A comparative transcriptomic analysis of astrocytes differentiation from human neural progenitor cells

Abstract

Astrocytes are a morphologically and functionally heterogeneous population of cells that play critical roles in neurodevelopment and in the regulation of central nervous system homeostasis. Studies of human astrocytes have been hampered by the lack of specific molecular markers and by the difficulties associated with purifying and culturing astrocytes from adult human brains. Human neural progenitor cells (NPCs) with self-renewal and multipotent properties represent an appealing model system to gain insight into the developmental genetics and function of human astrocytes, but a comprehensive molecular characterization that confirms the validity of this cellular system is still missing. Here we used an unbiased transcriptomic analysis to characterize in vitro culture of human NPCs and to define the gene expression programs activated during the differentiation of these cells into astrocytes using FBS or the combination of CNTF and BMP4. Our results demonstrate that in vitro cultures of human NPCs isolated during the gliogenic phase of neurodevelopment mainly consist of radial glial cells (RGCs) and glia-restricted progenitor cells. In these cells the combination of CNTF and BMP4 activates the JAK/STAT and SMAD signaling cascades, leading to the inhibition of oligodendrocytes lineage commitment and activation of astrocytes differentiation. On the other hand FBS-derived astrocytes have properties of reactive astrocytes. Our work suggests that in vitro culture of human NPCs represents a valuable cellular system to study human disorders characterized by impairment of astrocytes development and function. Our datasets represent an important resource for researchers studying human astrocytes development and might set the basis for the discovery of novel human-specific astrocyte markers.

Cocaine alters Homer1 natural antisense transcript in the nucleus accumbens

“…Moreover, many NATs are expressed in the CNS where they have been shown to play fundamental roles in a variety of biological processes. For example, our previous work demonstrates that NAT dysregulation is associated with Alzheimer’s disease pathophysiology (Modarresi et al , 2011; Magistri et al , 2016), and in separate reports we have also revealed that NATs contribute to Parkinson’s disease, Autism, Fragile X mental retardation, seizures and metabolism (Scheele et al , 2007; Faghihi et al , 2010a; Pastori et al , 2014; Hsiao et al , 2016; Silva et al , 2016). Mechanistically, we and others have shown that NATs can mediate epigenetic modification of genes in cis , making them unexplored targets to achieve specific up-regulation of gene expression (Modarresi et al , 2012).…”

Plasminogen binding inhibitors demonstrate unwanted activities on GABA A and glycine receptors in human iPSC derived neurons

“…New isolation and culture protocols maintaining a non-reactive state of primary human and mouse astrocytes demonstrate the importance of xeno-free and supportive cultures (154,208,248). Additionally, by applying defined conditions hPSC derived astrocytes differentiating via a glia restricted progenitor (GRP) state demonstrate strong transcriptional changes after FBS exposure (20), which is also observed for primary GRP (155). However, withdrawal of FBS induced changes indicating a conversion back towards a non-reactive state (20).…”

“…The morphology of isolated primary astrocyte is affected by in vitro culturing, which been demonstrated to be dependent on the application of fetal bovine serum (FBS). Defined medium or FBS containing conditions show significant differences on morphology, maintaining profound stellate morphology while inducing polygonal morphology, respectively (154,155). Protoplasmic astrocytes in vivo can lose their bushy morphology acquiring a reactive phenotype reducing SLC1A2 while increasing CD44 expression (121).…”

“…For example, if the astrocyte is at maximum reactivity before stimulation the inflammatory induction will not be representative, meaning that the stimulation response will depend on the basal level of reactivity during non-stimulated conditions. Several factors are associated with astrocyte reactivity including isolation procedure (248), extracellular matrix components (171), and the use of FBS (154,155). New isolation and culture protocols maintaining a non-reactive state of primary human and mouse astrocytes demonstrate the importance of xeno-free and supportive cultures (154,208,248).…”

Identification and epigenetic analysis of divergent long non-coding RNAs in multilineage differentiation of human Neural Progenitor Cells

“…The analysis of astrocyte-specific lncRNAs was done as already mentioned for neurons. Here we combined NPCs, Astrocytes and annotated lncRNAs RNAseq datasets [17]. We then extracted XH lncRNA expression, as already explained for neuronal differentiation.…”