Genomic study of human glial progenitor development

Abstract

The broad aim of this research project was to understand molecular signals which underlie the development of human oligodendrocytes. Oligodendrocytes, which are responsible for producing myelin in the brain, are lost in demyelinating diseases such as leukodystrophy and multiple sclerosis. In development, glial progenitor cells (GPCs) give rise to oligodendrocytes and, following demyelination GPCs may give rise to new oligodendrocytes in a process known as remyelination. As such, the identification and characterization of novel molecular pathways that regulate oligodendrocyte fate commitment in human cells may provide the means by which to induce remyelination in human demyelinating disease. However the molecular processes that regulate these processes appear to be species specific and are poorly understood in human cells. We have previously shown that human GPCs labeled with the CD140a antigen (expressed by the PDGFRA gene) develop into oligodendrocytes while O4 is expressed by the committed oligodendrocytes. The fetal human forebrain dissociated cells were resolved by fluorescent activated cell sorting into different stages of oligodendrocytic lineage commitment and differentiation using CD140a and O4 antigens and genome-wide microarray analysis was performed (n=6, fetal samples). In addition, we profiled human CD140a + GPCs undergoing oligodendrocyte differentiation in vitro and in the presence of PDGF-AA which blocks GPC differentiation (n=4, fetal samples). We analyzed these datasets using R/Bioconductor software and identified genes specific to different glial lineage stages. Furthermore we used Gene Set Enrichment Analysis for quantifying the population-specific enrichment of gene sets regulated in the rodent oligodendrocyte progenitor cells. A model fitting approach was used to compare and contrast gene profiles of sorted populations. To identify networks of genes relevant for oligodendrocyte differentiation weight-based correlational analysis (WGCNA) was used for network construction. The gene ontology (GO), KeGG pathway, gene homology and Ensembl databases were used for annotating and characterizing the regulated gene sets. Among cell surface receptors, the muscarinic acid type 3 receptor (CHRM3) was highly differentially regulated in the CD140a + O4 + population. As CHRM3 has been shown to regulate rodent oligodendrocyte differentiation and was topologically clustered with oligodendrocyte specific genes in our gene-gene networks we predicted that agonism of CHRM3 may regulate human GPCs fate. Indeed, we found that treatment with oxotremorine, a muscarinic receptor-specific agonist, inhibited GPCs differentiation into O4 + oligodendrocytes (n=3). Thus we validated an extensive resource for transcriptional study of the GPC development and established means to identify putative targets which can be regulated to mobilize GPCs for treating demyelinating diseases.