Stem Cell Neurodevelopmental Solutions for Restorative Treatments of the Human Trunk and Spine

Abstract

The ability to reliably repair spinal cord injuries (SCI) will be one of the greatest human achievements realized in regenerative medicine. Until recently, the cellular path to this goal has been challenging. However, as detailed developmental principles are revealed in mouse and human models, their application in the stem cell community brings trunk and spine embryology into efforts to advance human regenerative medicine. New models of posterior embryo development identify neuromesodermal progenitors (NMPs) as a major bifurcation point in generating the spinal cord and somites and is leading to production of cell types with the full range of axial identities critical for repair of trunk and spine disorders. This is coupled with organoid technologies including assembloids, circuitoids, and gastruloids. We describe a paradigm for applying developmental principles towards the goal of cell-based restorative therapies to enable reproducible and effective near-term clinical interventions.

Keywords: assembloids; cell therapy; differentiation; gastruloids; organoids; spine development; stem cells; trunk development.

Figure 1

Overview of vertebrate ontogeny of cell types in the head vs. trunk and spine. Anterior/rostral development from neuroectoderm vs. posterior/caudal development from neuromesodermal progenitor cells. Lineage ontogeny from distinct stem and progenitor pools is detailed. General cell sources (left column) and general cell types produced by their differentiation (middle column) along with resulting general tissues (right column) are listed. Differentiation potential varies along the anterior–posterior neuraxis such as in cell types of the neural crest that can otherwise behave similarly.NSPCs, neural stem/progenitor cells; NCCs, neural crest cells; Hox PG, Hox paralogous group; CSF, cerebrospinal fluid; BBB, blood-brain barrier; PA, pharyngeal arch.

Figure 2

Neuromesodermal progenitor (NMP) models of human trunk and spine development. (A) NMPs act upstream as multipotent building blocks to generate the trunk and spine. (B) Putative gene regulatory network (GRN) of NMP differentiation (Gouti-Briscoe model). RA (retinoic acid), NSPC (neural stem/progenitor cell). (C) Two proposed NMP developmental models. NMP model 1: NMPs in the caudolateral epiblast (CLE)/node streak border (NSB) act as caudal axial stem cells, that give rise to NPCs and the presomitic mesoderm (PSM) that undergoes somitogenesis in waves under a segmentation clock (Henrique et al., 2015). NMP model 2: NMPs form a barrier between distinct neural stem and mesoderm stem zones (Wood et al., 2020). Both models produce the spinal cord and paraxial mesoderm/somites from caudal stem cell pools. SOX2/Bra spatial heterogeneity in NMPs correlates with differential migratory velocity and PSM vs. spinal cord lineage commitment (Romanos et al., 2021). NCC, neural crest cell; aPSM/pPSM, anterior/posterior presomitic mesoderm; PNS, peripheral nervous system; CNS, central nervous system.

Figure 3

Developmental principles applied to stem cell differentiation. (A) Overview flow chart of human pluripotent stem cell (hPSC) differentiation through NMPs. Key differentiation factors and biomarkers are provided. It should be noted that induced protocols with forced transcription factor expression are also used. Variations in differentiation factors for each lineage have been performed in the literature. (B) Two routes to caudal neural crest cells (NCCs). Route 1 is to caudalize anterior neural crest progenitors (Hox negative) using retinoic acid (RA) to produce posterior cranial, vagal, and cardiac NCCs (Hox1–5). Route 2 is to first generate an NMP intermediate that can in turn produce trunk NCCs with broad axial identity. (C) Hox code of spinal cord motor columns along the neuraxis (Philippidou and Dasen, 2013). R (rostral), C (caudal) directions. (D) Recapitulating the in vivo somite segmentation clock in vitro (Diaz-Cuadros et al., ; Matsuda et al., 2020). (E) SMN co-culture with skeletal myotubes to model NMJ function and dysfunction. (F) Neural circuit signal propagation efficiency depends on multicellular interactions between neurons (blue/pink), oligodendrocytes (green), and astrocytes (purple). (G) Tripartite synapse model wherein astrocytes (purple) assist synapse formation, function, and homeostasis.

Figure 4

3D heterogenous cell culture developmental models and biotechnologies. Overview of stem cell integration with biotechnologies to produce developmental neurotechnologies. In vitro 3D heterogenous cell culture systems include gastruloids, organoids, and assembloids that can be merged with biomaterials, microfluidics, and devices. mESC, mouse embryonic stem cell; hiPSC, human induced pluripotent stem cell; hESC, human embryonic stem cell; NMJ, neuromuscular junction.

Figure 5

Clinical potential of stem cells and developmental neurotechnologies. (A) Conditions of the human trunk and spine that will benefit from developmental neurotechnologies. Examples include trauma and spinal cord injury (SCI), neuromuscular disorders such as amyotrophic lateral sclerosis (ALS) and myasthenia gravis, neural tube closure defects such as spina bifida that relate to PAX3, neurocristopathies such as Hirschsprung’s disease that result from NCC migration defects, CNS neurodegeneration, demyelinating disorders such as multiple sclerosis, and sensory disorders such as neuropathic pain. (B) Clinical stages of developmental neurotechnology discovery and testing from pre-clinical through to current good manufacturing processes (cGMP). (C) The developmental neurotechnology paradigm to generate clinical therapies and disease models by merging stem cell developmental principles with neurotechnologies.