Journal: bioRxiv

Article Title: Impact of histone deacetylase inhibition and arimoclomol on heat shock protein expression and disease biomarkers in primary culture models of familial ALS

doi: 10.1101/2023.12.13.571549

Figure Lengend Snippet: Effects of HDAC inhibitors, arimoclomol and combined treatments on HSPA1A in motor neurons expressing TDP-43 G348C . A Phase contrast image of spinal cord – DRG culture. Arrows point to motor neurons. B Small but significant increase in the percentage of motor neurons with HSPA1A immunoreactivity three days following microinjection of plasmid encoding TDP-43 G348C , compared to absence of labeling in neurons injected with mCherry (control). C-F Percentage of HSPA1A immunopositive motor neurons after three days of treatment with vehicle, the HDAC inhibitors C SAHA, D RGFP109, E RGFP963 or F Tubastatin A, or with arimoclomol alone or in combination with the respective HDAC inhibitor. HDAC inhibitors with class I activity ( C-E ) significantly increased the percentage of neurons expressing HSPA1A. Arimoclomol and the HDAC6 inhibitor Tubastatin A were ineffective. Data presented as mean ± S.D., n = 9 cultures. Statistical significance was evaluated by one-way ANOVA followed by Bonferroni post hoc analysis. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001. Scale bar = 20μm.

Article Snippet: Preparation and maintenance of dissociated spinal cord-DRG cultures from E13 CD1 mouse embryos (Charles River Laboratories, St.

Techniques: Expressing, Plasmid Preparation, Labeling, Injection, Activity Assay

Journal: bioRxiv

Article Title: Impact of histone deacetylase inhibition and arimoclomol on heat shock protein expression and disease biomarkers in primary culture models of familial ALS

doi: 10.1101/2023.12.13.571549

Figure Lengend Snippet: A-E ALS variants suppress Hspa8 mRNA levels in motor neurons. Hspa8 mRNA was quantified by smFISH three days after microinjection of TDP-43 G348C , FUS R521G or SOD1 G93A plus eGFP vectors. Microinjected motor neurons were identified by double-immunolabeling with antibodies to eGFP and TDP-43, FUS or SOD1. A Examples of Hspa8 smFISH. Significant decrease in RNA spot counts in B somata and C dendrites of motor neurons expressing ALS variants compared to eGFP control. D,E Reduction in Hspa8 mRNA only in somata of neurons expressing wild type (WT) TDP-43. Data presented as mean spot counts ± S.D., n = 11-61 neurons. F,G Reduction in HSPA8 protein in somata of motor neurons expressing ALS variants, compared to noninjected motor neurons in the same culture (CTL). F Representative micrographs of spinal cord-DRG culture double-labeled with anti-HSPA8 and anti-TDP-43. Arrow points to a neuron expressing TDP-43 G348C . G Graphed is mean pixel intensity of immunolabeling signal ± S.D., n = 9-24 neurons. Statistical significance was evaluated through one-way ANOVA followed by Bonferroni post hoc analysis. **p<0.01, ***p<0.001, ****p<0.0001. Scale bar = 15μm.

Article Snippet: Preparation and maintenance of dissociated spinal cord-DRG cultures from E13 CD1 mouse embryos (Charles River Laboratories, St.

Techniques: Immunolabeling, Expressing, Labeling