During neonatal and post natal development oligodendrocytes

During neonatal and post-natal development, oligodendrocytes are abundantly produced in three temporally distinct waves (Kessaris et al., 2006), and differentiated oligodendrocytes express specific markers according to the state of development (Zhang, 2001). In the initial stage of differentiation, several transcription factors are involved in the oligodendroglial lineage restriction, including the downregulation of Sox2 and upregulation of glial lineage-associated Sox family – Sox8 and Sox9 (Wegner and Stolt, 2005). Consequently, expression of transcription factors as NKX2.2, Olig 1 and Olig 2 and Sox 10 increase (Kuhlbrodt et al., 1998), and give rise to progenitors that proliferate and differentiate into pre-oligodendrocytes, which express e.g. platelet-derived growth factor receptor-α (PDGFRα) with a bipolar or tripolar morphology. The expression of PDGFRα is the first important step in OPCs differentiation, since this expression establishes the OPC fate acquisition (Hart et al., 1989). During the transition from OPCs to immature oligodendrocytes, a new set of transcription factors is required to promote the OPCs differentiation. These include the transcriptional repression of E2F1 and c-MYC and the expression of StemRegenin 1 inhibitors such as p27KIP1 (CDKN1B) and SOX17 (Chew et al., 2011; Larocque et al., 2005). The immature oligodendrocytes exhibit a multipolar morphology, and can be recognized by the O4 antibody. After establishment of the oligodendroglial fate, the immature oligodendrocytes undergo extensive cellular growth and mature into myelinating oligodendrocytes expressing myelin basic protein (MBP) and 2′,3′-Cyclic-nucleotide 3′-phosphodiesterase (CNPase) (Seiberlich et al., 2015). This occurs with the transcriptional activation of myelin regulatory factor (MYRF) and the consequent positive-regulation of its targets MBP, CNPase, proteolipid protein 1 (PLP1), myelin-associated glycoprotein (MAG), and myelin oligodendrocyte protein (MOG) (Emery, 2010). In the past years, several groups, including ours, generated protocols to differentiate OPCs from human pluripotent stem cells (PSCs) (Djelloul et al., 2015; Douvaras et al., 2014; Sim et al., 2011; Stacpoole et al., 2013) to use as models for several diseases involving impaired myelination. Moreover, Vítězslav and colleagues (Vítězslav Bryja, 2006) described a simple protocol for generation of mESC from embryonic day 3.5 blastocysts allowing a starting point for several applications, including the generation of oligodendrocytes. Chen and colleagues have published a protocol for an in vitro isolation and maintenance of enriched rat and mouse OPC, facilitating studies on OPC lineage progression and myelin injury-recovery assays (Chen et al., 2007). Douvaras and colleagues developed a reduced-time protocol to generate oligodendrocytes from human pluripotent stem cells with increased percentage of O4 positive cells isolated (Douvaras and Fossati, 2015). Also, Wang and colleagues were the first to demonstrate the beneficial effect of transplanting hiPSC-derived PDGFRα/CD140a+ cells into hypomyelinated mice (Wang et al., 2013).
Applications of our protocols Here we present, in the same report, procedures to generate and isolate OPCs from different sources: brain and primary cultures from mice, mESC, hPSC (Fig. 1). In addition, we describe a new protocol that allows the generation of cultures highly enriched in mESCs-derived O4+ oligodendrocytes, by use of retinoic acid (RA) and smoothen agonist (SAG). The presented methods will allow a rapid and comprehensive approach to a simple and step-by-step protocol to obtain purified oligodendrocytes. The generated oligodendrocytes can be used to study demyelinating and/or dysmyelinating disorders, such as Guillain-Barré syndrome and multiple sclerosis. Moreover, oligodendrocytes are also involved in several neurodegenerative diseases such as amyotrophic lateral sclerosis, Alzheimer\'s disease and multiple system atrophy, as well as brain trauma and white matter stroke (Behrendt et al., 2013). Recently, oligodendrocyte\'s dysfunction with abnormal myelin formation is suspected of contributing to several mental illnesses, including depression, schizophrenia, autism and bipolar disorders (Bernstein et al., 2015). Importantly, there is an open field of applications to explore the interaction and role of oligodendrocytes with several other neurodegenerative and neurological disorders.