tween-20 (TBST), which contained 5% bovine serum albumin, and then probed overnight at 4 °C with primary antibodies against PPARβ (1:2000; GTX113250) and FAS (1:1000; GTX13550) (both from GeneTex, Irvine, CA, USA) and ACC2 (1:500; A03668-2) and CPT1b (1:500; PB9491) (both from Boster Bio, Pleasanton, CA, USA).

Techniques: Amplification

Journal: Heliyon

Article Title: miR-122/PPARβ axis is involved in hypoxic exercise and modulates fatty acid metabolism in skeletal muscle of obese rats

doi: 10.1016/j.heliyon.2024.e26572

Figure Lengend Snippet: Expression of the lipid metabolism regulator PPARβ and downstream effectors (CPT1b, FAS, and ACC2). mRNA levels in obese rats under hypoxic conditions, with hypoxic training, and with regulated miR-122 expression were determined using qRT-PCR. OE, obese rats with miR-122 overexpression and hypoxic training; IE, obese rats with miR-122 depletion and hypoxic training; CE, obese rats with hypoxic training only; H, obese sedentary rats without regulation of miR-122 expression. Data are presented as mean ± SD. *p < 0.05, **p < 0.01 vs rats in group H; # p < 0.05, ## p < 0.01 vs rats in group CE; & p < 0.05, && p < 0.01 vs rats in group OE (1-way analysis of variance).

Article Snippet: The protein samples were then transferred to a PVDF membrane (0.45-μm pore size) at a constant pressure of 100 V for 10 min, followed by blocked overnight in Tris-buffered saline with 0.1% Tween-20 (TBST), which contained 5% bovine serum albumin, and then probed overnight at 4 °C with primary antibodies against PPARβ (1:2000; GTX113250) and FAS (1:1000; GTX13550) (both from GeneTex, Irvine, CA, USA) and ACC2 (1:500; A03668-2) and CPT1b (1:500; PB9491) (both from Boster Bio, Pleasanton, CA, USA).

Techniques: Expressing, Quantitative RT-PCR, Over Expression

Journal: Heliyon

Article Title: miR-122/PPARβ axis is involved in hypoxic exercise and modulates fatty acid metabolism in skeletal muscle of obese rats

doi: 10.1016/j.heliyon.2024.e26572

Figure Lengend Snippet: Expression of the lipid metabolism regulator PPARβ and downstream effectors (CPT1b, FAS, and ACC2). Protein levels in obese rats under hypoxic conditions, with hypoxic training, and with regulated miR-122 expression were determined using Western blot analysis. OE, obese rats with miR-122 overexpression and hypoxic training; IE, obese rats with miR-122 depletion and hypoxic training; CE, obese rats with hypoxic training only; H, obese sedentary rats without regulation of miR-122 expression. Data are presented as mean ± SD. *p < 0.05, **p < 0.01 vs rats in group H; # p < 0.05, ## p < 0.01 vs rats in group CE; & p < 0.05, && p < 0.01 vs rats in group OE (1-way analysis of variance).

Article Snippet: The protein samples were then transferred to a PVDF membrane (0.45-μm pore size) at a constant pressure of 100 V for 10 min, followed by blocked overnight in Tris-buffered saline with 0.1% Tween-20 (TBST), which contained 5% bovine serum albumin, and then probed overnight at 4 °C with primary antibodies against PPARβ (1:2000; GTX113250) and FAS (1:1000; GTX13550) (both from GeneTex, Irvine, CA, USA) and ACC2 (1:500; A03668-2) and CPT1b (1:500; PB9491) (both from Boster Bio, Pleasanton, CA, USA).

Techniques: Expressing, Western Blot, Over Expression