Journal: EMBO Molecular Medicine

Article Title: Identification of p38 MAPK inhibition as a neuroprotective strategy for combinatorial SMA therapy

doi: 10.1038/s44321-025-00303-6

Figure Lengend Snippet: ( A ) RT-qPCR analysis of Smn mRNA levels in untreated and Dox-treated NIH3T3-SMN2/Smn RNAi cells cultured for 5 days in the presence of either DMSO or EO1428, Clozapine, DHE and CGP7930 at a concentration of 10 μM. Mean, SEM, and individual values from Dox-treated cells normalized to DMSO-treated cells cultured without Dox are shown ( n = 3 independent biological replicates). One-way ANOVA followed by Tukey’s post hoc test. ns not significant. ( B ) RT-qPCR analysis of total (TOT) SMN2 mRNA levels in the same groups as in ( A ). Mean, SEM, and individual values from Dox-treated cells normalized to DMSO-treated cells cultured without Dox are shown ( n = 3 independent biological replicates). One-way ANOVA followed by Tukey’s post hoc test. ns, not significant. ( C ) RT-qPCR analysis of full-length (FL) SMN2 mRNA levels in the same groups as in ( A ). Mean, SEM, and individual values from Dox-treated cells normalized to DMSO-treated cells cultured without Dox are shown ( n = 3 independent biological replicates). One-way ANOVA followed by Tukey’s post hoc test. ns not significant. ( D ) Western blot analysis of SMN expression in the same groups as in ( A ). β-actin was used as a loading control. ( E ) Normalized mean, SEM, and individual values of SMN levels in Dox-treated relative to untreated NIH3T3-SMN2/Smn RNAi cells from three independent experiments as in ( D ). Kruskal–Wallis followed by Dunn’s post hoc test. ns not significant. ( F ) Western blot analysis of total and phosphorylated p38 MAPK in untreated and Dox-treated NIH3T3-SMN2/Smn RNAi cells. SMN and tubulin were used as controls. ( G ) Quantification of the levels of phosphorylated p38 MAPK relative to total p38 MAPK in NIH3T3-SMN2/Smn RNAi cells cultured with or without Dox from experiments as in ( B ). Normalized mean, SEM, and individual values from three independent biological replicates are shown. Two-tailed unpaired t -test. P = 0.031. ( H ) Dose-response analysis of the effects of the indicated p38 MAPK inhibitors on the proliferation of Dox-treated SMN2/Smn RNAi cells at day 5. The graphs show mean and SEM of normalized cell number relative to vehicle-treated cells at day 5 ( n = 6 biological replicates) for each tested concentration, and non-linear curve fitting. .

Article Snippet: EO1428 , Tocris , Cat#2908.

Techniques: Quantitative RT-PCR, Cell Culture, Concentration Assay, Western Blot, Expressing, Control, Two Tailed Test