Journal: Frontiers in Genetics

Article Title: RNA-seq analysis reveals a positive role for NGF in the myogenic differentiation of bovine skeletal muscle satellite cells

doi: 10.3389/fgene.2025.1713817

Figure Lengend Snippet: Role of NGF in the regulation of myogenic differentiation. (A) Determination of the mRNA levels of Pax7, a marker gene of proliferation. (B) Determination of the protein levels of Pax7. (C) Detection of the effect of the over-NGF on the proliferation of bSMSCs using the CCK8 method. (D) Detection of the effect of the si-NGF on the proliferation of bSMSCs using the CCK8 method. (E) Determination of the mRNA expression of MyHC, MyoG and MyoD1-myogenic differentiation marker genes. (F) Determination of the protein expression of MyHC, MyoG and MyoD1. ** p < 0.01; *** p < 0.001.

Article Snippet: Primary antibodies against NGF(Bioss, bs-10806R, Beijing, China, 1:2000 dilution), PAX7 (DSHB, Pax7, United States, 1:1500 dilution), MyoD1(Bioss, bs-62914R, Beijing, China, 1:1500 dilution), MyHC(DSHB,10F5, United States, 1:1500 dilution), MyoG (Bioss, bs-3550R, Beijing, China, 1:1500 dilution), Akt (CST, 9272S, United States, 1:2000 dilution), p-Akt (CST, 9271S, United States,1:2000 dilution), PI3K(CST, 4249S, United States, 1: 2000 dilution), p-PI3K(CST, 4228S, United States, 1: 2000 dilution)and GAPDH(Proteintech, 60004-1-Ig, United States, 1: 5000 dilution) were diluted with TBST, and the total protein concentrations of the samples were determined using a BCA assay.

Techniques: Cell Characterization, Marker, Expressing

Journal: Stem Cell Research & Therapy

Article Title: VEGF-B-mediated myofiber types involved in high-fat diet-induced hyperglycemia through PKA-NFATs signaling pathway

doi: 10.1186/s13287-025-04455-7

Figure Lengend Snippet: VEGF-B186 inhibits myoblast differentiation/fusion and slow-twitch myofiber formation through the PKA-NFAT-MyoG/MEF2C signaling pathway. ( A ) Heatmap showing the expression of NFAT signaling pathway genes in the gastrocnemius muscle tissue of WT and Vegfb −/− mice fed a ND or HFD. ( B ) Quantitative analysis of NFATc1 and NFATc2 expression in ( A ). n = 3. ** P < 0.01. ( C - E ) Western blot detection of Nfatc1 and Nfatc2 proteins in the tibialis anterior muscle of WT-ND, Vegfb −/− -ND, WT-HFD and Vegfb −/− -HFD mice. n = 3. * P < 0.05, ** P < 0.01. ( F - G ) Western blot detection of P-Nfatc1/c2 in the cytoplasm and NFATc1/c2 in the cell nucleus on the 6th day of differentiation in C2C12 myoblast cells treated with 100 ng/mL VEGF-B186 with or without N 6 -Bz-cAMP (10 − 5 mol/L) and 8-Bromo-cAMP (10 − 5 mol/L). n = 3. * P < 0.05, ** P < 0.01. (H) Western blot detection of MyHC and NOQ on the 6th day of differentiation in C2C12 cells treated with 100 ng/mL VEGF-B186 with or without AdNFATc1/c2. n = 3. * P < 0.05, ** P < 0.01. ( I ) Representative images of MyoG + and MEF2C + staining on the 6th day of differentiation in C2C12 myoblast cells treated with 100 ng/mL VEGF-B186 with or without AdNFATc1/c2. DAPI represents the nucleus. Bar = 100 µM

Article Snippet: Differentiated myoblasts were stained for MyoG, MEF2C, NOQ or My32 via the polyclonal primary antibodies MyoG (1:150, Sc-12732, Santa Cruz, Dallas, Texas, USA), MEF2C (1:200, 5030 S, Cell Signaling Technology Inc., Danvers, Massachusetts, USA), slow skeletal myosin (NOQ7.5.4D, NOQ, 1:200, ab11083, Abcam Corporation, Cambridge, England) or fast myosin skeletal heavy chain (My32, ab51263, 1:200, Abcam Corporation, Cambridge, England) and the appropriate fluorescence-conjugated secondary antibodies.

Techniques: Expressing, Western Blot, Staining

Journal: Frontiers in Cell and Developmental Biology

Article Title: Cellular and transcriptomic changes by the supplementation of aged rat serum in human pluripotent stem cell-derived myogenic progenitors

doi: 10.3389/fcell.2024.1481491

Figure Lengend Snippet: Expression levels of various markers in differentiated hPSC-derived myogenic progenitors treated with rat sera. The expression of Pax7 and Myogenin (MyoG) proteins were analyzed in the differentiated cells for 3 days with 20% young or aged rat sera. (A) The type of supplemented sera did not alter the percentage of total myogenic cells (expressing PAX7, MyoG, or both) (Student’s t -test, t = 1.115, df = 10, p = 0.291). There was no difference in the number (B) and proportions (C) of quiescent, activating and activated myogenic progenitors out of total cells (represented by cells expressing PAX7 + /MyoG − , PAX7 + /MyoG + and PAX7 − /MyoG + , respectively) (2-way ANOVA, F 1,30 = 1.958, p = 0.172 for (B) ; Student’s t -test, t < 0.001 df = 2, p > 0.999 for (C) . (D) On Day 7 of terminal differentiation, cells supplemented with aged rat sera showed an increased level of a cell proliferation marker Ki67 (t = 9.357, df = 10, p < 0.001; by unpaired two-tailed Student’s t-test). (E) Western blot analysis showed increased cyclin B1 and CDK 2 expression in hPSC-derived myogenic progenitors differentiated in 20% aged rat sera for 2 weeks. Beta-actin served as loading control. n = 6 for each group * p < 0.05.

Article Snippet: After rinsing with PBS, the cells were incubated with mouse monoclonal antibodies against PAX7 [1:40; Developmental Studies Hybridoma Bank (DSHB), Iowa City, IA] and rabbit polyclonal antibodies against MyoG (1:400; Santa Cruz, Dallas, TX) at 4°C overnight.

Techniques: Expressing, Derivative Assay, Marker, Two Tailed Test, Western Blot, Control