Introduction

NG2 cells are the source of oligodendrocytes (OLs) and thus termed oligodendrocyte precursor cells. NG2 cells are widely distributed in the gray and white matter of the central nervous system (CNS), comprising 4–8% of cells in the adult CNS (Dawson et al. 2003). NG2 cells are the fourth type of glial cell in the CNS, distinct from astrocytes, OLs, and microglia. Their morphology and functional properties differ among brain areas and environmental conditions (Chittajallu et al. 2004). NG2 cells can also differentiate into astrocytes and neurons in the presence of specific growth factors (Belachew et al. 2003). They divide, proliferate, and differentiate into mature OLs in various regions of the CNS throughout life (Li et al. 2020; Zhu et al. 2011) (Fig. 1).

Fig. 1
figure 1

Timing of the stages of NG2 cell differentiation

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Notch signaling can promote or suppress cell proliferation and differentiation, thus controlling cell fate during the development of organs, including the CNS (Givogri et al. 2002). Notch receptors, such as Notch1–4, are highly conserved transmembrane receptors. Notch ligands—including Jagged1, Jagged2, Delta-like1, Delta-like3, and Delta-like4—initiate Notch signaling via receptor binding (Gray et al. 1999). Multiple Notch receptors and ligands are expressed in the peripheral nervous system and CNS (Lindsell et al. 1996). Notch signal transduction relies on ligand–receptor binding, resulting in release of an active Notch fragment, which initiates intramembrane proteolysis. After release of the Notch intracellular domain (NICD) by proteolysis from a membrane tether, it travels to the nucleus, associates with a DNA-binding protein to assemble a transcription complex, and represses Hes target genes. This can be prevented by a gamma-secretase inhibitor (Jurynczyk et al. 2005).

NG2 cells proliferate and differentiate into mature OLs, which produce myelin basic protein (MBP) and proteolipid protein, which can wrap around axons. Hypomyelination may be caused by failed differentiation of NG2 cells, preventing the generation of new myelin. The Notch1 receptor is expressed in NG2 cells, the differentiation of which is controlled by the Notch pathway (Wang et al. 1998). Activation of Notch signaling suppresses differentiation of NG2 cells, leading to defective remyelination (Wang et al. 2017). Suppression of Notch1 expression in OLs results in an increased number of mature OLs in the gray matter of the developing CNS (Givogri et al. 2002).

As a kind of precursor cell, NG2 cells can not only differentiate into oligodendrocytes but also produce astrocytes and even neurons in special circumstances or pathological conditions (Diers-Fenger et al. 2001; Zhu et al. 2008). It has been found that activation of the Notch signaling pathway in the microenvironment after CNS injury leads to the differentiation of NG2 cells into more astrocytes (Khazaei et al. 2020), and astrocytes are the main cell types in glial scars (Hesp et al. 2018; Huang et al. 2018). Inhibition of the Notch signaling pathway inhibits the proliferation of reactive astrocytes and reduces inflammation-related secondary injury (Qian et al. 2019). After CNS injury, NG2 cells also have the potential to produce new neurons, which may be a potential strategy for neurofunctional repair (Boulanger and Messier 2017; Heinrich et al. 2014). However, the Notch signaling pathway has a negative effect on neuronal differentiation. Inhibition of the Notch signaling pathway promotes neuronal differentiation and maturation and the recovery of neural function (Chen et al. 2015).

Notch Signaling in the CNS

In the CNS, the ligand Jagged1 activates Notch1 receptors to inhibit the differentiation and maturation of NG2 cells. Notch1 receptors are expressed in NG2 cells, and Jagged1 is localized along the axons of retinal ganglion cells. In culture, NG2 cells do not express Notch ligands. Also, Notch and Jagged 1 expression decreased with age in the developing rat optic nerve (Wang et al. 1998).

Demyelinated axons are not a major source of Jagged1. However, Jagged1 is expressed at a high level in reactive astrocytes, whereas Notch1 receptors and Hes5 are expressed in NG2 cells in active multiple sclerosis (MS) lesions lacking remyelination. Jagged1 expression was increased in a broad zone from within the lesion center to the lesion border, and Jagged1 immunoreactivity decreased with distance from the area of demyelination. Astrocytes in remyelinated areas did not show Jagged1 immunoreactivity. Therefore, the expression of Jagged1 by astrocytes might be related to limited remyelination (John et al. 2002). Resting adult astrocytes do not express Notch1, but following demyelination, Notch1 is expressed by a small proportion of astrocytes at the lesion margins (Ge et al. 2002).

Jagged1 and Jagged2 are expressed constitutively in the trigeminal ganglion. Jagged1 is expressed in NG2 cells, OLs, and myelinating Schwann cells. Notch1 is present in post-mitotic neurons and OLs (Nonneman et al. 2018). In the SOD1-G93A model, Notch activation was universal in proliferating astrocytes, Jagged1 was upregulated only in proliferating microglia, and expression of the Notch ligand DLL4 was increased in activated astrocytes and degenerating OLs (Liu et al. 2020). The nigh-on ubiquitous expression of Notch1 and Jagged1 suggests a role for Notch–Jagged signaling in the CNS.

The Notch pathway is activated upon ligand (DLL1, Jagged1, or Jagged2) binding to a receptor (Notch1 or Notch2) (Gray et al. 1999). This is followed by a series of cleavage events, releasing the NICD, which translocates to the nucleus and modulates the expression of Hes1 and Hes5, basic helix loop–helix-type transcriptional repressors. This results in the inhibition of NeuroD and Mash1 expression (Ohtsuka et al. 1999). Gamma-secretase is a critical component of the Notch signaling pathway, and its inhibition may interfere with Notch-related processes. The Notch signaling inhibitor, DAPT, suppressed the expression of Hes1 and Hes5 (Palagani et al. 2012).

Effect of Notch Signaling on the Proliferation and Differentiation of NG2 Cells

The Notch pathway plays a key role in cell fate determination in the CNS and is involved in several crucial events in glial cell development, such as maintaining NG2 cells in an undifferentiated state.

A regulatory role for the Notch pathway in the differentiation of NG2 cells was suggested by the finding that their differentiation was inhibited by Notch ligands. Other evidence supports a role for the Notch pathway in controlling the timing of NG2 cell differentiation (Wang et al. 1998). A study using the Cre/loxP system in transgenic mice to selectively inhibit Notch1 signaling in NG2 cells showed that the function of Notch1 is crucial for OL development and differentiation in the brain and spinal cord. Also, ablation of Notch1 in NG2 cells led to ectopic production of prematurely differentiated immature OLs in the mouse spinal cord. Most of the prematurely differentiated OLs were found in the gray matter at P0, where immature OLs were scarce in control animals (Genoud et al. 2002). To confirm involvement of Notch1 signaling in remyelination in vivo, Zhang and colleagues generated an Olig1Cre: Notch112f/12f mouse model in which Notch1 was selectively inactivated throughout the oligodendrocyte lineage. In Notch-inactivated mice, the repair of demyelinated lesions in the corpus callosum (CC) was accelerated. In addition, experiments in vitro confirmed that Notch1 signaling promoted the expansion of NG2 cells but inhibited their differentiation and myelin formation (Zhang et al. 2009). To investigate whether Notch–Jagged signaling regulates the rate of remyelination, their expression was compared between young and older animals. However, no correlation between their expression and the remyelination rate was found, and the lesions underwent complete remyelination in older animals. Therefore, adult expression of Notch1 and Jagged1 neither prevented nor played a major rate-determining role in remyelination, in contrast to developmental myelination. Also, Notch1 ablation in NG2 cells of cuprizone-treated Plp-creER Notch1(lox/lox) transgenic mice did not significantly influence the remyelination parameters of knockout or control mice(Stidworthy et al. 2004).

Notch signaling also promotes differentiation of NG2 cells via other pathways. F3/contactin acts as a functional ligand of Notch, triggering gamma-secretase-dependent nuclear translocation of the NICD and recruitment of Deltex1 before or after releasing NICD into the cytoplasm. The NICD/RBP-J/Deltex1 complex may undergo specific but unknown modifications before moving to the nucleus, where it induces the expression of myelin-associated glycoprotein, promoting the differentiation of NG2 cells. This process can be blocked by dominant-negative expression of Notch1, Notch2, and two Deltex1 mutants lacking the RING-H2 finger motif, but not by dominant-negative expression of RBP-J or Hes1 antisense oligonucleotides. Therefore, F3/contactin initiates Notch/Deltex1 signaling, promoting oligodendrocyte maturation and myelination (Hu et al. 2003) (Fig. 2).

Fig. 2
figure 2

Roles of Jagged1/Notch and F3/Notch in the differentiation of NG2 cells. Jagged1/Notch signaling activates Notch signaling, release of the Notch intracellular domain (NICD), and recruitment of RBP-J. Next, RBP-J forms a complex with NICD, translocates to the nucleus, and induces Hes1/Hes5 expression to inhibit the differentiation of NG2 cells. In contrast, F3/contactin interacts with Notch receptors on the surface of NG2 cells, stimulating Notch signaling and recruiting Deltex1 before or after releasing NICD into the cytoplasm. The NICD/RBP-J/Deltex1 complex may undergo specific but unknown modifications before moving to the nucleus, where it induces the expression of myelin-associated glycoprotein (MAG) to promote the differentiation of NG2 cells

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Delta–Notch signaling is required for spinal cord oligodendrocyte specification. In a transgenic, conditional expression system, constitutive Notch activity promoted the generation of excess NG2 cells. Additionally, dla−/−, dld−/−, and mib−/− embryos did not produce NG2 cells or premyelinating OLs. Therefore, Delta–Notch signaling promotes the generation of NG2 cells. Notch signaling promotes the specification of neural precursors to an oligodendrocyte fate and subsequently regulates their differentiation, possibly matching the development of myelinating OLs to their target axons (Park and Appel 2003). In delta-like 1 mutant mice, neurospheres decreased the number of NG2 cells, and addition of a soluble Notch ligand to wild-type neurospheres enhanced their generation (Grandbarbe et al. 2003).

Hes1 and Hes5 are highly expressed in developing mammalian brain. Their activation by Notch signaling inhibits oligodendrocyte maturation and differentiation (Jarriault et al. 1998; Wang et al. 1998). The progressive decrease in Hes5 expression in vivo may also reflect a decrease in Notch signaling. Moreover, overexpression of Hes5 inhibits oligodendrocyte differentiation, which is induced by mitogen withdrawal or thyroid hormone addition. This suggests that Hes5 is important in Notch-mediated inhibition (Kondo and Raff 2000).

In zebrafish notch3 mutants, analysis of notch3st51 and an insertional allele of notch3 revealed that Notch3 is required for the development of, and MBP gene expression in, NG2 cells during larval development. Reduced MBP expression in embryos is associated with fewer NG2 cells in notch3 mutants (Zaucker et al. 2013).

Effect of Other Signaling Pathways on the Proliferation and Differentiation of NG2 Cells

The timely proliferation and differentiation of NG2 cells depend on a highly coordinated series of events regulated by multiple intracellular and extracellular factors (Miller 2002). The Wnt/β-catenin signaling pathway is an obvious negative regulator of NG2 cell proliferation and differentiation, and the Notch signaling pathway also regulates these processes (Fancy et al. 2009). Other regulatory factors, including the GSK3 and PDGF signaling pathways, are also involved in the regulation of NG2 cell proliferation and differentiation, and they interact with each other. The Wnt/β-catenin signaling pathway is a typical negative regulator of NG2 cell differentiation, and abnormalities in this pathway can also lead to myelin regeneration disorder in multiple sclerosis (Galimberti et al. 2011; Fancy et al. 2009). In developing CNS, the Wnt/β-catenin signaling pathway mainly inhibits the differentiation and maturation of NG2 cells but does not affect the proliferation of NG2 cells (Langseth et al. 2010). Activation of β-catenin, a signaling molecule of the Wnt signaling pathway, can regulate the differentiation of NG2 cells but does not affect the proliferation of NG2 cells (Dai et al. 2014). However, the role of the Wnt/β-catenin signaling pathway as an inhibitor of myelin formation was challenged by a study that showed that inhibition of the Wnt/β-catenin signaling pathway prevented the formation of mature myelin. It was speculated that the Wnt/β-catenin signaling pathway might initially regulate the maturation of NG2 cells by inhibiting differentiation but later become a driving force in the later stage of myelin formation (Tawk et al. 2011). GSK3 is a key regulator of the Wnt/β-catenin signaling pathway that participates in the regulation of NG2 cell differentiation and myelin formation (Fancy et al. 2009). GSK3 is widely involved in a variety of cell biological processes, including division, proliferation, and differentiation, the regulation of which is of great significance in the treatment of diseases such as multiple sclerosis and white matter damage (Kockeritz et al. 2006). GSK3β is a negative regulator of cell fate and the target of many signaling pathways (Cohen and Goedert 2004). When GSK3β inhibitors are given, a large number of mature oligodendrocytes are formed (Azim and Butt 2011). This finding is helpful to promote the differentiation, maturation, and myelin regeneration of NG2 cells. The positive effect of GSK3β inhibitors is to counteract and overcome the negative effects of the Wnt/β-catenin signaling pathway on the differentiation and myelination of NG2 cells, and studies have also shown that inhibition of GSK3β reduces the activation of the Notch signaling pathway and promotes the differentiation of NG2 cells (Fancy et al. 2009). NG2 cells retain the expression of the PDGF-α receptor, which is also a specific index for the identification of NG2 cells (Sim et al. 2011). PDGF is an important neurotrophic factor that promotes the differentiation of NG2 cells (Mohapel et al. 2005). Activation of the PDGF signaling pathway can significantly increase the number of NG2 cells in an animal model of demyelination (Hill et al. 2013). There is also an interaction between the PDGF signaling pathway and the Notch signaling pathway that affects the transcriptional level of target genes in the Notch signaling pathway (Liang et al. 2017). In this chapter, we mainly discuss the effect of the Notch signaling pathway on the differentiation of NG2 cells in demyelinating diseases.

Crosstalk Between Other Molecular Events and Notch Signaling in the CNS

Diverse molecular events influence Notch signaling and the differentiation of NG2 cells, but the role of other molecular events in the CNS is unclear (Fig. 3).

Fig. 3
figure 3

Regulation of the differentiation of NG2 cells by the Notch signaling pathway. The differentiation and myelination status of NG2 cells can be determined by the balance between opposing signaling systems. Positive regulators of Notch signaling promote, and negative regulators inhibit, the differentiation of NG2 cells

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F3/Contactin and NB-3

F3/contactin and NB-3 are members of the F3/contactin family of the immunoglobulin superfamily, and F3/contactin is expressed in various regions of the brain. F3/contactin and NB-3, as functional ligands of Notch, enhance Notch1 and Notch2 expression, promoting oligodendrocyte maturation. All of these events require Deltex1 as an intermediate factor (Hu et al. 2006).

Myelin Protein 36 K

36 K is one of the most abundant proteins in the zebrafish brain. Identifying the function of 36 K in zebrafish myelin would enhance understanding of demyelinating diseases and remyelination in the human CNS. 36 K plays an important role in oligodendrocyte differentiation by inhibiting the Notch signaling pathway. 36 K also regulates the synthesis of transmembrane Notch ligands and promotes intramembrane gamma-secretase processing of Notch. A gamma-secretase inhibitor prevented activation of Notch, rescuing the number of NG2 cells in 36 K morphants (Nagarajan et al. 2020).

Astrocyte-Derived Endothelin 1 (ET-1)

ET-1 is a secreted signaling peptide that suppresses remyelination and is highly expressed in reactive astrocytes of demyelinated lesions. ET-1 promotes Notch activation in NG2 cells during remyelination by inducing Jagged1 expression in reactive astrocytes. Inhibiting ET signaling prevents Notch activation in demyelinated lesions and accelerates remyelination. Genetic ablation of ET-1 also modulates Jagged1/Notch1 signaling and the differentiation of NG2 cells. PD142,893, a potent inhibitor of ET-1 signaling, prevents Jagged1 induction and Notch activation (Hammond et al. 2014). However, although ET-1 inhibits differentiation of NG2 cells via an astrocyte-dependent pathway, it may also signal directly to NG2 cells, which also express ET receptors (Gadea et al. 2009). Endothelin-2 reportedly promotes remyelination in the rat cerebellum (Yuen et al. 2013).

Mutation of fbxw7

fbxw7 encodes the substrate recognition component of a ubiquitin ligase, which targets Notch and other proteins for degradation. Fbxw7 attenuates Notch signaling during zebrafish neural development, thereby suppressing generation of excess NG2 cells. Notch signaling is elevated in fbxw7-mutant embryos, indicating that Notch proteins are functionally relevant targets of Fbxw7-mediated ubiquitination during oligodendrocyte specification (Snyder et al. 2012).

Fibroblast Growth Factor 2 (FGF2)

FGF2 inhibits the differentiation of NG2 cells into myelinating OLs during development and remyelination by activating fibroblast growth factor receptor signaling, predominantly via fibroblast growth factor receptor 1, in oligodendrocyte-lineage cells (Zhou et al. 2006). FGF2 induces Notch1 expression in immature OLs (Faux et al. 2001). Notch1 signaling increases the responsiveness to FGF2 in telecephalic progenitors (Yoon et al. 2004). FGF2 signaling may interact with downstream components of the Notch signaling pathway, including Maml1 and Hes5. Therefore, the interaction of FGF2 with Notch signaling components may be critical for the regulation of differentiation and myelination of NG2 cells during CNS development (Zhou and Armstrong 2007).

Sox17

SRY-Box (Sox)-containing transcription factors are evolutionarily conserved proteins essential for the differentiation and maturation of the developing nervous system. Sox17 is the only member of the SoxF family involved in CNS glial development, and it is upregulated during postnatal oligodendrocyte development and promotes oligodendrocyte differentiation (Sohn et al. 2006). Sox17 also promotes the differentiation of cultured NG2 cells (Chew et al. 2011). The numbers of Olig2-expressing cells and mature OLs are decreased by Sox17 ablation, leading to hypomyelination and motor dysfunction. Following lysolecithin (LPC)-induced demyelination, Sox17 deficiency significantly inhibits oligodendrocyte regeneration. Sox17 promotes progenitor expansion and differentiation via Notch signaling and thus contributes to oligodendrocyte generation. TCF7L2 expression is also regulated by both Sox17 and Notch, and Sox17 regulates Notch1 receptor and Hes effectors (Chew et al. 2019).

Tocopherol Derivative TFA-12

The tocopherol long-chain fatty alcohol TFA-12 is a synthetic molecule that combines an α-tocopherol moiety and a neurotrophic ω-alkanol side chain with 12 carbon atoms. TFA-12 is a member of the vitamin E family and a modulator of MS because of its antioxidant and anti-inflammatory effects; it also ameliorates white matter damage in experimental models. TFA-12 is an inhibitor of microglial activation in vitro and decreases secretion of nitric oxide and tumor necrosis factor α (Muller et al. 2004). In two rodent models of MS (an experimental autoimmune encephalomyelitis model and LPC-induced demyelination model), TFA-12 promotes differentiation of NG2 cells into mature OLs and remyelination by inhibiting Notch/Jagged1 signaling (Blanchard et al. 2013). TFA-12 represses expression of the Notch downstream effectors Hes1 and Hes5 and concomitantly upregulates Mash1, a bHLH transcription factor involved in oligodendrocyte differentiation. TFA-12 also reverses the Jagged1-mediated inhibition of NG2 cell differentiation (Parras et al. 2007). TFA-12 directly binds to Notch receptors, inhibiting gamma-secretase activity and the nuclear translocation of NICD (Blanchard et al. 2013).

Transforming Growth Factor β (TGF-β1)

TGF-β1 has been detected in a range of CNS conditions with a traumatic or inflammatory etiology and induces Jagged1 expression in primary cultures of human astrocytes. In contrast, astrocytes do not show Jagged1 immunoreactivity in remyelinated areas. The effect of TGF-β1 on Jagged1 inhibits the maturation of NG2 cells but does not induce Delta1 ligand. Therefore, TGF-β1 is associated with activation of the Notch pathway, inhibiting oligodendrocyte maturation and myelination (John et al. 2002). However, other findings contradict the above notion that TGF-β promotes NG2 cell proliferation in 8-day adult brain SVZ neurosphere cultures and triggers their differentiation into mature OLs. The gamma-secretase inhibitor DAPT significantly decreased the population of PDGFRα+ NG2 cells in TGF-β-treated neurospheres (Gomez et al. 2018).

Testicular Orphan Receptor 4 (TR4)

TR4 is an orphan nuclear receptor important in the development and maturation of the CNS, particularly in the differentiation and maturation of OLs in the forebrain (Young et al. 1997). Hypomyelination in TR4−/− forebrains was associated with a decreased number of mature OLs, possibly mediated by the Jagged1–Notch signaling pathway. Jagged1 expression is higher in axon fiber-enriched regions in the developing TR4−/− forebrain, indicating that Notch signaling is enhanced when NG2 cells contact these axons. This inhibits oligodendrocyte maturation and is correlated with decreased myelination but promotes astrocyte generation (Tanigaki et al. 2001). In addition, the timely downregulation of Jagged1 in axons is disrupted in the TR4−/− forebrain, implicating TR4 in the regulation of Jagged1. Notch signaling is more highly activated in TR4−/− brains in which Hes1 is not expressed, indicating that Hes1 is not a major effector. Regulation of Hes5 by TR4 via a non-Notch-related pathway is possible. Therefore, crosstalk between TR4 and Notch signaling is important for oligodendrocyte differentiation and maturation. The altered Jagged1–Notch signaling in the TR4−/− forebrain indicates that TR4 is necessary for proper myelination in the CNS (Zhang et al. 2007).

Notch and Demyelinating Diseases

MS

MS is an immune-mediated disorder characterized by inflammation and multifocal demyelination accompanied by progressive neurodegeneration, and it typically affects young people. MS is related to OL damage, failed remyelination, and axon degeneration, but these lost functions can be restored by generating OLs to place new myelin sheaths on demyelinated axons. NG2 cells give rise to OLs during CNS development, remain quiescent and undifferentiated during adulthood, and are recruited and undergo proliferation and differentiation during CNS injury. MS lesions contain numerous NG2 cells with myelination ability that do not differentiate into mature OLs, indicating no myelin synthesis. Activation of Notch signaling may hamper NG2 cell differentiation and remyelination, and inhibition of Notch could promote axon myelination, indicating therapeutic potential for MS (Zhang et al. 2009).

A model of focal demyelination induced by LPC injection in the CC has shown that a single intracranial injection of apotransferrin (aTf) induces an increase in F3/contactin levels and in myelin-associated glycoprotein gene expression at 24 h. DAPT injection reverses aTf-induced remyelination, implicating Notch signaling in the promyelination activity of aTf (Aparicio et al. 2013). A mouse model of acute demyelination has enabled exploration of the effect of Notch1 on remyelination in MS. Inhibition of Notch1 suppressed the Hes and Jagged‐1 protein levels and promoted the differentiation of NG2 cells and formation of myelin (Fan et al. 2018).

The toxic cuprizone-induced demyelination model, which mimics pattern III lesions of MS, shows an increased proportion of Jagged1+/GFAP+ cells. This is associated with enhanced Jagged1-driven Notch signaling activation in NG2 cells during early demyelination, subsequently increasing the proportion of F3/contactin+/NG2+ cells and promoting F3/contactin transcription during remyelination in the CC (Mathieu et al. 2019). Therefore, Notch signaling mediates degenerative disorders by regulating the proliferation, migration, and differentiation of NG2 cells and could be a therapeutic target in conditions related to demyelination.

Amyotrophic Lateral Sclerosis (ALS)

ALS is a late-onset degenerative disease affecting mainly motor neurons with an oligodendrocyte pathology and reactive astrocytes and microglia. The clinical features include muscle denervation, muscle weakness and atrophy, and ultimately paralysis and denervation of the respiratory muscles, leading to respiratory failure and death. In a model of ALS, the Notch signaling pathway is abnormally activated in the spinal cord of SOD1G93A mice, as well as in that of patients with sporadic ALS. The Notch ligand Jagged1 is highly expressed in reactive astrocytes in the spinal cord of mice and patients with ALS (Nonneman et al. 2018). Therefore, abnormal Notch signaling activation contributes to the pathogenesis of ALS, participates in other pathogenetic processes, and influences the proliferation and differentiation of NG2 cells into mature OLs (Philips et al. 2013).

Exposure to Hyperoxia

Hyperoxia damages the immature brain by inducing an inflammatory response (Nasoohi et al. 2012). Death of NG2 cells has been reported in mice exposed to 80% oxygen for 48 h (Schmitz et al. 2011). Hyperoxia reduces the number of mature OLs (MBP+) and increases that of NG2 cells after hyperoxia on postnatal day 12. DAPT pretreatment significantly ameliorates hyperoxia-induced disruption of oligodendrocyte maturation, increases the expression of MBP, and alleviates necrosis, cytoplast swelling, and karyoplast dissolution of immature brain cells in white matter. Mice pretreated with DAPT before hyperoxia showed significant decreases in the escape latency time and distance, dwell time, and frequency of crossing the platform, suggesting that the Notch pathway contributes to these cognitive changes in the immature brain. Therefore, Notch activation could hinder NG2 cell differentiation and induce dysregulation of oligodendrocyte maturation in the immature brain after hyperoxia and lead to behavioral abnormalities. Selective pharmacological inhibition of the Notch signaling pathway could ameliorate the adverse effects of hyperoxia. The Notch signaling pathway may be an important target for ameliorating hyperoxia-induced damage and behavioral changes in the neonatal brain (Du et al. 2017).

Schizophrenia

The key features of schizophrenia are white matter disturbances and myelin impairment. Electron microscopy of postmortem samples from patients with schizophrenia showed aberrant myelination of synaptic terminals, increased density of concentric lamellar bodies, and apoptosis/necrosis of OLs (Uranova et al. 2001). The Notch4 locus is a candidate susceptibility gene for schizophrenia (Stefansson et al. 2009), and the mRNA level of Notch1 is aberrant in parvalbumin-immunoreactive neurons in patients with schizophrenia (Pietersen et al. 2014). Quetiapine is a novel second-generation antipsychotic used to treat schizophrenia that promotes NG2 maturation, OL regeneration, and myelin repair (Zhang et al. 2012). Quetiapine exerted antipsychotic and myelin-protective effects in a Notch-dependent manner. Moreover, quetiapine ameliorated the cuprizone-induced inhibition of Notch signaling factors, such as Notch1, Hes1, and Hes5, in the forebrain, and MW167 suppressed these protective effects of quetiapine. Therefore, the antipsychotic and myelin-protective effects of quetiapine are mediated by Notch signaling, which thus has potential as a target for the development of antipsychotic drugs (Wang et al. 2015).

Stroke and Ischemia

Stroke is a neurological disease caused by many factors, such as cerebral artery stenosis, occlusion, or rupture, and its clinical manifestation is transient or permanent brain dysfunction. As one of the three most common diseases in the world, stroke has a high mortality and disability rate and is a serious threat to human life and health. Ischemic stroke is the most common form of stroke. White matter is particularly sensitive to stroke. Because the blood flow of white matter is lower than that of gray matter, deep white matter has a lower blood supply (Pantoni et al. 1996). Oligodendrocytes are the main cell types in white matter, and the differentiation and maturation of oligodendrocytes in the brain are very important for the maintenance and repair of white matter (Lo et al. 2003). NG2 cells are one of the cell types that respond quickly to ischemic injury and can differentiate into mature oligodendrocytes in the preparation for the formation of a new myelin sheath (Zhang et al. 2013). It was shown that at 7 days after cerebral ischemia–reperfusion injury, the number of NG2 cells in the area around the ischemic focus had increased significantly, and there were morphological changes that included enlarged cell bodies and hypertrophic protuberances. However, the number of NG2 cells in the central area of the ischemic focus decreased significantly, and the number of NG2 cells in the contralateral brain area did not change. It has been suggested that after cerebral ischemia–reperfusion, the NG2 cells proliferating in the surrounding area may differentiate into mature oligodendrocytes, thus supplementing dead oligodendrocytes and participating in the myelination of axons, which is of great significance to the regeneration and repair of brain tissue after ischemic injury (Matsumoto et al. 2008; Tanaka et al. 2001). Another study found that the number of NG2 cells in the injured juvenile striatum increased significantly 7 days after transient middle cerebral artery occlusion, indicating that NG2 cells responded to ischemic injury and increased their proliferation rate. In adult mice, NG2 cells undergo morphological changes after ischemic injury. NG2 cells show high resistance to ischemic damage in the juvenile striatum (Ahrendsen et al. 2016; Levine 2016). In ischemic injury, NG2 cells also exhibit changes in ion channels and membrane receptors as well as cell death induced by excitotoxicity (Boda et al. 2011; Pivonkova et al. 2010). The Notch signaling pathway in glial cells is activated after ischemia–reperfusion. Treatment with DAPT maintains the proliferation of NG2 cells in the area around the ischemic focus but also reduces the proliferation of reactive astrocytes, enabling NG2 cells to retain the ability to promote myelin repair, which has a beneficial effect on the recovery of ischemic injury (Marumo et al. 2013). These findings suggest that NG2 cells will be a potential therapeutic target for the treatment of ischemic injury.

Reaction of Notch Signaling and NG2 Cells in Other CNS Diseases

NG2 cells respond not only to demyelinating diseases but also to a variety of CNS injuries, including brain injury, spinal cord injury, inflammation, and other neuropathological processes, by changing their own morphology, proliferation, differentiation, and migration, which plays an important role in functional recovery, myelin repair, and immune regulation (Dawson et al. 2003; Levine et al. 2001). NG2 cells can increase rapidly within 1–3 days after CNS injury at almost 100 times the basic proliferation rate. With increasing time after injury, NG2 cells gradually decrease and basically return to their original density within a few weeks (Simon et al. 2011). When brain injury occurred, the NG2 cells around the injury showed morphological changes including enlarged cell bodies and hypertrophic protuberances, and immunoreactivity was significantly enhanced (Tanaka et al. 2001). In a model of acupuncture injury of the cerebellar cortex, the number of NG2 cells increased at the site of injury, and the number of irregularly shaped NG2 cells increased at 48 h after injury (Levine 1994). Jones et al. found that NG2 cells increased significantly after spinal cord injury and reached a peak on the 7th day after injury (Jones et al. 2002). After spinal cord injury, NG2 cells proliferate and differentiate into oligodendrocytes, which directly and indirectly affect many aspects of spinal cord injury, including hemorrhage, angiogenesis, glial scar formation, laminin deposition, astrocyte reaction, and axonal growth (Hesp et al. 2018). In CNS injury, NG2 cells participate in myelin regeneration and axon protection, but NG2 cells also seem to participate in glial scar formation and hinder axon regeneration (Tran et al. 2018). Glial scars formed after CNS injury include a large number of NG2 cells (Tran et al. 2018). NG2 proteoglycan is a major obstacle to axonal growth (Gaudet and Fonken 2018). In vitro, when neurons were grown in an environment containing scattered NG2 proteoglycan, axon growth bypassed the area of NG2 proteoglycan distribution and extended to other regions (Dou and Levine 1994). Some studies related to axon regeneration have found that neutralizing NG2 proteoglycan with an anti-NG2 antibody after injury can promote sensory axon regeneration and functional recovery (Tan et al. 2006). However, other studies have found that NG2 proteoglycans have the opposite effect, suggesting that NG2 cells provide adhesion substrates for axonal growth (Yang et al. 2006). It has also been found that activation of the Notch signaling pathway in the microenvironment after CNS injury induces NG2 cells to differentiate into astrocytes, and astrocytes are the main cell types in glial scars (Huang et al. 2018; Khazaei et al. 2020). Reactive astrocytes lose their normal cellular function and have cytotoxic effects on local neurons and oligodendrocytes, and these effects are strongly associated with pathogenic progression (Xu et al. 2018). Inhibition of the Notch signaling pathway can inhibit the proliferation of reactive astrocytes and reduce inflammation-related secondary injury (Qian et al. 2019). In injured tissues, the immune response of NG2 cells often changes earlier than that of neurons and even earlier than that of other glial cells and macrophages, indicating that NG2 cells play an active role in neuropathology and may buffer the cytotoxins produced in the process of injury by releasing some biological factors and eliminating denatured tissue through inflammatory reactions (Wang and He 2009). However, the response of NG2 cells depends on the type of injury and the stage of development. Moreover, the significance of NG2 cell activation under injury conditions is uncertain. Therefore, in future research, it will be of great significance to explore the changes in and the role of NG2 cells in CNS diseases to fully utilize their protective functions and reduce their impairment, which will be of great significance for the treatment and recovery of CNS diseases.

Conclusions

Notch signaling may represent a novel therapeutic target in demyelination disorders and enhances the understanding of demyelination/remyelination. However, caution is needed because interfering with Notch signaling could have harmful effects.