Journal: International Journal of Molecular Sciences

Article Title: Global Impairment of Immediate-Early Genes Expression in Rett Syndrome Models and Patients Linked to Myelination Defects

doi: 10.3390/ijms24021453

Figure Lengend Snippet: IEGs transcriptional changes in prefrontal cortex and hippocampus of Mecp2 y/− mice. n.d = not differentially expressed.

Article Snippet: Specific primary antibodies used were Rb-Egr2 (EPR4004, AbCam, Cambridge, UK); Rb-JunB (ab128878, AbCam); Rb-Fos (#2250, Cell Signaling, Danvers, MA, USA); Rb-Jun (#9165S, Cell Signalling); b-actin peroxidase conjugated as control loading (A3854; Sigma-Aldrich).

Techniques: Activity Assay

Journal: International Journal of Molecular Sciences

Article Title: Global Impairment of Immediate-Early Genes Expression in Rett Syndrome Models and Patients Linked to Myelination Defects

doi: 10.3390/ijms24021453

Figure Lengend Snippet: Validation of IEGs expression in Mecp2 -null mice. ( a , b ) IEGs expression in the hippocampus and prefrontal cortex of 8-weeks mice. Graphs show mean ± SEM of n = 5–6 animals per condition. Student’s t -tests were used (* p < 0.05, ** p < 0.01, *** p < 0.001). ( c ) Western Blot analysis of EGR2, c-FOS, c-Jun and NPAS4 in the hippocampus of 8-weeks mice. β-ACTIN was used as loading control. Samples from two animals per condition are shown. Graphs on the right represent quantitation of band intensity (mean values ± SEM, two tailed Mann-Whitney test, * p < 0.05, ns = not significant). ( d ) Representative immunostaining of EGR2 protein (red) in the hippocampal regions of WT or Mecp2 -KO 8-week mice, counterstained with DAPI (blue). All images were processed with ImageJ Fiji version 1.50 g. Scale bar represents 200 µm. The graph corresponds to quantitation of n = 10 images per condition (mean values ± SEM, two-tailed unpaired t test, **** p < 0.001).

Article Snippet: Specific primary antibodies used were Rb-Egr2 (EPR4004, AbCam, Cambridge, UK); Rb-JunB (ab128878, AbCam); Rb-Fos (#2250, Cell Signaling, Danvers, MA, USA); Rb-Jun (#9165S, Cell Signalling); b-actin peroxidase conjugated as control loading (A3854; Sigma-Aldrich).

Techniques: Expressing, Western Blot, Quantitation Assay, Two Tailed Test, MANN-WHITNEY, Immunostaining

Journal: International Journal of Molecular Sciences

Article Title: Global Impairment of Immediate-Early Genes Expression in Rett Syndrome Models and Patients Linked to Myelination Defects

doi: 10.3390/ijms24021453

Figure Lengend Snippet: IEGs gene expression response to Forskolin is dysregulated in 8-weeks Mecp2 -KO mice neurons. ( a ) Mecp2 expression in cortical and hippocampal neurons, as measured by RT-qPCR. ( b ) Schematics indicating treatment of cultured primary neurons with forskolin and time-course of sample collection. ( c , d ) Time-course of Fos , Junb , Egr2 and Npas4 transcript levels analysis for both WT and Mecp2 -KO hippocampal ( c ) or cortical ( d ) neurons upon treatment with forskolin. All expression data are relative to WT unstimulated values ( n = 3 biological replicates for each condition, means ± SEM are represented, * p < 0.05 and *** p < 0.001 in Student’s t -tests).

Article Snippet: Specific primary antibodies used were Rb-Egr2 (EPR4004, AbCam, Cambridge, UK); Rb-JunB (ab128878, AbCam); Rb-Fos (#2250, Cell Signaling, Danvers, MA, USA); Rb-Jun (#9165S, Cell Signalling); b-actin peroxidase conjugated as control loading (A3854; Sigma-Aldrich).

Techniques: Expressing, Quantitative RT-PCR, Cell Culture

Journal: International Journal of Molecular Sciences

Article Title: Global Impairment of Immediate-Early Genes Expression in Rett Syndrome Models and Patients Linked to Myelination Defects

doi: 10.3390/ijms24021453

Figure Lengend Snippet: IEGs expression in WT and Mecp2 -KO, 8-week mice following kainic acid (KA) administration. ( a , c ) Mecp2 transcript levels in the hippocampus ( a ) or frontal cortex ( c ) of KA-treated versus untreated WT mice. ( b , d ) Expression levels of Fos , Junb , Egr2 and Npas4 one hour after KA administration in the hippocampus ( b ) or frontal cortex ( d ) of WT and Mecp2 -KO mice ( n = 5–6 animals for each condition, graphs represent means ± SEM, * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001 in Student’s t -tests).

Article Snippet: Specific primary antibodies used were Rb-Egr2 (EPR4004, AbCam, Cambridge, UK); Rb-JunB (ab128878, AbCam); Rb-Fos (#2250, Cell Signaling, Danvers, MA, USA); Rb-Jun (#9165S, Cell Signalling); b-actin peroxidase conjugated as control loading (A3854; Sigma-Aldrich).

Techniques: Expressing

Journal: International Journal of Molecular Sciences

Article Title: Global Impairment of Immediate-Early Genes Expression in Rett Syndrome Models and Patients Linked to Myelination Defects

doi: 10.3390/ijms24021453

Figure Lengend Snippet: IEGs expression in human RTT patients and cellular models. ( a , c ) RT-qPCR analysis of EGR2 , JUN , FOS and JUNB transcript levels in the hippocampus ( a ) or cerebellum ( c ) of post-mortem RTT patients or healthy control samples ( n = 3 subjects per condition, graphs represent means ± SEM, * p < 0.05, ** p < 0.01 in Student’s t -tests). ( b , d ) Western blot analysis of EGR2, JUN and FOS protein levels in the hippocampus ( b ) or the cerebellum ( d ) of post-mortem RTT patients or healthy control samples. Graphs on the right indicate quantitation of band intensity (mean ± SEM, * p < 0.05, *** p < 0.001 in Student’s t -tests) of the three biological replicates. ( e ) RT-qPCR analysis of FOS , JUNB and EGR2 expression upon overexpression of MeCP2 in a human neural progenitor model ( n = 3 biological replicates, graphs represent means ± SEM, ** p < 0.01, **** p < 0.0001, in Student’s t -tests). ( f ) RT-qPCR analysis of EGR2 , JUN , FOS and JUNB transcript levels in the peripheral blood of RTT patients or healthy control samples ( n ≥ 10 biological replicates, graphs represent means ± SEM, * p < 0.05 and ** p < 0.01 in Student’s t -tests).

Article Snippet: Specific primary antibodies used were Rb-Egr2 (EPR4004, AbCam, Cambridge, UK); Rb-JunB (ab128878, AbCam); Rb-Fos (#2250, Cell Signaling, Danvers, MA, USA); Rb-Jun (#9165S, Cell Signalling); b-actin peroxidase conjugated as control loading (A3854; Sigma-Aldrich).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Quantitation Assay, Over Expression