Molecular Components of Store-Operated Calcium Channels in the Regulation of Neural Stem Cell Physiology, Neurogenesis, and the Pathology of Huntington's Disease

Abstract

One of the major Ca²⁺ signaling pathways is store-operated Ca²⁺ entry (SOCE), which is responsible for Ca²⁺ flow into cells in response to the depletion of endoplasmic reticulum Ca²⁺ stores. SOCE and its molecular components, including stromal interaction molecule proteins, Orai Ca²⁺ channels, and transient receptor potential canonical channels, are involved in the physiology of neural stem cells and play a role in their proliferation, differentiation, and neurogenesis. This suggests that Ca²⁺ signaling is an important player in brain development. Huntington's disease (HD) is an incurable neurodegenerative disorder that is caused by polyglutamine expansion in the huntingtin (HTT) protein, characterized by the loss of γ-aminobutyric acid (GABA)-ergic medium spiny neurons (MSNs) in the striatum. However, recent research has shown that HD is also a neurodevelopmental disorder and Ca²⁺ signaling is dysregulated in HD. The relationship between HD pathology and elevations of SOCE was demonstrated in different cellular and mouse models of HD and in induced pluripotent stem cell-based GABAergic MSNs from juvenile- and adult-onset HD patient fibroblasts. The present review discusses the role of SOCE in the physiology of neural stem cells and its dysregulation in HD pathology. It has been shown that elevated expression of STIM2 underlying the excessive Ca²⁺ entry through store-operated calcium channels in induced pluripotent stem cell-based MSNs from juvenile-onset HD. In the light of the latest findings regarding the role of Ca²⁺ signaling in HD pathology we also summarize recent progress in the in vitro differentiation of MSNs that derive from different cell sources. We discuss advances in the application of established protocols to obtain MSNs from fetal neural stem cells/progenitor cells, embryonic stem cells, induced pluripotent stem cells, and induced neural stem cells and the application of transdifferentiation. We also present recent progress in establishing HD brain organoids and their potential use for examining HD pathology and its treatment. Moreover, the significance of stem cell therapy to restore normal neural cell function, including Ca²⁺ signaling in the central nervous system in HD patients will be considered. The transplantation of MSNs or their precursors remains a promising treatment strategy for HD.

Keywords: Ca2+ homeostasis; Huntington’s disease; brain organoids; induced pluripotent stem cells; neural stem cells; store-operated Ca2+ channels; store-operated Ca2+ entry.

FIGURE 1

SOCE regulates neural stem cell physiology. Store-operated Ca²⁺ entry (SOCE) and its molecular components positively regulate the proliferation of embryonic stem cells (ESCs) and neural stem cell (NSCs)/neural progenitor cells (NPCs), differentiation of ECSs, and NSCs/NPCs as well as neurogenesis. References that correspond to respective discoveries are shown in the figure. The upward arrows indicate that SOCE and its molecular players positively regulate proliferation, differentiation, and neurogenesis. Compared with other studies Gopurappilly et al. (2018) (reference indicated in the figure by a down arrow) detected a decrease in NSC differentiation.

FIGURE 2

Neural stem cells in HD pathology and contribution of SOCE to HD MSN pathology. Under physiological conditions, neural stem cells (NSCs)/neural progenitor cells (NPCs) are characterized by proper cell organization and development. The dysfunction of NSCs/NPCs under conditions of HD pathology, including both juvenile- and adult-onset, is illustrated. Juvenile-onset HD, in which mutant HTT (mHTT) contains over 60 CAG repeats, is characterized by a decrease in the differentiation of NSCs/NPCs in different in vitro models, with the exception of data that were published by Zhang et al. (2019) who discovered premature neurogenesis and neuronal differentiation (the reference is indicated in the figure by an up arrow). Furthermore, NSCs/NPCs exhibited decreases in neurogenesis and disruption in cell organization in juvenile-onset HD. In adult-onset HD, in which mHTT carries equally or more than 39 CAG repeats but not more than 60 repeats, Barnat et al. (2020) reported that mHTT dysregulates neuronal precursors differentiation and directs neurogenesis toward the neuronal lineage. In contrast Conforti et al. (2018) observed an attenuation of NSC differentiation (the reference is indicated in the figure by a down arrow). Additionally, a decrease in cell proliferation, and abnormal cell organization in adult-onset HD NSCs/NPCs in vitro and iPSC-derived cortical organoids were reported. References that correspond to the discoveries in juvenile- and adult-onset HD are shown. Under physiological conditions in wildtypes, in GABAergic medium spiny neurons (MSNs), synaptic spine stability is maintained by the SOCE process. In HD MSNs, supranormal synaptic neuronal SOCE (nSOCE) can lead to spine loss and subsequently neurodegeneration. The references that are shown in the figure relate to published data that demonstrate abnormal SOCE in induced pluripotent stem cell (iPSC)-derived MSNs from juvenile- and adult-onset HD and MSNs from the YAC128 transgenic mouse model of HD. Wu et al. (2016) detected an increase in nSOCE in synaptic spines of YAC128 MSNs. The thick red arrow represents the elevation of Ca²⁺ influx via nSOCE. The up or down arrows indicate an increase, a decrease, or dysregulation in proliferation, differentiation, neurogenesis, or cell organization in juvenile- and adult-onset HD.

FIGURE 3

Generation of human striatal MSNs from fibroblasts and stem cells. To reprogram fibroblasts into induced pluripotent stem cells (iPSCs), four transcription factors are used to transduce cells from HD patients (Victor et al., 2018). Indicated in red are the most commonly used factors for neural induction of iPSCs or embryonic stem cells (ESCs) to obtain neural stem cells (NSCs)/neural progenitor cells (NPCs), and for differentiation of progenitors and mature GABA-ergic medium spiny neurons, pMSNs and MSNs, respectively. Marked in black are factors used for neural induction or differentiation only in one or two protocols, which are discussed in the manuscripts. For direct differentiation, microRNA-9/9*-124, and other factors that are indicated in blue were used (Victor et al., 2014). The duration of each of the differentiation steps differs according to the various protocols. Figure summarizes fourteen protocols describing the generation of human striatal MSNs from iPSCs (Zhang et al., 2010; Delli Carri et al., 2013; Arber et al., 2015; Nekrasov et al., 2016; Adil et al., 2018; Comella-Bolla et al., 2020; Grigor’eva et al., 2020; Vigont et al., 2021), fibroblasts (Victor et al., 2014), ESCs (Nicoleau et al., 2013; Arber et al., 2015; Wu M. et al., 2018), NSCs (El-Akabawy et al., 2011), and NPCs (Lin et al., 2015). Using bFGF and neural media for culturing, it is possible to differentiate iPSCs to induced neural stem cells (iNSCs) and then transplant them into the striatal region of the mouse brain where they differentiate into MSNs (not shown) (Al-Gharaibeh et al., 2017).

FIGURE 4

Generation of human iPSC-derived three-dimensional brain structures modeling HD. All illustrated HD organoids, including two region-specific brain organoids that consist of the three-dimensional (3D) human striatal spheroid (hStrS) and human cortical spheroid (hCS) that were used to create the assembloid by Miura et al. (2020), originated from induced pluripotent stem cells (iPSCs). Cortical organoids (Conforti et al., 2018; Zhang et al., 2019) and the chimeric organoids (Zhang et al., 2019) represent 3D cell models of Huntington’s disease. Motor nerve organoids extended axons as a result of their culture in the special chamber with microchannels (Kawada et al., 2017). The organoid formation time differs depending on the protocol. Neural differentiation inducers were used in the organoids formation process: retinoic acid (RA) (Kawada et al., 2017; Conforti et al., 2018; Zhang et al., 2019), retinoid X receptor γ (RXRG) agonist, SR-11237 (Miura et al., 2020), glycogen synthase kinase (GSK) inhibitor, CHIR-99021 (Zhang et al., 2019), and sonic hedgehog signaling agonist (SAG) (Kawada et al., 2017).