ct26 colon cancer cells  (ATCC)

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: Ellagic Acid Alleviates Abnormal Fat Reduction by Activating the RXRβ–PPARγ Pathways in a CT26 Tumour‐Induced Cachexia Mouse Model

doi: 10.1002/jcsm.70176

Figure Lengend Snippet: EA restores reduced adipocyte function in CM‐stimulated white adipocytes. (A) The experimental scheme for the preparation of the CT26 CM is shown. (B) IL‐6 levels were measured in CT26 CM with ELISA kits ( n = 3). (C) Intracellular lipid droplets were stained with Oil Red O (magnification 400×, scale bar 75 = μm). (D) The quantification of intracellular lipid was detected at 500 nm in 3T3‐L1 cells differentiated into white adipocytes, and treated with 50% of CT26 CM or EA (3.1, 6.3 and 12.5 μM) ( n = 3). (E) The levels of intracellular and extracellular free fatty acids were measured in 3T3‐L1 cells differentiated into white adipocytes, and treated with 50% of CT26 CM or EA (6.3 and 12.5 μM) ( n = 3). (F, G) The expression of adipokines was analysed using a Mouse Adipokine Proteome Array kit. (H) The protein expression of IGFBP‐3 and lipocalin‐2 was analysed with ImageJ ( n = 2). All data are expressed as the mean ± SEM. Statistical significance was determined using a one‐way ANOVA with Tukey's post hoc test for multigroup comparisons. * p < 0.05, ** p < 0.01, *** p < 0.001 or **** p < 0.0001 were considered statistically significant. CM, conditioned medium. DM (Wh), differentiation medium. EA, ellagic acid.

Article Snippet: Briefly, all mice were first randomized by body weight and divided into a non‐tumour‐bearing vehicle group, which received a subcutaneous injection of PBS, and a tumour‐induction group, which was injected with 5 × 10 5 CT26 colon cancer cells (CRL‐2638, ATCC, Rockville, MD, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Staining, Expressing

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: Ellagic Acid Alleviates Abnormal Fat Reduction by Activating the RXRβ–PPARγ Pathways in a CT26 Tumour‐Induced Cachexia Mouse Model

doi: 10.1002/jcsm.70176

Figure Lengend Snippet: EA‐mediated RXRβ‐PPARγ axis activation mitigates the reduction of adipogenesis by CT26 CM. (A) Overview of the crystal structure of the complex between RXRB (PDB ID: 7A78) and EA, and expected intermolecular interactions are shown. (B) Relative mRNA expression of Rxrb was measured by RT‐PCR in 3T3‐L1 differentiated into white adipocytes with or without si Rxrb , and data are normalized to Gapdh ( n = 3). (C) Representative morphological images of 3T3‐L1 treated with siRxrb and/or EA are shown (magnification 400×, scale bar = 75 μm). (D) The lipid droplets were stained with BODIPY‐Green (magnification 1000×, scale bar = 25 μm). Lipid droplet sizes and area were measured using the ImageJ software ( n = 3). (E) Protein expression of PPARγ and ACC was measured by Western blot analysis. (F) Intensities of the protein bands were measured with ImageJ and normalized to β‐actin ( n = 6). All data are expressed as the mean ± SEM. Statistical significance was determined using a one‐way ANOVA with Tukey's post hoc test for multigroup comparisons and the non‐parametric Mann–Whitney U test for two‐group comparisons. * p < 0.05, ** p < 0.01 or *** p < 0.001 were considered statistically significant. CM, conditioned medium. DM (Wh), differentiation medium. EA, ellagic acid.

Article Snippet: Briefly, all mice were first randomized by body weight and divided into a non‐tumour‐bearing vehicle group, which received a subcutaneous injection of PBS, and a tumour‐induction group, which was injected with 5 × 10 5 CT26 colon cancer cells (CRL‐2638, ATCC, Rockville, MD, USA).

Techniques: Activation Assay, Expressing, Reverse Transcription Polymerase Chain Reaction, Staining, Software, Western Blot, MANN-WHITNEY

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: Ellagic Acid Alleviates Abnormal Fat Reduction by Activating the RXRβ–PPARγ Pathways in a CT26 Tumour‐Induced Cachexia Mouse Model

doi: 10.1002/jcsm.70176

Figure Lengend Snippet: EA protects against body weight loss in CT26 tumour‐induced cachectic mice. (A) The experimental scheme of the in vivo study is shown. BALB/c mice were subcutaneously inoculated with 5 × 10 5 CT26 cells (CT26 group), except for the vehicle group. EA administration (10 mg/kg) via oral gavage was started 1 week after tumour cell injection (CT26 + EA group). 0.9% Normal saline (vehicle group and CT26 group) or EA (CT26 + EA group) was fed five times per week for 2 weeks. (B) The tumour‐free weight was calculated by subtracting the isolated tumour weight from the body weight ( n = 4). (C) The combined bilateral weight of the iWAT is shown ( n = 4). (D) The H&E‐stained image of the iWAT (magnification 400×, scale bar = 75 μm) is shown, and lipid droplet sizes were calculated using the ImageJ software. (E) The protein levels of C/EBPα, PPARγ, and pACC and ACC were analysed by Western blot analysis. Signal intensities of the protein bands were measured with ImageJ and normalized to β‐actin ( n = 4). (F) The paraffin‐embedded iWAT was stained with SREBP1 (green) and DAPI (blue) (magnification 1000×, scale bar = 25 μm), and representative images are shown. Fluorescence intensity of SREBP1 was quantified using the ImageJ software ( n = 4). Data are expressed as the mean ± SEM. Statistical significance was determined using a one‐way ANOVA with Tukey's post hoc test for multigroup comparisons. * p < 0.05, ** p < 0.01 or *** p < 0.001 were considered statistically significant. EA, ellagic acid. eWAT, epididymal white adipose tissue. iWAT, inguinal white adipose tissue.

Article Snippet: Briefly, all mice were first randomized by body weight and divided into a non‐tumour‐bearing vehicle group, which received a subcutaneous injection of PBS, and a tumour‐induction group, which was injected with 5 × 10 5 CT26 colon cancer cells (CRL‐2638, ATCC, Rockville, MD, USA).

Techniques: In Vivo, Injection, Saline, Isolation, Staining, Software, Western Blot, Fluorescence

Journal: Journal of Cachexia, Sarcopenia and Muscle

Article Title: Ellagic Acid Alleviates Abnormal Fat Reduction by Activating the RXRβ–PPARγ Pathways in a CT26 Tumour‐Induced Cachexia Mouse Model

doi: 10.1002/jcsm.70176

Figure Lengend Snippet: EA increases the expression of RXRβ in the iWAT of the CT26 cachexia model. (A) The tumour‐free weight was measured ( n = 7), and the percentage of fat in the total body was measured with DEXA analysis ( n = 3). (B) The combined bilateral weight of the iWAT and eWAT is shown ( n = 7). (C) The combined bilateral weight of TA was measured, and grip strength was measured ( n = 6–7). (D) The paraffin‐embedded iWAT was stained with RXRβ (green) and DAPI (blue) (magnification 1000×, scale bar = 25 μm), and representative images are shown. The bottom panels show zoomed views of the boxed areas in the top panels. (E) Index of correlation (IC) between RXRβ and nuclear (DAPI) was measured with the Colocalization Colormap plugin using ImageJ ( n = 4). (F) Schematic of the experimental models and the mechanism of action for EA. Data are expressed as the mean ± SEM. Statistical significance was determined using a one‐way ANOVA with Tukey's post hoc test for multigroup comparisons. * p < 0.05, ** p < 0.01 or *** p < 0.001 were considered statistically significant. EA, ellagic acid. eWAT, epididymal white adipose tissue. iWAT, inguinal white adipose tissue.

Article Snippet: Briefly, all mice were first randomized by body weight and divided into a non‐tumour‐bearing vehicle group, which received a subcutaneous injection of PBS, and a tumour‐induction group, which was injected with 5 × 10 5 CT26 colon cancer cells (CRL‐2638, ATCC, Rockville, MD, USA).

Techniques: Expressing, Staining