Journal: Nature Communications

Article Title: Activin A more prominently regulates muscle mass in primates than does GDF8

doi: 10.1038/ncomms15153

Figure Lengend Snippet: Changes in TA muscle weight following administration of increasing doses of ( a ) ActRIIB.hFc ( n =7), ( b ) α-GDF8 ( n =10, except for 0.1 and 0.75 mg kg −1 doses, n =5), ( c ) α-GDF8 in the presence of 10 mg kg −1 α-Act A ( n =7, except for 2.5, 25 and 50 mg kg −1 doses, n =8) and ( d ) α-Act A in the presence of 10 mg kg −1 of α-GDF8 ( n =5, except for 0.3 and 1.0 mg kg −1 doses, n =6) in male SCID mice for 21 days. Change in TA muscle weight was calculated as the difference versus isotype control group mean. ( e ) Representative images of laminin staining of TA muscle from mice injected with 10 mg kg −1 of α-Act A, α-GDF8, the combination of α-Act A and α-GDF8 or ActRIIB.hFc for 21 days ( n =6, except for ActRIIB.hFc, n =9). Mean fibre cross-sectional area ( f ) and fibre number per section ( g ) from studies described in e . mRNA levels of Mstn ( h ) and Inhba ( i ) from TA muscle of SCID mice treated with α-Act A or α-GDF8, the combination of the antibodies or ActRIIB.hFc for 21 days (10 mg kg −1 each, n =7). Data expressed in reads per kilobase of transcript per million mapped reads (RPKM). ELISA data for GDF8 ( j ) and activin A ( k ) from SCID mice treated for 21 days with α-GDF8, α-Act A or the combination (control group in j , n =5, all others n =6). ( l ) Change in TA muscle weights in C57BL/6 mice compared to SCID mice after 21 days of treatment ( n =5 for C57BL/6 study; n =12 for the control, n =5 for the α-GDF8 and combination groups, n =10 for the ActRIIB.hFc in SCID study). ( m ) Change in TA muscle weights in SCID mice following treatment with α-Act A and α-GDF8 combination (10 mg kg −1 each, n =7) in the absence or presence of α-GDF11 antibody (10 mg kg −1 , n =6). Data are mean±s.e.m. * P <0.05, ** P <0.01, *** P <0.001, **** P <0.0001 versus control ( a , b , f , j – m ), 10 mg kg −1 α-Act A ( c ) or 10 mg kg −1 α-GDF8 ( d ) by one-way ANOVA and Bonferroni post hoc test.

Article Snippet: Activin A, B, AB, AC, GDF8 and GDF11 protein reagents and the α-GDF11 antibody (MAB19581) were obtained from R&D Systems (Minneapolis, MN).

Techniques: Staining, Injection, Enzyme-linked Immunosorbent Assay

Journal: Nature Communications

Article Title: Activin A more prominently regulates muscle mass in primates than does GDF8

doi: 10.1038/ncomms15153

Figure Lengend Snippet: ( a ) Per cent change in lean mass over baseline in male and female cynomolgus monkeys ( n =5 per gender per group). Doses shown in mg kg −1 . Basal serum levels of ( b ) GDF8, ( c ) activin A and ( d ) GDF11 in SCID mice ( n =8), rats ( n =8), cynomolgus monkeys ( n =6) and humans ( n =5 per gender; 18–43 years of age). Data are shown as mean±s.e.m. *** P <0.001, **** P <0.0001 versus saline control by one-way ANOVA and Bonferroni post hoc test.

Article Snippet: Activin A, B, AB, AC, GDF8 and GDF11 protein reagents and the α-GDF11 antibody (MAB19581) were obtained from R&D Systems (Minneapolis, MN).

Techniques: Saline

Journal: Cell Reports Medicine

Article Title: Exercise-induced crosstalk between immune cells and adipocytes in humans: Role of oncostatin-M

doi: 10.1016/j.xcrm.2023.101348

Figure Lengend Snippet:

Article Snippet: GDF15 , Bio-Techne LTD , 9279-GD-050.

Techniques: In Vitro, Recombinant, Plasmid Preparation, Fluorescence, Blocking Assay, Protease Inhibitor, Enzyme-linked Immunosorbent Assay, Bicinchoninic Acid Protein Assay, Multiplex Assay, Software, Real-time Polymerase Chain Reaction, Microscopy

Journal: Biology

Article Title: Transdifferentiation of Human Fibroblasts into Skeletal Muscle Cells: Optimization and Assembly into Engineered Tissue Constructs through Biological Ligands

doi: 10.3390/biology10060539

Figure Lengend Snippet: Determination of efficacy of C2C12 differentiation in conjunction with the exposure to ligand combinations. C2C12s were differentiated for 7 days and treated with combination ligands of GDF11 (G), TMSB4X (T), IL6 (I), and TNF-α (F) at 10 ng/mL for seven additional days. ( A ) Fusion index was calculated from total myotube nuclei vs. total nuclei ( n = 16, mean + SD). ( B ) Multinucleation of C2C12 myotubes were quantified ( n = 11, mean + SD). ( C ) Nuclear density was evaluated from nuclear count per field of 5x microscopy ( n = 4, mean + SD). ( D ) C2C12 exposed to ligand combinations were stained to express nuclear MYOD1 ( n = 6, mean + SD). ( E ) Cells were stained with Ki67, and where similarly quantified based on average total nuclear count ( n = 6, mean + SD). ( F ) ACTN2 and Ki67 immunostaining of control cells. ( G ) Cells exposed to GTF showed decreases in fusion index and myonucleation levels, although no change in nuclear density and Ki67+ expression was detected. ( H ) GTIF supplementation significantly reduced skeletal muscle differentiation parameters fusion index and multinucleation, in addition to decreasing average nuclear density. ( I ) Control C2C12s expressing nuclear MYOD1. ( J ) GTF treatment greatly reduced nuclear fusion and showed limited differentiation capacity while expressing comparable levels of nuclear MYOD1. ( K ) Exposure of C2C12s to GTIF combination significantly inhibited skeletal muscle differentiation. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Proteins utilized included recombinant human Follistatin (Fs, 669-FO-025), recombinant human Myostatin (GDF8, 788-G8-010) or Growth Differentiation factor (GDF8), recombinant human basic Fibroblast Growth Factor 2 (FGF2, 233-FB-025), recombinant human GDF11 (1958-GD-010), recombinant human GDF15 (957-GD-025/CF), recombinant human Bone Morphogenetic Protein 4 (BMP4, 314-BP-010/CF), recombinant human BMP7 (354-BP-010), recombinant human Growth Hormone (hGH, 1067-GH-025), recombinant human Interleukin 6 (IL6, 206-IL-010), recombinant human Tumor Necrosis Factor Alpha (TNF-α, 210-TA-005) (All R&D Systems), and Thymosin β (TOCRIS, 3390).

Techniques: Microscopy, Staining, Immunostaining, Control, Expressing

Journal: Biology

Article Title: Transdifferentiation of Human Fibroblasts into Skeletal Muscle Cells: Optimization and Assembly into Engineered Tissue Constructs through Biological Ligands

doi: 10.3390/biology10060539

Figure Lengend Snippet: Effect of ligand combination exposure on differentiation of skeletal muscle cells derived from tHFs. Cells were transduced with MYOD1 fragments and induced to express the skeletal muscle phenotype via the induction of doxycycline and SB431542 over a 7-day period. Ligand combinations of GDF11 (G), TMSB4X (T), IL6 (I), and TNF-α (F) at 10 ng/mL were introduced for an additional week, and SB and Dox administration was discontinued. Skeletal muscle cells were fixed and stained on day 14 and characterized by various differentiation and proliferation parameters from 5× microscopy. ( A ) Fusion index of tHFs was evaluated by determining the ratio of myotube nuclei vs total nuclear count ( n = 16, mean + SD). ( B ) Cellular multinucleation was quantified to assess tHF development of differentiation ( n = 22, mean + SD). ( C ) Nuclear density was similarly assessed by quantifying nuclear count per field ( n = 4, mean + SD). ( D ) Nuclear MYOD1 was quantified ( n = 6, mean + SD). ( E ) Ki67 nuclei were also assessed with a nuclear count ( n = 6, mean + SD). ( F ) Control tHF myotubes were immunostained with ACTN2 and Ki67. ( G ) IL6 and TNF-α combination demonstrated significant decrease in differentiation parameters fusion index, multinucleation, myotube length, and diameter , although Ki67+ expression had increased. ( H ) Exposure of tHFs to combined GDF11, TMSB4X, IL6, and TNF-α showed similar results, however nuclear Ki67 expression was unchanged. ( I ) Untreated tHFs with ACTN2 and MYOD1 nuclear stains. ( J ) Cells treated with IF showed a decrease in MYOD1 nuclear expression. ( K ) Additionally, GDF11, TMSB4X, and IL6 exposure yielded similar results with respect to MYOD1+. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001.

Article Snippet: Proteins utilized included recombinant human Follistatin (Fs, 669-FO-025), recombinant human Myostatin (GDF8, 788-G8-010) or Growth Differentiation factor (GDF8), recombinant human basic Fibroblast Growth Factor 2 (FGF2, 233-FB-025), recombinant human GDF11 (1958-GD-010), recombinant human GDF15 (957-GD-025/CF), recombinant human Bone Morphogenetic Protein 4 (BMP4, 314-BP-010/CF), recombinant human BMP7 (354-BP-010), recombinant human Growth Hormone (hGH, 1067-GH-025), recombinant human Interleukin 6 (IL6, 206-IL-010), recombinant human Tumor Necrosis Factor Alpha (TNF-α, 210-TA-005) (All R&D Systems), and Thymosin β (TOCRIS, 3390).

Techniques: Derivative Assay, Transduction, Staining, Microscopy, Control, Expressing

Journal: Biology

Article Title: Transdifferentiation of Human Fibroblasts into Skeletal Muscle Cells: Optimization and Assembly into Engineered Tissue Constructs through Biological Ligands

doi: 10.3390/biology10060539

Figure Lengend Snippet: Skeletal muscle tissues were engineered from a composite fibrin/Matrigel hydrogel mixture with mouse skeletal myoblasts C2C12s, and subject to 10 ng/mL biological ligands. C2C12s were encapsulated and differentiated in a fibrin-based hybrid hydrogel over a 7-day period, 10 ng/mL biological ligands GDF11, TMSB4X, IL6 or TNF-α were administered after a week of tissue plating. ( A ) Immunohistochemical staining of C2C12 skeletal muscle constructs with ACTN2 and DAPI, demonstrated high cellular density. ( B ) Skeletal myotubes increased compactness and alignment towards central pillar regions where tensile force is maximal ( C ) Structural organization of C2C12s at pillar regions appeared disrupted due to gel contraction. ( D ) Cross-striated, multinucleated skeletal muscle form condensed tissues as demonstrated with high magnification 60× confocal microscopy. ( E ) Myotube diameter (µm) was not affected by one-week exposure to 10 ng/mL ligands. ( n > 32, mean + SD). ( F ) Nuclear density of skeletal muscle C2C12s within tissue were not impacted with ligand administration. ( n = 6, mean + SD).

Article Snippet: Proteins utilized included recombinant human Follistatin (Fs, 669-FO-025), recombinant human Myostatin (GDF8, 788-G8-010) or Growth Differentiation factor (GDF8), recombinant human basic Fibroblast Growth Factor 2 (FGF2, 233-FB-025), recombinant human GDF11 (1958-GD-010), recombinant human GDF15 (957-GD-025/CF), recombinant human Bone Morphogenetic Protein 4 (BMP4, 314-BP-010/CF), recombinant human BMP7 (354-BP-010), recombinant human Growth Hormone (hGH, 1067-GH-025), recombinant human Interleukin 6 (IL6, 206-IL-010), recombinant human Tumor Necrosis Factor Alpha (TNF-α, 210-TA-005) (All R&D Systems), and Thymosin β (TOCRIS, 3390).

Techniques: Immunohistochemical staining, Staining, Construct, Confocal Microscopy