Journal: Glia

Article Title: N-myc downstream regulated family member 1 (NDRG1) is enriched in myelinating oligodendrocytes and impacts myelin degradation in response to demyelination.

doi: 10.1002/glia.24108

Figure Lengend Snippet: ( A ) Schematic illustrating the conditional knock-out breeding strategy used to generate the Olig1cre;Ndrg1floxed line, leading to deletion of Ndrg1 exons 6 and 7 in oligodendroglial lineage cells. ( B ) Representative western blot of NDRG1 in adult corpus callosum protein lysate from control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice. Tubulin was used as loading control. ( C ) Quantification of NDRG1 protein levels normalized to Tubulin- expression; histograms represent average values ± SD for n=3 samples (**** p < 0.0001 , unpaired t-test). ( D ) Quantitative real-time PCR analysis of Ndrg1, Ndrg2, Ndrg3 and Ndrg4 on corpus callosum tissues of adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice. Data represents average values normalized to control samples ± SD for n=4 mice ( *p < 0.05 , Student’s t-test). ( E-F ) Quantitative real-time PCR analysis of progenitors ( E ) and oligodendroglial ( F ) transcripts in corpus callosum samples of adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice. Data represents average values normalized to control samples ± SD for n=4 mice (* p < 0.05 , ** p < 0.01 , Student’s t-test). ( G ) Representative western blot of PDGFRA and OLIG2 in adult corpus callosum total lysate from control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice. Tubulin was used as control. ( H-I ) Quantification of PDGFRA and OLIG2 protein levels. Data were normalized to Tubulin-α expression; histograms represent average values ± SD for n=3 samples.

Article Snippet: Lineage specific knockout lines were generated by crossing Ndrg1 fl/fl females with Olig1 tm1(cre)Rth/J males heterozygous for cre (referred to Olig1 cre/+ in the manuscript) (RRID:IMSR_JAX:011105), to generate the conditional mutants ( Olig1 cre/+ ;Ndrg1 fl/fl ).

Techniques: Knock-Out, Western Blot, Control, Mutagenesis, Expressing, Real-time Polymerase Chain Reaction

Journal: Glia

Article Title: N-myc downstream regulated family member 1 (NDRG1) is enriched in myelinating oligodendrocytes and impacts myelin degradation in response to demyelination.

doi: 10.1002/glia.24108

Figure Lengend Snippet: ( A ) Schematics of experimental plan conducted in adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice, fed with cuprizone diet (n=3) for three weeks. ( B ) Representative confocal image of coronal brain sections from adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice after control chow or cuprizone diet, stained for OLIG2 (green), CC1 (red) and DAPI (blue). Scale bar = 500μm. ( C-E ) Quantification of the density of OLIG2+ cells and CC1+ cells in the corpus callosum of adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice. Data represents average number of OLIG2+ and CC1+ cells per mm 2 in the corpus callosum area ± SD for n=3 mice (* p < 0.05 , ** p < 0.01 , *** p < 0.001 , **** p < 0.0001, Two-way ANOVA with Tukey’s post-hoc test). ( D-F ) Percentage of OLIG2+ cells and of CC1+ cells in the corpus callosum of adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice compared to their respective genotype under control chow diet. Data represents average percentage of OLIG2+ and CC1+ cells per mm 2 in the corpus callosum area ± SD for n=3 mice (** p < 0.01 , *** p < 0.001 , unpaired t-test). ( G ) Representative western blot of CNPase, MBP and TUBULIN in adult corpus callosum total protein lysates from control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice either on chow or on cuprizone diet for the indicated time period. ( H-J ) Quantification of CNPase and MBP protein levels, normalized to tubulin expression. Data represents average values ± SD for n=3 mice. (* p < 0.05 , ** p < 0.01 , *** p < 0.001 and *** p < 0.0001, Two-way ANOVA with Tukey’s post-hoc test). ( I-K ) Percentage of CNPase and MBP protein levels in the corpus callosum of adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice compared to their respective genotype under control chow diet. Data represents average percentage of CNPase and MBP levels in the corpus callosum area ± SD for n=3 mice (* p < 0.05 , ** p < 0.01 , *** p < 0.001 , unpaired t-test) ( L ) Representative images of coronal brain sections stained with Luxol fast blue from adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice, fed either chow diet or 0.2% cuprizone diet. Scale bars = 2mm on gross image (left) and 50μm on high magnification (right). ( M ) Quantification of Luxol fast blue (LFB) intensity in the corpus callosum (CC) of adult control Olig1 +/+ ;Ndrg1 fl/fl and mutant Olig1 cre/+ ;Ndrg1 fl/fl mice. Data represents average intensity percentage in corpus callosum regions, normalized to the intensity the control group under chow diet, ± SD for n=3 mice. ((* p < 0.05 and ** p < 0.01 , Two-way ANOVA with Tukey’s post-hoc test).

Article Snippet: Lineage specific knockout lines were generated by crossing Ndrg1 fl/fl females with Olig1 tm1(cre)Rth/J males heterozygous for cre (referred to Olig1 cre/+ in the manuscript) (RRID:IMSR_JAX:011105), to generate the conditional mutants ( Olig1 cre/+ ;Ndrg1 fl/fl ).

Techniques: Control, Mutagenesis, Staining, Western Blot, Expressing