Journal: Communications Biology

Article Title: Irisin regulates the phosphorylation of glucocorticoid receptor Ser212 and Ser234 and mediates glucocorticoid-induced muscle atrophy in mice

doi: 10.1038/s42003-025-09203-4

Figure Lengend Snippet: A Western blot analyses of P-GR(Ser234), P-GR(Ser212), P-GR(Ser143) and GAPDH in Dex and irisin-administered myotubes. B , C Protein levels were analyzed, n = 4 independent replicates. D Western blot analyses of P-GR(Ser234), P-GR(Ser212), GR and GAPDH in lentiviral constructs of C2C12 cell lines expressing wild-type GR (WT), serine-to-alanine phosphorylation-deficient (S234A, S212A, S234/212A) mutants, or serine-to-aspartate persistently phosphorylated (S234D, S212D, S234/212D) mutants. E The mRNA expressions of Trim63 and Fbxo32 were analyzed by qRT-PCR in Dex treatment lentiviral constructs of overexpression C2C12 cell lines, n = 4 independent replicates. F The mRNA expressions of Fbxo32 were analyzed by qRT-PCR in Dex and irisin treatment lentiviral constructs of overexpression GR S234D, S212D, S234/212D C2C12 cell lines, n = 4 independent replicates. G The mRNA expressions of Trim63 were analyzed by qRT-PCR in Dex and irisin treatment lentiviral constructs of overexpression GR S234A, S212A, S234/212A C2C12 cell lines, n = 4 independent replicates. Data are presented as means ± SEM. One-way ANOVA was used for statistical analysis ( B , C ), two-way ANOVA was used for statistical analysis ( E – G ).

Article Snippet: NC membranes were incubated with the following primary antibodies overnight at 4 °C: anti-irisin (1:1500, Phoenix Biotech, H-067-17), anti-Atrogin-1 (1:1000, abcam, ab168372), anti-MuRF-1 (1:1000, abcam, ab183094), anti-p-AKT(Ser473) (1:1000, CST, 9271S), anti-AKT (1:1000, CST, 9272S), anti-GAPDH(1:1000, ZSGBBIO, TA-08), anti-GR (1:1000, CST, 12041S), anti-Histone H3 (1:1000, CST, 3638S), anti-MSTN (1:250, abcam, ab98337), anti-p-Smad3 (1:1000, CST, 9520S), anti-Smad3 (1:1000, CST, 9523S), anti-IGF-1(1:1000, BOSTER, BA0939), anti-p-GR Ser234 (1:1000, orthologous to serine 226 in human, CST, 97285S), anti-p-GR Ser212 (1:1000, orthologous to serine 203 in human, Thermo Fisher, PA5-104446), anti-p-GR Ser134 (1:1000, orthologous to serine 143 in human, CST, 85060S), anti-p-JNK(Thr183/Tyr185) (1:1000, CST, 9251S), anti-p44/42 MAPK (Erk1/2) (1:1000, CST, 4695T), anti-integrin αV(1:1000, abcam, ab179475), anti-integrin β5(1:1000, CST, 3629S).

Techniques: Western Blot, Construct, Expressing, Phospho-proteomics, Quantitative RT-PCR, Over Expression

Journal: Communications Biology

Article Title: Irisin regulates the phosphorylation of glucocorticoid receptor Ser212 and Ser234 and mediates glucocorticoid-induced muscle atrophy in mice

doi: 10.1038/s42003-025-09203-4

Figure Lengend Snippet: A Western blot analyses of P-JNK (Thr183/Tyr185), P-ERK and GAPDH in Dex and irisin-administered myotubes. B Protein levels were analyzed, n = 4 independent replicates. C , D Western blot analyses of P-GR(Ser234) and P-GR(Ser212) in myotubes with JNK inhibitors and combined JNK and ERK inhibitors. E – H The mRNA expressions of Fbxo32, Trim63, IGF-1 and GAPDH were analyzed by qRT-PCR in myotubes treated with JNK inhibitors and ERK inhibitors, alone or in combination, n = 4 independent replicates. Data are presented as means ± SEM. One-way ANOVA was used for statistical analysis ( B ), two-way ANOVA was used for statistical analysis ( E – H ).

Article Snippet: NC membranes were incubated with the following primary antibodies overnight at 4 °C: anti-irisin (1:1500, Phoenix Biotech, H-067-17), anti-Atrogin-1 (1:1000, abcam, ab168372), anti-MuRF-1 (1:1000, abcam, ab183094), anti-p-AKT(Ser473) (1:1000, CST, 9271S), anti-AKT (1:1000, CST, 9272S), anti-GAPDH(1:1000, ZSGBBIO, TA-08), anti-GR (1:1000, CST, 12041S), anti-Histone H3 (1:1000, CST, 3638S), anti-MSTN (1:250, abcam, ab98337), anti-p-Smad3 (1:1000, CST, 9520S), anti-Smad3 (1:1000, CST, 9523S), anti-IGF-1(1:1000, BOSTER, BA0939), anti-p-GR Ser234 (1:1000, orthologous to serine 226 in human, CST, 97285S), anti-p-GR Ser212 (1:1000, orthologous to serine 203 in human, Thermo Fisher, PA5-104446), anti-p-GR Ser134 (1:1000, orthologous to serine 143 in human, CST, 85060S), anti-p-JNK(Thr183/Tyr185) (1:1000, CST, 9251S), anti-p44/42 MAPK (Erk1/2) (1:1000, CST, 4695T), anti-integrin αV(1:1000, abcam, ab179475), anti-integrin β5(1:1000, CST, 3629S).

Techniques: Western Blot, Quantitative RT-PCR

Journal: Communications Biology

Article Title: Irisin regulates the phosphorylation of glucocorticoid receptor Ser212 and Ser234 and mediates glucocorticoid-induced muscle atrophy in mice

doi: 10.1038/s42003-025-09203-4

Figure Lengend Snippet: A Western blot analyses of integrin αV, integrin β5 and GAPDH in Dex and irisin-administered myotubes. B Protein levels were analyzed, n = 4 independent replicates. C Western blot analyses of P-ERK, P-GR(Ser212), P-JNK (Thr183/Tyr185), P-GR(Ser234), IGF-1, MSTN and GAPDH in integrin αVβ5 inhibitor cliengitide treatment myotube. D – I Protein levels were analyzed, n = 4 independent replicates. Data are presented as means ± SEM. One-way ANOVA was used for statistical analysis ( B ), Two-way ANOVA was used for statistical analysis ( D – I ).

Article Snippet: NC membranes were incubated with the following primary antibodies overnight at 4 °C: anti-irisin (1:1500, Phoenix Biotech, H-067-17), anti-Atrogin-1 (1:1000, abcam, ab168372), anti-MuRF-1 (1:1000, abcam, ab183094), anti-p-AKT(Ser473) (1:1000, CST, 9271S), anti-AKT (1:1000, CST, 9272S), anti-GAPDH(1:1000, ZSGBBIO, TA-08), anti-GR (1:1000, CST, 12041S), anti-Histone H3 (1:1000, CST, 3638S), anti-MSTN (1:250, abcam, ab98337), anti-p-Smad3 (1:1000, CST, 9520S), anti-Smad3 (1:1000, CST, 9523S), anti-IGF-1(1:1000, BOSTER, BA0939), anti-p-GR Ser234 (1:1000, orthologous to serine 226 in human, CST, 97285S), anti-p-GR Ser212 (1:1000, orthologous to serine 203 in human, Thermo Fisher, PA5-104446), anti-p-GR Ser134 (1:1000, orthologous to serine 143 in human, CST, 85060S), anti-p-JNK(Thr183/Tyr185) (1:1000, CST, 9251S), anti-p44/42 MAPK (Erk1/2) (1:1000, CST, 4695T), anti-integrin αV(1:1000, abcam, ab179475), anti-integrin β5(1:1000, CST, 3629S).

Techniques: Western Blot

Journal: Scientific Reports

Article Title: Anxiolytic effects of Enterococcus faecalis 2001 on a mouse model of colitis

doi: 10.1038/s41598-024-62309-3

Figure Lengend Snippet: Effects of EF-2001 on serum LPS and corticosterone, and p-GR in the brain. Effects of chronic treatment with Enterococcus faecalis 2001 (EF-2001) on increased serum LPS ( a ) and corticosterone ( b ) concentration, and enhanced p-GR in the prefrontal cortex (PFC) ( c ) and ventral hippocampus ( d ) of DSS-treated mice. Quantification of the normalized values of p-GR with t-GR. Original immunoblot images were shown in Supplementary Fig. . ( e ) Microscopy images of p-GR (Ser211) (green), DAPI (blue), and NeuN, GFAP or Iba1 (red) immunostaining in the PFC of DSS-treated mice. Bars represent means ± standard error of mean (SEM). *p < 0.05 and **p < 0.01 vs. water-treated water group, # p < 0.05 and ## p < 0.01 vs. water-treated DSS group (n = 6–8 per group).

Article Snippet: After washing three times every 5 min, the sections were incubated with PBS containing 1% normal goat serum (Life Technologies Corporation, Carlsbad, CA, USA) or 3% bovine serum albumin and 0.3% Triton X-100 (PBSGT or PBSBT) at room temperature (23 ± 1 °C) for 2 h. The sections were incubated overnight at 4 °C with rabbit anti-p-GR (Ser211) (1:200; Cell Signaling Technology, #4161), rabbit anti-p-CAMKII (1:500; Cell Signaling Technology, #3361), rabbit anti-p-CREB (1:200; Cell Signaling Technology, #9198), mouse anti-NeuN (1:200; Millipore, MAB377), mouse anti-glial fibrillary acidic protein (GFAP; 1:200; Millipore, MAB360), or goat anti-ionized calcium-binding adaptor molecule 1 (Iba1; 1:200, Abcam, ab5076).

Techniques: Concentration Assay, Western Blot, Microscopy, Immunostaining