Journal: Sports Medicine and Health Science

Article Title: Exercise preconditioning prevents immobilization-induced skeletal muscle atrophy by activating Prmt1-p38/ATF2-Sesn1 signaling axis in C57BL/6J mice

doi: 10.1016/j.smhs.2025.04.001

Figure Lengend Snippet: Exercise preconditioning prevented muscle atrophy through protein arginine methyltransferase 1 (Prmt1)-Sestrin1 (Sesn1)-transcriptional co-activator PPAR-γ co-activator-1 α (PGC-1α)-mediated skeletal muscle regeneration. (A–B) Western blot results of MyoD, Myf5, MEF2 and MyoG in GAS muscle. (C–D) Co-IP results, IP: MyoD, GAS muscle was used. A-B, ∗ p ​< ​0.05 vs. C; ∗∗ p ​< ​0.01 vs. C; ## p ​< ​0.01 vs. Im; $ p ​< ​0.05 vs. E ​+ ​Im; $$ p ​< ​0.01 vs. E ​+ ​Im. Two-way ANOVA was used, and data are shown as means ​± ​standard error of the mean ( SEM ) (C, n ​= ​8, Im, n ​= ​8, E ​+ ​Im, n ​= ​8, E+5003+Im, n ​= ​8). (E–F) H&E staining of myotubes at each differentiation time points of C2C12 myoblasts overexpressing Sesn1. Scale bar ​= ​100 ​μm ∗ p ​< ​0.05 vs. Ad-Sesn1-, unpaired Student's t-test was used and data are shown as means ​± ​ SEM ( n ​= ​3 in each group). (G–H) Western blot results of Sesn1, Prmt1, PGC-1α, MyoD, MEF2, Myf5, and MyoG in C2C12 myoblasts at each time points of differentiation. ∗ p ​< ​0.05 vs. D0, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group). (I–J) Western blot results of PGC-1α in nucleus of C2C12 myoblasts overexpressing Sesn1 at each time points of differentiation. ∗∗ p ​< ​0.01 vs. Ad-Sesn1, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group). (K–L) Western blot results of PGC-1α in cytoplasm of C2C12 myoblasts overexpressing Sesn1 at each time points of differentiation. ∗∗ p ​< ​0.01 vs. Ad-Sesn1-, unpaired Student's t -test was used and data are shown as means ​± ​ SEM ( n ​= ​6 in each group).

Article Snippet: The antibodies are listed below: aDMA (Anti-Asymmetric Di-Methyl Arginine Motif) (1:1 500, Rabbit, Cell Signal Tech, USA), Akt (protein kinase B) (1:2 000, Mouse, Proteintech, USA), pAkt-Ser473 (1:2 000, Rabbit, Cell Signal Tech, USA), AMPKα2 (1:2 000, Rabbit, Cell Signal Tech, USA), pAMPK-Thr172 (1:2 000, Rabbit, Cell Signal Tech, USA), ATF2 (1:2 000, Rabbit, Proteintech, USA), Atrogin-1 (FBXO32) (1:15 000, Mouse, Proteintech, USA), FoxO3a (Forkhead box O3) (1:1 000, Mouse, Proteintech, USA), pFoxO3a-Ser315 (1:2 000, Rabbit, Proteintech, USA), GAPDH (1:5 000, Rabbit, Utibody, CN), IGF-1 (insulin-like growth factor 1) (1:2 000, Mouse, Proteintech, USA), LaminB (1:2 000, Rabbit, Abcam, USA), MEF2 (myocyte enhancer factor 2) (1:2 000, Rabbit, Proteintech, USA), Myf5 (myogenic factor 5) (1:2 000, Rabbit, Abclonal, CN), MyoD (myogenic differentiation antigen) (1:2 000, Rabbit, Proteintech, USA), MyoG (myogenin) (1:2 000, Rabbit, Proteintech, USA). mTOR (mammalian target of rapamycin) (1:2 000, Rabbit, Cell Signal Tech, USA), p38 (1:2 000, Rabbit, WANLEIBIO, CN), p-p38-Thr180/Tyr182 (1:2 000, Rabbit, WANLEIBIO, CN), PGC-1α (1:1 000, Rabbit, Abcam, USA), Prmt1 (1:2 000, Rabbit, Cell Signal Tech, USA), Raptor (1:1 000, Rabbit, Cell Signal Tech, USA), Sesn1 (1:1 000, Rabbit, HUABIO, CN), Sesn1 (1:1 000, Rabbit, Abcam, USA), TRIM63 (MuRF1) (1:2 000, Rabbit, Proteintech, USA).

Techniques: Western Blot, Co-Immunoprecipitation Assay, Staining

Journal: Journal of cachexia, sarcopenia and muscle

Article Title: Survival motor neuron deficiency slows myoblast fusion through reduced myomaker and myomixer expression.

doi: 10.1002/jcsm.12740

Figure Lengend Snippet: Figure 4 SMN deficiency alters the expression of MyoD and myogenin. (A) Schematic showing binding of MyoD and myogenin to E-box elements up- stream of the myomaker and myomixer promoters. MyoD (B) and myogenin (C) mRNA expression in wild-type and SMN-deficient C2C12 cells after 7 days of differentiation. Data were analysed using an unpaired t-test (MyoD: P < 0.0001; myogenin: P < 0.0001). N = 5. (D) Myomaker and myomixer mRNA expression 48 h after MyoD or myogenin overexpression. Data were analysed using a one-way ANOVA [myomaker: F(5, 23) = 5.358, P = 0.0021; myomixer: F(5, 23) = 3.667, P = 0.0138] followed by post hoc Tukey’s test for multiple comparisons (myomaker: WT vs. WT + MyoD: P > 0.9999; WT vs. WT + Myogenin: P = 0.9996; SMN KD vs. SMN KD + MyoD: P = 0.0236; SMN KD vs. SMN KD + Myogenin: P = 0.0434) (myomixer: WT vs. WT + MyoD: P = 0.9725; WT vs. WT + Myogenin: P = 0.8755; SMN KD vs. SMN KD + MyoD: P = 0.0251; SMN KD vs. SMN KD + Myogenin: P = 0.2222). N = 5. (E) MyoD or myogenin were overexpressed in wild-type and SMN-deficient C2C12 cells and differentiated for 5 days. Cells were fixed and stained for myosin and DAPI (20×; scale bar: 50 μM) and quantification of the fusion index and the per cent of total myotubes with 2–3, 4–7, and 8+ nuclei were determined to measure myotube maturity. Fusion index data were analysed using a one-way ANOVA [F(5, 24) = 28.86, P < 0.0001] followed by post hoc Tukey’s test for multiple comparisons (WT vs. WT + MyoD: P = 0.8238; WT vs. WT + Myogenin: P = 0.4409; SMA vs. SMA + MyoD: P = 0.0185; SMA vs. SMA + Myogenin: P = 0.0127). Myotube maturity data were analysed using a one-way ANOVA [(2–3 nuclei: F(3, 16) = 38.62, P < 0.0001; 4–7 nuclei: F(3, 16) = 25.65, P < 0.0001; 8+ nuclei: F(3, 16) = 36, P < 0.0001] followed by post hoc Tukey’s test for multiple comparisons (2–3 nuclei SMN KD vs. SMN KD + MyoD: P < 0.0001; 2–3 nuclei SMN KD vs. SMN KD + myogenin: P = 0.0007; 4–7 nuclei SMN KD vs. SMN KD + MyoD: P < 0.0001; 4–7 nuclei SMN KD vs. SMN KD + myogenin: P = 0.0017; 8+ nuclei SMN KD vs. SMN KD + MyoD: P > 0.9999; 8+ nuclei SMN KD vs. SMN KD + myogenin: P > 0.9999). N = 5. (F) Sequences of the mouse MyoD and myogenin promoters that were probed in ChIP experiments. The primers bound to the sequences underlined in red and E-boxes within these regions are outlined in a black rectangle. The PCR product from the MyoD promoter begins 104 bp upstream of the MyoD start sequence and is 92 bp long. The PCR product from the myogenin promoter begins 142 bp upstream of the myogenin start sequence and is 140 bp long. (G) Wild-type C2C12 cells were differentiated for 48 h and fixed. Chromatin co-immunoprecipitated with SMN protein was probed with primers to the MyoD and myogenin promoters. Data were analysed using an unpaired t-test (MyoD: P = 0.0031; Myogenin: P = 0.4158). n = 4. All data are represented as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Article Snippet: Wild-type and SMN-deficient C2C12 cells were plated and after 24 h, transfected with 1 μg MyoD (Cat. #14710, Addgene, Watertown, MA, USA) or myogenin (Cat. #MR227400, Origene Technologies Inc., Rockville, MD, USA) using Lipofectamine 3000 (Thermo Fisher).

Techniques: Expressing, Binding Assay, Over Expression, Staining, Sequencing, Immunoprecipitation

Journal: The Journal of Veterinary Medical Science

Article Title: Primary pharyngeal alveolar rhabdomyosarcoma in an adolescent Japanese black heifer

doi: 10.1292/jvms.20-0305

Figure Lengend Snippet: Primary antibodies, antigen retrieval conditions, and immunohistochemistry results

Article Snippet: Myogenin , F5D , Origene, Rockville, MD, USA , 1:200 , 121°C, 15 min, in CB , +/– , + , + , +.

Techniques: Immunohistochemistry

Journal: Journal of Biomedical Science

Article Title: Hypoxia and therapeutic treatment of EV-A71 with an immune modulator TLR7 agonist in a new immunocompetent mouse model

doi: 10.1186/s12929-019-0585-y

Figure Lengend Snippet: A “white-jade” muscle phenotype in EV-A71-infected mouse strain 129. a Left panel : We coined a “white-jade” phenotype to describe the pathological changes in the color of the hindlimb muscle in EV-A71-infected mice (indicated by arrow). This phenotype showed characteristic muscle with locally whitened color in appearance and hardened tissue mass. In the saline control, tendons (not muscle) can be seen in white color. Right panel : An anatomical sketch of the hindlimb muscle. GM: gluteal muscle, QF: quadratus femoris muscle, BF: Biceps femoris muscle. b Both VP1 protein (upper) and myogenin protein (lower) were detected by IHC staining in the muscles, indicating EV-A71 infection, muscle injury and regeneration. The magnification is 200X. c Striking expression of HIF1A (hypoxia inducible factor 1-α) protein (brown color) was detected by IHC staining in the white-jaded tissue, but not in the non-white-jaded tissue from the same mouse. d Specific association between HIF1A expression and white-jaded muscle

Article Snippet: Slides were washed with PBS containing 0.1% Tween 20 (PBST), followed by incubation with specific antibodies, including rabbit anti-VP1 polyclonal antibody (PB7631, Abnova, Taiwan), rabbit anti-HIF1A polyclonal antibody (NB100–479, Novus), rabbit anti-VEGFA polyclonal antibody (ab46154, Abcam), rabbit anti-myogenin monoclonal antibody (GTX 63352, Genetex, Taiwan), as well as antibodies specific for the immune system, CD3 (MA1–90582, Thermo), CD19 (PAB19567, Abnova), CD68 (ab955, Abcam), and CD163 (bs-2527, Bioss), respectively.

Techniques: Infection, Saline, Control, Immunohistochemistry, Muscles, Expressing

Journal: Journal of Biomedical Science

Article Title: Hypoxia and therapeutic treatment of EV-A71 with an immune modulator TLR7 agonist in a new immunocompetent mouse model

doi: 10.1186/s12929-019-0585-y

Figure Lengend Snippet: Hypoxia in EV-A71-infected muscle was rescued by timely treatment with IFNαA. a A schematic diagram of mouse IFNαA treatment after EV-A71 infection. Six different experimental groups of mice received mIFNαA (300 IU/g of mouse) at different dpi and frequencies of treatments. Therapeutic efficacies were assayed by survival rates. In some cases, infected and treated mice ( n = 12) were monitored for up to 1.5 years. b Clinical scores were significantly reduced only in Exp. I (green) and Exp. II (red), when treated at dpi 1, or dpi 1, 2, 3. c Survival rates of Exp. I (green) (100%) and Exp. II (red) (88%) were significantly higher than that of the control group with no IFNαA treatment. d No VP1-positive signal was detected in the muscle by IHC at 25 dpi and 41 dpi in mice recovered from IFNαA treatment in Exp. 1. e The myogenin protein (black arrowhead) was expressed in recovered muscle via IHC staining, indicating muscle regeneration after injury. f HIF1A and VEGFA proteins were both expressed in newly recovered muscle (dpi = 25), but not in the fully recovered muscle (dpi = 41)

Article Snippet: Slides were washed with PBS containing 0.1% Tween 20 (PBST), followed by incubation with specific antibodies, including rabbit anti-VP1 polyclonal antibody (PB7631, Abnova, Taiwan), rabbit anti-HIF1A polyclonal antibody (NB100–479, Novus), rabbit anti-VEGFA polyclonal antibody (ab46154, Abcam), rabbit anti-myogenin monoclonal antibody (GTX 63352, Genetex, Taiwan), as well as antibodies specific for the immune system, CD3 (MA1–90582, Thermo), CD19 (PAB19567, Abnova), CD68 (ab955, Abcam), and CD163 (bs-2527, Bioss), respectively.

Techniques: Infection, Control, Immunohistochemistry