The online version of this article (doi:10.1186/s13229-017-0134-z) contains supplementary material, which is available to authorized users.
Rett syndrome (RTT) is a severe, neurodevelopmental disorder primarily affecting girls, characterized by progressive loss of cognitive, social, and motor skills after a relatively brief period of typical development. It is usually due to de novo loss of function mutations in the X-linked gene, MeCP2, which codes for the gene expression and chromatin regulator, methyl-CpG binding protein 2. Although the behavioral phenotype appears to be primarily due to neuronal Mecp2 deficiency in mice, other cell types, including astrocytes and oligodendrocytes, also appear to contribute to some aspects of the RTT phenotype. In addition, microglia may also play a role. However, the effect of Mecp2 deficiency in microglia on RTT pathogenesis is controversial.
In the current study, we applied whole transcriptome analysis using RNA-seq to gain insight into molecular pathways in microglia that might be dysregulated during the transition, in female mice heterozygous for an Mecp2-null allele (Mecp2 +/−; Het), from the pre-phenotypic (5 weeks) to the phenotypic phases (24 weeks).
We found a significant overlap in differentially expressed genes (DEGs) with genes involved in regulating the extracellular matrix, and those that are activated or inhibited when macrophages and microglia are stimulated towards the M1 and M2 activation states. However, the M1- and M2-associated genes were different in the 5- and 24-week samples. In addition, a substantial decrease in the expression of nine members of the heat shock protein (HSP) family was found in the 5-week samples, but not at 24 weeks.
These findings suggest that microglia from pre-phenotypic and phenotypic female mice are activated in a manner different from controls and that pre-phenotypic female mice may have alterations in their capacity to response to heat stress and other stressors that function through the HSP pathway.
Additional file 1: Figure S1. Representative histograms from the FACS analysis CD11b+ and CD11b− fractions obtained in the microglial isolation procedure. (JPG 62 kb)13229_2017_134_MOESM1_ESM.jpg
Additional file 2: Table S1. Total number of reads and mapped reads in 5- and 24-week Het females and WT controls. The mapping statistics were from tophat2 alignment report. QC of mapping results was done with RSeQC [ 82]. (XLSX 11 kb)
Additional file 3: Table S2. Entire gene list for Het vs WT in 5-week (sheet 1) and 24-week microglia (sheet 2). The genes are listing in descending order according to the adjusted p value (padj). Sheet 3 shows the number of DEGs in the 5-week samples after repeating the analysis five times by randomly sampling from the 5-week samples the numbers of RNA-seq reads matching to the read depth of the week 24 samples. (XLSX 7264 kb)13229_2017_134_MOESM3_ESM.xlsx
Additional file 4: Figure S2. Principal components analysis (PCA) was carried out on the top 5000 most variable genes as defined by the software DESeq2. The samples could be separated using PC1 and PC3. (JPG 47 kb)13229_2017_134_MOESM4_ESM.jpg
Additional file 5: Figure S3. A heat map shows the expression level of all week 5 DEGs across all samples, where expression levels here are quantified by normalized read counts (log10-transformed). As seen in the heat map, similar levels of expression were found in the week 4 and week 24 samples. (JPG 103 kb)13229_2017_134_MOESM5_ESM.jpg
Additional file 6: Table S3. Enrichment analysis of DEGs using the ToppGene suite. 5-week down-regulated DEGs (sheet 1); 5-week up-regulated DEGs (sheet 2); 24-week down-regulated DEGs (sheet 3); and 24-week up-regulated DEGs (sheet 4). (XLSX 7206 kb)13229_2017_134_MOESM6_ESM.xlsx
Additional file 7: Table S4. DEGs that overlap with M1- and M2-activated genes. Week 5 and week 24 DEGs were compared with genes activated in mouse macrophage following M1 and M2 activation found by Jablonski et al. as described in the Methods section [ 36]. Each sheet contains the overlap of up- or down-regulated genes with M1- or M2-activated genes. (XLSX 502 kb)
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- Transcriptome analysis of microglia in a mouse model of Rett syndrome: differential expression of genes associated with microglia/macrophage activation and cellular stress
Herbert M. Lachman
- BioMed Central