r d • mab353  (Developmental Studies Hybridoma Bank)

Journal: Molecular Therapy. Nucleic Acids

Article Title: Restoration of myogenesis in ALS-myocytes through miR-26a-5p-mediated Smad4 inhibition and its impact on motor neuron development

doi: 10.1016/j.omtn.2025.102581

Figure Lengend Snippet: PCM derived from muscles of hSOD1(G93A) Tg mice show impaired in vitro differentiation compared to the hSOD1(WT) counterpart (A) The expression of EmbMyHC, Pax7, MyoD, and myogenin was evaluated by WB in hSOD1(WT) and hSOD1(G93A) PCM at different DIV. The left shows representative WBs and the corresponding Coomassie blue staining of the membrane, while the graphs on the right report the densitometric analysis for each protein. n = 3 for each DIV and hSOD1 genotype. (B) p38 (MAPK) phosphorylation (Thr180/Tyr182) and the expression of p21 were analyzed by WB at different DIV. The top shows representative WBs, and the graphs on the bottom show the corresponding densitometric analysis by reporting the ratio between phosphorylated and total protein amounts for p38 while the abundance of p21 was determined by normalizing the optical density of immunoreactive bands to that of the corresponding Coomassie blue-stained lanes. n = 4 (different PCM) for each DIV and hSOD1 genotype. (C) 4 DIV PCM protein extracts were analyzed for phosphorylation of PKA substrates by WB using α-pPKAs as antibody. Loading control with the Coomassie blue staining. n = 4 (different PCM) for each DIV and hSOD1 genotype. (D) CREB phosphorylation (Ser133) was analyzed by WB at different DIV. The left shows representative WBs, and the graph on the right show the corresponding densitometric analysis by reporting the ratio between phosphorylated and total protein amount. n = 4 (different PCM) for each DIV and hSOD1 genotype. ∗ indicates bands considered for protein quantification. In (A, B, and D), ∗ p < 0.05 and ∗∗ p < 0.01, hSOD1(WT) vs. hSOD1(G93A) in each DIV; § p < 0.05 and §§ p < 0.01 with respect to hSOD1(WT) in 1 DIV; and # p < 0.05 and ## p < 0.01 with respect to hSOD1(G93A) in 1 DIV; two-way ANOVA followed by Sidak’s multiple comparison test. In (C), ∗ p < 0.05, Mann-Whitney test.

Article Snippet: The following primary Abs were used: anti-EmbMyHC mouse mAb (cl.BF-G6), 1:5,000 (Developmental Studies Hybridoma Bank, University of Iowa); anti-CREB rabbit mAb, 1:1,000 (Cell Signaling Technology, cat. no. 9197s); anti-pSer133 CREB mouse mAb, 1:1,000 (Cell Signaling Technology, cat. no. 9196s); anti-p38 rabbit polyclonal antibody (pAb), 1:1,000 (Cell Signaling Technology; cat. no. 9212); anti-pThr180/Tyr182 p38 rabbit mAb, 1:1,000 (Cell Signaling Technology; cat. no. 9211); anti-Pax7 mouse mAb, 1:300 (Santa Cruz Biotechnology, cat. no. sc-81648); anti-MyoD mouse mAb (cl.

Techniques: Derivative Assay, Muscles, In Vitro, Expressing, Staining, Membrane, Phospho-proteomics, Control, Comparison, MANN-WHITNEY

Journal: Molecular Therapy. Nucleic Acids

Article Title: Restoration of myogenesis in ALS-myocytes through miR-26a-5p-mediated Smad4 inhibition and its impact on motor neuron development

doi: 10.1016/j.omtn.2025.102581

Figure Lengend Snippet: hSOD1(G93A) co-cultured with hSOD1(WT) PCM recover the defective differentiation phenotype (A) Representative fluorescent images of the three different co-culture conditions (WT WT , top; WT G93A , middle; and G93A G93A , bottom) (see ). At 4 DIV, target cells were co-stained with anti-desmin antibody (red signal) and the Hoechst 33342 dye (blue signal). Scale bars, 20 μm. (B) The graph reports the frequency distribution of the myotube area under the three different culturing conditions. (C) The diagram reports the average myotube area in the three co-culture settings normalized to the mean value of the WT WT cultures. n = 21 for WT WT and WT G93A, while is 35 for G93A G93A . (D and E) The bar diagrams show the differentiation index and the fusion index under the three co-culture conditions. (F) At 4 DIV, the EmbMyHC expression was assessed by WB in the three co-culture conditions. The left shows a representative WB and the corresponding Coomassie blue staining of the membrane, while graphs on the right report the densitometric analysis. (G) The activation of CREB and the expression of myogenin were assessed at 4 DIV co-cultures by WB as described in . The left shows representative WBs and the corresponding Coomassie staining of the membrane, while the bar diagrams on the right show the corresponding densitometric analysis. n is reported inside the bars. For all panels, n.s., not significant, ∗ p < 0.05 ∗∗ p < 0.01, and ∗∗∗∗ p < 0.0001. One-way ANOVA followed by Tukey’s multiple comparison test.

Article Snippet: The following primary Abs were used: anti-EmbMyHC mouse mAb (cl.BF-G6), 1:5,000 (Developmental Studies Hybridoma Bank, University of Iowa); anti-CREB rabbit mAb, 1:1,000 (Cell Signaling Technology, cat. no. 9197s); anti-pSer133 CREB mouse mAb, 1:1,000 (Cell Signaling Technology, cat. no. 9196s); anti-p38 rabbit polyclonal antibody (pAb), 1:1,000 (Cell Signaling Technology; cat. no. 9212); anti-pThr180/Tyr182 p38 rabbit mAb, 1:1,000 (Cell Signaling Technology; cat. no. 9211); anti-Pax7 mouse mAb, 1:300 (Santa Cruz Biotechnology, cat. no. sc-81648); anti-MyoD mouse mAb (cl.

Techniques: Cell Culture, Co-Culture Assay, Staining, Expressing, Membrane, Activation Assay, Comparison

Journal: bioRxiv

Article Title: Complement inhibition by C3-siRNA treatment prevents AChR loss and reduces complement activation in the rat Passive Transfer Myasthenia Gravis (PTMG)

doi: 10.1101/2025.08.31.673367

Figure Lengend Snippet: A) After baseline measurements on day 0, subcutaneous treatment injections were administered on day 1 and 8. Intermediate testing and blood collection were performed on day 4, 7 and 11 (not shown). On day 15, indicated with a red arrow, PTMG was induced using mAb35, while non-MG animals received a saline injection of the same volume. B-C) Plasma C3 expression was reduced by the sc administration of C3-siRNA at various dose levels in B) non-MG animals and C) PTMG animals. For animals receiving 2 x 30 mg/kg treatment, doses were administered on days 1 and 8. For animals receiving 1 x 10 or 1 x 30 mg/kg doses, administration occurred on day 8, as indicated by the black arrows on the graphs. Saline-treated animals showed no significant change in C3 levels over time. Group averages are shown. D) Plasma C3 level changes in individual treatment groups on day 17, normalized to baseline levels on day 0. Individual plasma C3 levels per animal are shown. Statistical analysis was performed with Two-Way ANOVA (B-D), followed by One-Way ANOVA and post-hoc Bonferroni’s multiple comparison test (D); α=0.05; ****p<0.0001. A) Created in BioRender. Schöttler, A. (2025) https://BioRender.com/b4xxodd .

Article Snippet: PTMG was induced at 11-weeks of age by sc injection of 40 pmol/100 g body weight of mAb35 [deposited to the DSHB by Lindstrom, J. (Developmental Studies Hybridoma Bank (DSHB) product mAb35) on day 15, after C3-siRNA treatment.

Techniques: Saline, Injection, Clinical Proteomics, Expressing, Comparison

Journal: bioRxiv

Article Title: Complement inhibition by C3-siRNA treatment prevents AChR loss and reduces complement activation in the rat Passive Transfer Myasthenia Gravis (PTMG)

doi: 10.1101/2025.08.31.673367

Figure Lengend Snippet: A) After baseline measurements on day 0, subcutaneous treatment injections were administered on day 1 and 8. Intermediate testing and blood collection were performed on day 4, 7 and 11 (not shown). On day 15, indicated with a red arrow, PTMG was induced using mAb35, while non-MG animals received a saline injection of the same volume. B-C) Plasma C3 expression was reduced by the sc administration of C3-siRNA at various dose levels in B) non-MG animals and C) PTMG animals. For animals receiving 2 x 30 mg/kg treatment, doses were administered on days 1 and 8. For animals receiving 1 x 10 or 1 x 30 mg/kg doses, administration occurred on day 8, as indicated by the black arrows on the graphs. Saline-treated animals showed no significant change in C3 levels over time. Group averages are shown. D) Plasma C3 level changes in individual treatment groups on day 17, normalized to baseline levels on day 0. Individual plasma C3 levels per animal are shown. Statistical analysis was performed with Two-Way ANOVA (B-D), followed by One-Way ANOVA and post-hoc Bonferroni’s multiple comparison test (D); α=0.05; ****p<0.0001. A) Created in BioRender. Schöttler, A. (2025) https://BioRender.com/b4xxodd .

Article Snippet: PTMG was induced at 11-weeks of age by sc injection of 40 pmol/100 g body weight of mAb35 [deposited to the DSHB by Lindstrom, J. (Developmental Studies Hybridoma Bank (DSHB) product mAb35) on day 15, after C3-siRNA treatment.

Techniques: Saline, Injection, Clinical Proteomics, Expressing, Comparison