Neural stem cells: developmental mechanisms and disease modeling

Cell Tissue Res. Author manuscript; available in PMC 2018 May 22.

Published in final edited form as:

PMCID: PMC5963504

NIHMSID: NIHMS967727

1Department of Neuroscience, University of Wisconsin-Madison, Madison, WI 53705, USA

2Waisman Center, University of Wisconsin-Madison, Madison, WI 53705, USA

1Department of Neuroscience, University of Wisconsin-Madison, Madison, WI 53705, USA

1Department of Neuroscience, University of Wisconsin-Madison, Madison, WI 53705, USA

2Waisman Center, University of Wisconsin-Madison, Madison, WI 53705, USA

The astonishing progress in the field of stem cell biology during the past 40 years has transformed both science and medicine. Neural stem cells (NSCs) are the stem cells of the nervous system. During development they give rise to the entire nervous system. In adults, a small number of NSCs remain and are mostly quiescent; however, ample evidence supports their important roles in plasticity, aging, disease, and regeneration of the nervous system. Because NSCs are regulated by both intrinsic genetic and epigenetic programs and extrinsic stimuli transduced through the stem cell niche, dysregulation of NSCs due to either genetic causes or environmental impacts may lead to disease. Therefore, extensive investigations in the past decades have been devoted to understanding how NSCs are regulated. On the other hand, ever since their discovery, NSCs have been a focal point for cell-based therapeutic strategies in the brain and spinal cord. The limited number of NSCs residing in the tissue has been a limiting factor for their clinical applications. Although recent advancements in embryonic and induced pluripotent stem cells have provided novel sources for NSCs, several challenges remain. In this special issue, leaders and experts in NSCs summarize our current understanding of NSC molecular regulation and the importance of NSCs for disease modeling and translational applications.

The term stem cells first appeared in the scientific literature in 1868 by the German biologist Ernst Haeckel (Haeckel, 1868). In his writings (Haeckel, 1868), stem cells had two distinct meanings: one is the unicellular evolutionary origin of all multicellular organisms, and the other is the fertilized egg giving rise to all other cell types of the body. The latter definition has evolved into the modern definition of stem cells - cells that can divide to self-renew and to differentiate into other cell types in tissues and organs (Li and Zhao, 2008, Ramalho-Santos and Willenbring, 2007).

The behavior and fate of stem cells are strongly influenced by their specific anatomical locations and surrounding cell types, called the stem cell niche. The niche provides physical support to host or anchor stem cells, and supplies factors to maintain and regulate them (Li and Zhao, 2008). Stem cells are also regulated by intrinsic signaling cascades and transcriptional mechanisms, some of which are common among all stem cells, and others that are unique to specific types. Some of the best known regulators include TGF-, BMP, Smad, Wnt, Notch, EGF fibroblast growth factors (Jobe, et al., 2012, Li and Zhao, 2008). Therefore, stem cells are regulated by complex mechanisms in both temporal- and context-specific manners to maintain their unique characteristics. Understanding stem cell regulation gives us the opportunity to explore mechanisms of development, as well as disorders resulting from their dysfunction.

During development, the central nervous system (CNS) is generated from a small number of neural stem cells (NSCs) lining the neural tube (Kriegstein and Alvarez-Buylla, 2009). A great deal of experimental evidence has demonstrated that radial glia, the NSCs during mammalian CNS development, undergo both symmetric divisions to expand the NSC pool, and asymmetric divisions to give rise to intermediate progenitors (IPCs) and the differentiated cell types. The three major cell types in the CNS arise from NSCs in a temporally defined sequence, with neurons appearing first, followed by astrocytes, and then oligodendrocytes (Okano and Temple, 2009). The technical advancement of live imaging and genomic tools have allowed for the identification of human-specific NSC populations (e.g. outer radial glia, or oRG) located at the outer subventricular zone (SVZ) (Gertz, et al., 2014). These oRG are essential for cortical expansion to achieve the large size of the human cortex. Single-cell genomic technologies have identified specific oRG markers that might be used for further characterization of these cells (Liu, et al., 2016, Pollen, et al., 2014). Investigating the regulatory mechanisms governing the self-renewal and fate specification of NSCs, especially human-specific developmental features, has significantly contributed to our understanding of human brain development and developmental diseases. In addition, this knowledge also has helped scientists refine protocols for pluripotent stem cell differentiation into specific nervous system cell types for both therapeutic goals and disease modeling.

In adult brains, NSCs are reduced and become restricted to specific brain regions. In rodents, both NSCs and ongoing neurogenesis have been widely documented in the SVZ of the lateral ventricles and the subgranular zone (SGZ) of the dentate gyrus (DG) of the hippocampus (Kempermann, et al., 2015). In humans, experimental evidence has supported ongoing neurogenesis in the hippocampus (Eriksson, et al., 1998, Spalding, et al., 2013). The confirmation of mammalian adult neurogenesis in the 1990s was one of the most exciting moments in science in the 21st century. Not only did it overthrow the prevailing dogma suggesting no neurons were made in the adult brain, but also it hinted that these adult NSCs could be utilized for neural repair in disease and following injury. Forty years later, we have learned a lot about NSCs. In the adult rodent SVZ, neurogenesis has been shown to be important for olfactory function and olfactory learning (Alonso, et al., 2006). During development, a subset of slowly-dividing NSCs are set aside to be the NSCs of the SVZ in the postnatal and adult brain (Fuentealba, et al., 2015, Furutachi, et al., 2015). The majority of neurogenic radial glia, however, become astrocytes and ependymal cells at the end of embryonic neurogenesis (Noctor, et al., 2004). A subset of these astrocytes persist as NSCs in specialized niches in the adult brain and continuously generate neurons that functionally integrate into restricted brain regions (Doetsch, 2003). In the hippocampus, radial glia-like stem cells of the SGZ make newborn neurons throughout life (Goritz and Frisen, 2012). These newborn neurons integrate into the circuity of the DG, contributing to behaviors such as pattern separation (Aimone, et al., 2011) and spatial learning (Dupret, et al., 2008), as well as hippocampus-associated learning, memory, and executive functions (Kempermann, Song and Gage, 2015).

Significant effort has been devoted into understanding the regulation of adult neurogenesis. As a result, we now know that many extrinsic stimuli and intrinsic mechanisms can affect this process. Mouse genetic studies have clearly demonstrated the important role of transcriptional regulation of NSCs through intrinsic genetic mechanisms (Hsieh and Zhao, 2016). Some examples include SOXC family proteins [Kavyanifar et al, in this issue (Kavyanifar, et al., 2018)], Bmi-1 (Molofsky, et al., 2003), Sox2 (Ferri, et al., 2004, Graham, et al., 1999), PTEN (Bonaguidi, et al., 2011), and Notch [Zhang et al, in this issue (Zhang, et al., 2018)]. In addition epigenetic regulation by DNA methylation pathways (e.g. Mbd1, Mecp2, Dnmt, Tet) (Noguchi, et al., 2015, Smrt, et al., 2007, Tsujimura, et al., 2009, Zhang, et al., 2013, Zhao, et al., 2003), chromatin remodeling (e.g. BAF, BRG1) (Ninkovic, et al., 2013, Petrik, et al., 2015, Tuoc, et al., 2017), and noncoding RNAs (Liu, et al., 2010)[Anderson and Lim, in this issue (Anderson and Lim, 2018)] play important roles. Many growth factors, signaling molecules, and neurotransmitters have been shown to regulate neurogenesis (Kempermann, Song and Gage, 2015). Catavero et al [in this issue (Catavero, et al., 2018)] review the role of GABA circuits, signaling, and receptors in regulating development of adult born cells, as well as the molecular players that modulate GABA signaling. Because progenitors with multipotent differentiation potentials have been found in brain regions without active neurogenesis (Palmer, et al., 1997), it is hypothesized that these progenitors might be manipulated to become neuron-competent in vivo so that they can contribute to brain generation [Wang et al, in this issue (Wang and Zhang, 2018)].

A great amount of literature has documented how physiological activities and enriched environment influences adult neurogenesis (Kempermann, Song and Gage, 2015). However, as summarized by Eisinger and Zhao [in this issue (Eisinger and Zhao, 2018)], the genes and gene network involved in these changes within NSCs have not been systematically analyzed at genome wide levels. Adult neurogenesis is also influenced by diseases including epilepsy (Parent and Lowenstein, 1997), stroke (Zhang and Chopp, 2016), depression (Dranovsky and Hen, 2006, Kempermann, et al., 2003), and injury [(Morshead and Ruddy, in this issue (Morshead and Ruddy, 2018) in this issue). Thodeson et al [in this issue (Thodeson, et al., 2018)] further summarize the contribution and dysregulation of adult neurogenesis in epilepsy and discuss how we can translate these findings to human therapeutics by using patient-derived neurons to study monogenic epilepsy-in-a-dish.

Aging affects every individual and is a major risk factor for many diseases. One of the strongest negative regulators of adult neurogenesis is aging. Both intrinsic and extrinsic components regulate the limitations of NSC proliferation and function (Moore and Jessberger, 2017, Seib and Martin-Villalba, 2015). In this issue, Mosher and Schaffer (Mosher and Schafer, 2018) and Ruddy and Morshead (Morshead and Ruddy, 2018) examine factors such as secreted signals, cell contact- dependent signals, and extracellular matrix cues that control neurogenesis in an age-dependent manner, and define these signals by the extrinsic mechanism through which they are presented to the NSCs. Smith et al [in this issue (Smith, et al., 2018)] discuss how age-related changes in the blood, such as blood-borne-factors, and peripheral immune cells, contribute to the age-related decline in adult neurogenesis in the mammalian brain.

Despite the extensive knowledge we have gained regarding adult neurogenesis, critical questions remain. For example, the control of the functional integration of new neurons remains a mystery. It has been shown that adult NSC-differentiated newborn neurons exhibit a critical period for sensitivity to external stimuli (Bergami, et al., 2015), and a heightened sensitivity to seizures (Kron, et al., 2010). It remains unclear how new neurons choose their connections. Jahn and Bergami [in this issue (Jahn and Bergami, 2018)] further discuss the critical period and its regulators during adult newborn neuron development.

Understanding the extrinsic and intrinsic regulation of adult NSCs and their newborn progeny, and their response to both positive and negative stimuli will further illuminate their role in disease, injury, stress, and brain function.

Human pluripotent stem cells (PSCs), including human embryonic stem cells (ESCs) and induced PSCs (iPSCs), offer a model system to reveal cellular and molecular events underlying normal and abnormal neural development in humans. ESCs are pluripotent cells derived from the inner cell mass of blastocyst stage preimplantation embryos, which were first isolated from mouse by Evans and Kaufman in 1981 (Evans and Kaufman, 1981) and later from humans by James Thompson in 1998 (Thomson, et al., 1998). Human ESCs are invaluable in the study of early embryonic development, allowing us to identify critical regulators of cell commitment, differentiation, and adult cell reprogramming (Dvash, et al., 2006, Ren, et al., 2009). iPSCs are reprogrammed from somatic cells by forced expression of stem cell genes and have the characteristics of ESCs (Okita, et al., 2007, Yu, et al., 2007). The development of iPSC technology has allowed us access to cells of the human nervous system through reprogramming of patient-derived cells, revolutionizing our ability to study human development and diseases.

To generate neural cells from either ESCs or iPSCs, the first step is neural induction. Through actions of a number of activators and inhibitors of cell signaling pathways, this process yields neural epithelial cell-like NSCs and then intermediate neural progenitors, resembling embryonic development. Despite many advances, a major hurdle of neural differentiation is lineage control. Using a standard dorsal forebrain neural differentiation protocol, most neural progenitors obtained are forebrain excitatory progenitors that produce mostly forebrain glutamatergic excitatory neurons. However, the purity and layer-specific composition of these progenitors, as well as neurons, vary significantly from experiment to experiment, cell line to cell line, and lab to lab. In addition, differentiation into specific types of neurons with high purity has always been a challenging goal. Much effort has been devoted into improving the efficiency of dopaminergic neuron and GABAergic neuron differentiation with great success (Hu, et al., 2010). However, the brain has many other types of neurons. Vadodaria et al [in this issue (Vadodaria, et al., 2018)] discuss how to generate serotonergic neurons, a type of neuron highly relevant to psychiatric disorders. To better understand the molecular control of human PSC and NSC differentiation, where protocols result in a large amount of cellular heterogeneity in identity and response, analysis must be done at the level of single cells. Harbom et al [in this issue (Harbom, et al., 2018)] summarizes how new state-of-the-art single-cell analysis methods may help to define differentiation from pluripotent cells.

The advancement in iPSC and gene editing technology has transformed the field of human disease modeling. As in many human disorders, especially neuropsychiatric disorders, mouse models have been useful. Yet there are several critical reasons why it is necessary to use human cells to define the underlying mechanisms that lead to human patient characteristics, particularly those affecting the nervous system. For example, in fragile X syndrome (FXS), the epigenetic silencing of the Fragile X Mental Retardation Gene 1 (FMR1) gene that causes FXS occurs only in humans. Mice engineered to mimic the human mutation in the FMR1 gene do not show the same methylation and silencing characteristics of the gene as in humans (Brouwer, et al., 2007). These results indicate that some epigenetic mechanisms in human and mice are different and preclude the ability to study epigenetic mechanisms of FMR1 silencing in mouse models of FXS (Bhattacharyya and Zhao, 2016). In this issue, Li and Shi discuss disease modeling using human PSC-differentiated neural progenitors (Li, et al., 2018), and Brito et al specifically focus on modeling autism spectrum disorder (Brito, et al., 2018).

The use of NSCs as a treatment strategy in CNS disease and injury has been tested for decades. Parkinsons disease specifically has gained the most momentum for potential therapeutic benefits (Studer, 2017); however, similar work has been performed in Huntingtons disease, stroke, and following spinal cord injury [for a review on this topic, see (Vishwakarma, et al., 2014)]. In some of these paradigms, NSCs are expected to differentiate into a specific cell type in the local CNS environment; in other cases, they are in a supportive role. In this issue, Kameda et al explores progress in using NSCs as a therapy following spinal cord injury (Kameda, et al., 2018).

While the development of PSC technologies has been a scientific breakthrough for future studies, there are limitations and risks that may be associated with their use. ESCs, iPSCs, and their differentiated NSCs are dividing cells. Either transplantation of NSCs or in vivo reprogramming of endogenous cells into NSCs could lead to tumorigenesis. In addition, reprogramming somatic cells into iPSCs results in a loss of some epigenetic signatures of disease and aging which are critical for disease modeling (Mertens, et al., 2015, Miller, et al., 2013, Ocampo, et al., 2016). In recent years, direct reprogramming of fibroblasts or other cell types into induced neurons (iN) has been developed (for review see (Mertens, et al., 2016)). Remarkably a growing number of studies have demonstrated that such direct reprogramming also can be effective in vivo. Wang et al [(Wang and Zhang, 2018) in this issue] will summarize recent progress of in vivo reprogramming into new neurons and present how this method can be used for spinal cord injury.

In cellular reprogramming, the cells targeted and the genetic factors used vary; however, the biggest difference is that some protocols push cells through a NSC stage, whereas others skip these stages (Gascon, et al., 2017, Guo, et al., 2014, Wang, et al., 2016). Bypassing this developmental stage has both pros and cons, and may lead to a completely novel path towards lineage commitment [discussed by Falk and Karow (Falk and Karow, 2018) in this issue].

NSCs are fascinating and promising cells because of their capability, flexibility, and multiplicity. Understanding how NSCs function provides important knowledge in development, adaptation, disease, regeneration, and rehabilitation of the nervous system. The studies of cortical development and adult neurogenesis using rodent models have contributed significantly to our knowledge about NSCs and will continually yield important new information, taking advantage of novel genetic and imaging technologies. However, using human NSCs provides us with a window to investigate human-specific aspects of development and disease mechanisms, which is potentiated by the fast advancement of stem cell and gene editing technologies. Challenges still remain regarding cell lineage control, in vivo NSC behavior, three dimensional cellular interactions, and preservation of epigenetic and aging marks.

We thank Klaus Unsicker for his encouragement and support and Jutta Jger for her help with invitations, and communications with authors and reviewers. This work was supported by grants from the US National Institutes of Health (R01MH078972, R56MH113146, R21NS098767, and R21NS095632 to X.Z, U54HD090256 to the Waisman Center), University of Wisconsin (UW)-Madison Vilas Trust (Kellett Mid-Career Award to X.Z.) and UW-Madison and Wisconsin Alumni Research Foundation (WARF to X.Z.), Jenni and Kyle Professorship (to X.Zhao), a Sloan Research Fellowship (to D.L.M.), a Junior Faculty Grant from the American Federation for Aging Research (to D.L.M.), and startup funds from UW-Madison School of Medicine and Public Health, WARF, and the Neuroscience Department (to D.L.M.).

Read more:
Neural stem cells: developmental mechanisms and disease modeling