Journal: Frontiers in Nutrition
Article Title: Coptis japonica Makino ethanol extracts attenuates cancer cachexia induced muscle and fat wasting through inhibition of the STAT3 signaling pathway
doi: 10.3389/fnut.2025.1509086
Figure Lengend Snippet: The effects of CJME on CT26 conditioned medium (CM) induced-myotube atrophy. The C2C12 myoblasts were differentiated for 2 days and induced to form myotubes. (A) Effect of CJME on differentiated myotubes viability were determined using WST-1 assay. (B) Indicated CJME concentrations were used for pretreatment 1 h prior to treatment with CT26 CM for 48 h in differentiated myotubes. mRNA levels of (B) Atrogin-1 and (C) MuRF1 were determined using real-time PCR. Relative mRNA expression levels were normalized to those of GAPDH. (D) Protein levels of MyHC, Atrogin-1 and MuRF1 were analyzed via immunoblotting in cells cultured as described in B. (E) Differentiated myotubes were cultured as described in B, fixed with 4% PAF and stained with Crystal violet. Representative images of stained myotubes were shown (left) . The scale bar represents 200 μm. Myotube thickness was measured using Image J 1.53 software (right) (F) Differentiated myotubes were pretreated with 20 μg/mL of CJME for 1 h and exposed to CT26 CM for various times (0, 3, 6, 12, 24, and 48 h). Then, phosphorylation of STAT1, STAT2, STAT3, ERK, JNK, p38, IKKα/β and IκBα was examined using immunoblotting (left) . Quantified data for STAT3 were measured using Image J 1.53 software (right) (G) Differentiated myotubes were cultured as described in B, secreted levels of IL-6 in the culture medium were measured using ELISA. Data are representative of either two independent experiments. Data are presented in terms of the mean ± standard deviation and analyzed using a one-wayANOVA or Student's t -test. P value of <0.05 (*), <0.01 (**), <0.001 (***), or <0.0001 (****) were considered statistically significant.
Article Snippet: Antibodies targeting p-STAT1, p-STAT2, p-STAT3, p-ERK, p-JNK, p-p38, p-IKKα/β, and p-IκBα were sourced from Cell Signaling Technology (CST, Danvers, MA, USA).
Techniques: WST-1 Assay, Real-time Polymerase Chain Reaction, Expressing, Western Blot, Cell Culture, Staining, Software, Phospho-proteomics, Enzyme-linked Immunosorbent Assay, Standard Deviation